http://journals.iucr.org/f/journalhomepage.html Acta Crystallographica Section F Structural Biology and Crystallization Communications aims to provide a home for communications on the crystallization and structure determination of biological macromolecules. It will commence publication in January 2005 and will include three categories of publication: Structural genomics communications, Protein structure communications, Crystallization communications. Articles will be available online when ready, making publication as fast as possible, and will include unlimited free colour illustrations, movies and other enhancements. The editorial process will be completely electronic with respect to deposition, submission, refereeing and publication. en-gb Copyright (c) 2009 International Union of Crystallography International Union of Crystallography International Union of Crystallography http://journals.iucr.org urn:issn:1744-3091 Acta Crystallographica Section F Structural Biology and Crystallization Communications aims to provide a home for communications on the crystallization and structure determination of biological macromolecules. It will commence publication in January 2005 and will include three categories of publication: Structural genomics communications, Protein structure communications, Crystallization communications. Articles will be available online when ready, making publication as fast as possible, and will include unlimited free colour illustrations, movies and other enhancements. The editorial process will be completely electronic with respect to deposition, submission, refereeing and publication. text/html Open access article in Acta Crystallographica Section F Structural Biology and Crystallization Communications text monthly 1 2002-01-01T00:00+00:00 med@iucr.org Acta Crystallographica Section F Structural Biology and Crystallization Communications Copyright (c) 2009 International Union of Crystallography urn:issn:1744-3091 http://journals.iucr.org/logos/rss10f.gif http://journals.iucr.org/f/journalhomepage.html Still image http://scripts.iucr.org/cgi-bin/paper?ll5193 ADAMTS13 is a reprolysin-type metalloproteinase belonging to the ADAMTS (a disintegrin and metalloproteinase with thrombospondin type 1 motif) family. It specifically cleaves plasma von Willebrand factor (VWF) and regulates platelet adhesion and aggregation. ADAMTS13 is a multi-domain enzyme. In addition to the N-terminal metalloproteinase domain, the ancillary domains, including a disintegrin-like domain, a thrombospondin-1 type 1 repeat, a Cys-rich domain and a spacer domain, are required for VWF recognition and cleavage. In the present study, a fragment of the ADAMTS13 ancillary domains (ADAMTS13-DTCS; residues 287–685) was expressed using CHO Lec cells, purified and crystallized. Diffraction data sets were collected using the SPring-8 beamline. Two ADAMTS13-DTCS crystals with distinct unit-cell parameters generated data sets to 2.6 and 2.8 Å resolution, respectively. http://creativecommons.org/licenses/by/2.0/uk urn:issn:1744-3091 Akiyama, M. Takeda, S. Kokame, K. Takagi, J. Miyata, T. 2009-07-01 doi:10.1107/S1744309109023410 International Union of Crystallography A fragment of the ADAMTS13 ancillary domains (ADAMTS13-DTCS) has been expressed, purified and crystallized and the crystals have been characterized by X-ray diffraction. en VON WILLEBRAND FACTOR-CLEAVING PROTEINASE; ADAMTS13; METALLOPROTEINASES; ANCILLARY DOMAINS ADAMTS13 is a reprolysin-type metalloproteinase belonging to the ADAMTS (a disintegrin and metalloproteinase with thrombospondin type 1 motif) family. It specifically cleaves plasma von Willebrand factor (VWF) and regulates platelet adhesion and aggregation. ADAMTS13 is a multi-domain enzyme. In addition to the N-terminal metalloproteinase domain, the ancillary domains, including a disintegrin-like domain, a thrombospondin-1 type 1 repeat, a Cys-rich domain and a spacer domain, are required for VWF recognition and cleavage. In the present study, a fragment of the ADAMTS13 ancillary domains (ADAMTS13-DTCS; residues 287–685) was expressed using CHO Lec cells, purified and crystallized. Diffraction data sets were collected using the SPring-8 beamline. Two ADAMTS13-DTCS crystals with distinct unit-cell parameters generated data sets to 2.6 and 2.8 Å resolution, respectively. text/html Production, crystallization and preliminary crystallographic analysis of an exosite-containing fragment of human von Willebrand factor-cleaving proteinase ADAMTS13 text 7 65 2009-07-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications 1744-3091 crystallization communications 739 med@iucr.org 742 1744-3091 http://scripts.iucr.org/cgi-bin/paper?sw5030 Two crystal structures of recombinant Geobacillus stearothermophilus 6-phosphogluconate dehydrogenase (Gs6PDH) in complex with the substrate 6-­phosphogluconate have been determined at medium resolution. Gs6PDH shares significant sequence identity and structural similarity with the enzymes from Lactococcus lactis, sheep liver and the protozoan parasite Trypanosoma brucei, for which a range of structures have previously been reported. Comparisons indicate that amino-acid sequence conservation is more pronounced in the two domains that contribute to the architecture of the active site, namely the N-terminal and C-terminal domains, compared with the central domain, which is primarily involved in the subunit–subunit associations required to form a stable dimer. The active-site residues are highly conserved, as are the interactions with the 6-phosphogluconate. There is interest in 6PDH as a potential drug target for the protozoan parasite T. brucei, the pathogen responsible for African sleeping sickness. The recombinant T. brucei enzyme has proven to be recalcitrant to enzyme–ligand studies and a surrogate protein might offer new opportunities to investigate and characterize 6PDH inhibitors. The high degree of structural similarity, efficient level of expression and straightforward crystallization conditions mean that Gs6PDH may prove to be an appropriate model system for structure-based inhibitor design targeting the enzyme from Trypanosoma species. http://creativecommons.org/licenses/by/2.0/uk urn:issn:1744-3091 Cameron, S. Martini, V.P. Iulek, J. Hunter, W.N. 2009-05-01 doi:10.1107/S1744309109012767 International Union of Crystallography The structure of 6-phosphogluconate dehydrogenase from a moderate thermophile, G. stearothermophilus, is presented and compared with those of orthologous enzymes. en PENTOSE PHOSPHATE PATHWAY; 6-PHOSPHOGLUCONATE DEHYDROGENASE; GEOBACILLUS STEAROTHERMOPHILUS Two crystal structures of recombinant Geobacillus stearothermophilus 6-phosphogluconate dehydrogenase (Gs6PDH) in complex with the substrate 6-­phosphogluconate have been determined at medium resolution. Gs6PDH shares significant sequence identity and structural similarity with the enzymes from Lactococcus lactis, sheep liver and the protozoan parasite Trypanosoma brucei, for which a range of structures have previously been reported. Comparisons indicate that amino-acid sequence conservation is more pronounced in the two domains that contribute to the architecture of the active site, namely the N-terminal and C-terminal domains, compared with the central domain, which is primarily involved in the subunit–subunit associations required to form a stable dimer. The active-site residues are highly conserved, as are the interactions with the 6-phosphogluconate. There is interest in 6PDH as a potential drug target for the protozoan parasite T. brucei, the pathogen responsible for African sleeping sickness. The recombinant T. brucei enzyme has proven to be recalcitrant to enzyme–ligand studies and a surrogate protein might offer new opportunities to investigate and characterize 6PDH inhibitors. The high degree of structural similarity, efficient level of expression and straightforward crystallization conditions mean that Gs6PDH may prove to be an appropriate model system for structure-based inhibitor design targeting the enzyme from Trypanosoma species. text/html Geobacillus stearothermophilus 6-phosphogluconate dehydrogenase complexed with 6-phosphogluconate text 5 65 2009-05-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications 1744-3091 structural communications 450 med@iucr.org 454 1744-3091 http://scripts.iucr.org/cgi-bin/paper?gx5140 GlmU is a bifunctional enzyme that catalyzes the final two steps in the biosynthesis of UDP-GlcNAc. Crystals of GlmU from Mycobacterium tuberculosis obtained using ammonium sulfate as a precipitant diffracted poorly (to 3.4 Å resolution) and displayed an unusually high solvent content (>80%) with sparse crystal packing that resulted in large solvent channels. With one molecule per asymmetric unit, the monomers from three neighbouring asymmetric units related by the crystal threefold formed a biological trimer. Although this is the first report of the structure of GlmU determined in a cubic crystal form, the trimeric arrangement here is similar to that observed for other GlmU structures determined in hexagonal (H3, H32, P6322) space groups. http://creativecommons.org/licenses/by/2.0/uk urn:issn:1744-3091 Verma, S.K. Jaiswal, M. Kumar, N. Parikh, A. Nandicoori, V.K. Prakash, B. 2009-05-01 doi:10.1107/S1744309109010252 International Union of Crystallography The structure of M. tuberculosis N-acetylglucosamine-1-phosphate uridyltransferase (GlmU) was determined by the molecular-replacement method to 3.4 Å resolution in space group I432 and was refined to a final Rwork and Rfree of 0.285 and 0.321, respectively. en MYCOBACTERIUM TUBERCULOSIS; BIFUNCTIONAL ENZYMES; ACETYLTRANSFERASES; URIDYLTRANSFERASES; GLMU GlmU is a bifunctional enzyme that catalyzes the final two steps in the biosynthesis of UDP-GlcNAc. Crystals of GlmU from Mycobacterium tuberculosis obtained using ammonium sulfate as a precipitant diffracted poorly (to 3.4 Å resolution) and displayed an unusually high solvent content (>80%) with sparse crystal packing that resulted in large solvent channels. With one molecule per asymmetric unit, the monomers from three neighbouring asymmetric units related by the crystal threefold formed a biological trimer. Although this is the first report of the structure of GlmU determined in a cubic crystal form, the trimeric arrangement here is similar to that observed for other GlmU structures determined in hexagonal (H3, H32, P6322) space groups. text/html Structure of N-acetylglucosamine-1-phosphate uridyltransferase (GlmU) from Mycobacterium tuberculosis in a cubic space group text 5 65 2009-05-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications 1744-3091 structural communications 435 med@iucr.org 439 1744-3091 http://scripts.iucr.org/cgi-bin/paper?sw5028 The structure of the MarR-family transcription factor NMB1585 from Neisseria meningitidis has been solved using data extending to a resolution of 2.1 Å. Overall, the dimeric structure resembles those of other MarR proteins, with each subunit comprising a winged helix–turn–helix (wHtH) domain connected to an α-helical dimerization domain. The spacing of the recognition helices of the wHtH domain indicates that NMB1585 is pre-configured for DNA binding, with a putative inducer pocket that is largely occluded by the side chains of two aromatic residues (Tyr29 and Trp53). NMB1585 was shown to bind to its own promoter region in a gel-shift assay, indicating that the protein acts as an auto-repressor. http://creativecommons.org/licenses/by/2.0/uk urn:issn:1744-3091 Nichols, C.E. Sainsbury, S. Ren, J. Walter, T.S. Verma, A. Stammers, D.K. Saunders, N.J. Owens, R.J. 2009-03-01 doi:10.1107/S174430910900414X International Union of Crystallography The structure of the MarR-family regulator NMB1585 from N. meningitidis has been solved using data extending to 2.1 Å resolution. en MARR; NEISSERIA MENINGITIDIS; TRANSCRIPTION FACTORS The structure of the MarR-family transcription factor NMB1585 from Neisseria meningitidis has been solved using data extending to a resolution of 2.1 Å. Overall, the dimeric structure resembles those of other MarR proteins, with each subunit comprising a winged helix–turn–helix (wHtH) domain connected to an α-helical dimerization domain. The spacing of the recognition helices of the wHtH domain indicates that NMB1585 is pre-configured for DNA binding, with a putative inducer pocket that is largely occluded by the side chains of two aromatic residues (Tyr29 and Trp53). NMB1585 was shown to bind to its own promoter region in a gel-shift assay, indicating that the protein acts as an auto-repressor. text/html The structure of NMB1585, a MarR-family regulator from Neisseria meningitidis text 3 65 2009-03-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications 1744-3091 structural communications 204 med@iucr.org 209 1744-3091 http://scripts.iucr.org/cgi-bin/paper?en5349 Enterotoxigenic Escherichia coli (ETEC), a major global cause of diarrhea, initiates the pathogenic process via fimbriae-mediated attachment to the small intestinal epithelium. A common prototypic ETEC fimbria, colo­nization factor antigen I (CFA/I), consists of a tip-localized minor adhesive subunit CfaE and the stalk-forming major subunit CfaB, both of which are necessary for fimbrial assembly. To elucidate the structure of CFA/I at atomic resolution, three recombinant proteins were generated consisting of fusions of the minor and major subunits (CfaEB) and of two (CfaBB) and three (CfaBBB) repeats of the major subunit. Crystals of CfaEB diffracted X-rays to 2.1 Å resolution and displayed the symmetry of space group P21. CfaBB exhibited a crystal diffraction limit of 2.3 Å resolution and had the symmetry of space group P21212. CfaBBB crystallized in the monoclinic space group C2 and diffracted X-­rays to 2.3 Å resolution. These structures were determined using the molecular-replacement method. http://creativecommons.org/licenses/by/2.0/uk urn:issn:1744-3091 Li, Y.-F. Poole, S. Rasulova, F. McVeigh, A.L. Savarino, S.J. Xia, D. 2009-03-01 doi:10.1107/S1744309109001584 International Union of Crystallography Three fusion proteins were generated in order to resolve the atomic structure of the CFA/I fimbriae of enterotoxigenic E. coli. CfaEB is a fusion of the minor and major CFA/I subunits, while CfaBB and CfaBBB are tandem fusions of two and three repeats, respectively, of the major subunit. Each protein was crystallized and the crystal structures of each of these fusions were determined successively by the molecular-replacement method using the CfaE crystal structure as an initial phasing model. en COLONIZATION FACTOR ANTIGEN I FIMBRIAE; CFAB SUBUNIT; ENTEROTOXIGENIC ESCHERICHIA COLI Enterotoxigenic Escherichia coli (ETEC), a major global cause of diarrhea, initiates the pathogenic process via fimbriae-mediated attachment to the small intestinal epithelium. A common prototypic ETEC fimbria, colo­nization factor antigen I (CFA/I), consists of a tip-localized minor adhesive subunit CfaE and the stalk-forming major subunit CfaB, both of which are necessary for fimbrial assembly. To elucidate the structure of CFA/I at atomic resolution, three recombinant proteins were generated consisting of fusions of the minor and major subunits (CfaEB) and of two (CfaBB) and three (CfaBBB) repeats of the major subunit. Crystals of CfaEB diffracted X-rays to 2.1 Å resolution and displayed the symmetry of space group P21. CfaBB exhibited a crystal diffraction limit of 2.3 Å resolution and had the symmetry of space group P21212. CfaBBB crystallized in the monoclinic space group C2 and diffracted X-­rays to 2.3 Å resolution. These structures were determined using the molecular-replacement method. text/html Crystallization and preliminary X-ray diffraction analyses of several forms of the CfaB major subunit of enterotoxigenic Escherichia coli CFA/I fimbriae text 3 65 2009-03-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications 1744-3091 crystallization communications 242 med@iucr.org 247 1744-3091 http://scripts.iucr.org/cgi-bin/paper?tb5009 The binding of (+)-camphor to cytochrome P450cam (P450cam) expels a cluster of waters at the active site, raising the redox potential of the haem to an extent that allows reduction by the electron-transfer system. This binding was reported to involve no significant structural changes in the protein. Here, two ferric P450cam structures partially complexed with (+)-camphor were determined by X-­ray crystallography at 1.30–1.35 Å resolution, revealing the structures of the substrate-free and substrate-bound forms. (+)-Camphor binding induces rotation of Thr101 to form a hydrogen bond that acts as a hydrogen donor to a peripheral haem propionate. This bonding contributes to the redox-potential change. http://creativecommons.org/licenses/by/2.0/uk urn:issn:1744-3091 Sakurai, K. Shimada, H. Hayashi, T. Tsukihara, T. 2009-02-01 doi:10.1107/S1744309108044114 International Union of Crystallography X-ray structures of ferric cytochrome P450cam partially complexed with the substrate (+)-camphor to two different extents were determined at 1.30–1.35 Å resolution, revealing the protein structures of the substrate-free and substrate-bound forms. en CYTOCHROME P450CAM; (+)-CAMPHOR; REDOX POTENTIAL The binding of (+)-camphor to cytochrome P450cam (P450cam) expels a cluster of waters at the active site, raising the redox potential of the haem to an extent that allows reduction by the electron-transfer system. This binding was reported to involve no significant structural changes in the protein. Here, two ferric P450cam structures partially complexed with (+)-camphor were determined by X-­ray crystallography at 1.30–1.35 Å resolution, revealing the structures of the substrate-free and substrate-bound forms. (+)-Camphor binding induces rotation of Thr101 to form a hydrogen bond that acts as a hydrogen donor to a peripheral haem propionate. This bonding contributes to the redox-potential change. text/html Substrate binding induces structural changes in cytochrome P450cam text 2 65 2009-02-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications 1744-3091 protein structure communications 80 med@iucr.org 83 1744-3091 http://scripts.iucr.org/cgi-bin/paper?ll5172 Recombinant human interferon α-2A (rhIFN-α-2A) has been crystallized in complex with the recombinantly produced Fab fragment of a therapeutic monoclonal antibody (MEDI545; IgG1/κ) which targets several human inter­feron α subtypes. This constitutes the first reported crystals of a human type I interferon bound to an antibody. The orthorhombic crystals belonged to either space group I222 or I212121, with unit-cell parameters a = 134.82, b = 153.26, c = 163.49 Å. The diffraction of the crystals extended to 3.0 Å resolution. The asymmetric unit contained two Fab–rhIFN-α-2A complexes. This corresponded to a crystal volume per protein weight (VM) of 3.02 Å3 Da−1 and a solvent content of 59.3%. The corresponding three-dimensional structure is expected to shed light on the mechanism of action of MEDI545 and the molecular basis of its specificity. http://creativecommons.org/licenses/by/2.0/uk urn:issn:1744-3091 Oganesyan, V. Damschroder, M.M. Cook, K.E. Wu, H. Dall'Acqua, W.F. 2009-01-01 doi:10.1107/S1744309108037925 International Union of Crystallography Crystals of the complex between the Fab fragment of a human anti-interferon α therapeutic antibody and human interferon α-2A have been obtained and diffracted to 3.0 Å resolution. en INTERFERON [ALPHA]-2A; ANTIBODIES; FAB FRAGMENTS Recombinant human interferon α-2A (rhIFN-α-2A) has been crystallized in complex with the recombinantly produced Fab fragment of a therapeutic monoclonal antibody (MEDI545; IgG1/κ) which targets several human inter­feron α subtypes. This constitutes the first reported crystals of a human type I interferon bound to an antibody. The orthorhombic crystals belonged to either space group I222 or I212121, with unit-cell parameters a = 134.82, b = 153.26, c = 163.49 Å. The diffraction of the crystals extended to 3.0 Å resolution. The asymmetric unit contained two Fab–rhIFN-α-2A complexes. This corresponded to a crystal volume per protein weight (VM) of 3.02 Å3 Da−1 and a solvent content of 59.3%. The corresponding three-dimensional structure is expected to shed light on the mechanism of action of MEDI545 and the molecular basis of its specificity. text/html Crystallization and preliminary X-ray diffraction analysis of the complex between a human anti-interferon antibody fragment and human interferon α-2A text 1 65 2009-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications 1744-3091 crystallization communications 14 med@iucr.org 16 1744-3091 http://scripts.iucr.org/cgi-bin/paper?kw5005 Caenorhabditis elegans expresses two manganese superoxide dismutase enzymes (MnSOD-2 and MnSOD-3) that are targeted to the mitochondrion. MnSOD-2 is constitutively expressed, while synthesis of MnSOD-3 is inducible. The structures of these two mononuclear metalloenzymes have been determined to 1.8 and 1.7 Å resolution, respectively. Pink crystals formed in space group P41212 for each, with unit-cell parameters a = b = 81.0, c = 137.4 Å for MnSOD-2 and a = b = 81.8, c = 136.0 Å for MnSOD-3. The final structure of MnSOD-3 was refined to R = 21.6% and Rfree = 26.2% at 293 K, and R = 18.9% and Rfree = 22.6% at 100 K, while that of MnSOD-2 was refined to R = 16.9% and Rfree = 20.1% at 100 K. The asymmetric unit cell is comprised of two subunits. The resulting structures are very similar to that of human MnSOD and form a tetramer corresponding to a dimer of dimers. The subunit interface between dimers is comprised of two four-helix bundles that stabilize the biologically significant homotetramer. http://creativecommons.org/licenses/by/2.0/uk urn:issn:1744-3091 Trinh, C.H. Hunter, T. Stewart, E.E. Phillips, S.E.V. Hunter, G.J. 2008-12-01 doi:10.1107/S1744309108037056 International Union of Crystallography Two manganese superoxide dismutase enzymes isolated from the eukaryote C. elegans have been characterized and their structures determined. The closely related structures reveal a striking similarity to manganese superoxide dismutase found in humans. en MANGANESE SUPEROXIDE DISMUTASES; CAENORHABDITIS ELEGANS Caenorhabditis elegans expresses two manganese superoxide dismutase enzymes (MnSOD-2 and MnSOD-3) that are targeted to the mitochondrion. MnSOD-2 is constitutively expressed, while synthesis of MnSOD-3 is inducible. The structures of these two mononuclear metalloenzymes have been determined to 1.8 and 1.7 Å resolution, respectively. Pink crystals formed in space group P41212 for each, with unit-cell parameters a = b = 81.0, c = 137.4 Å for MnSOD-2 and a = b = 81.8, c = 136.0 Å for MnSOD-3. The final structure of MnSOD-3 was refined to R = 21.6% and Rfree = 26.2% at 293 K, and R = 18.9% and Rfree = 22.6% at 100 K, while that of MnSOD-2 was refined to R = 16.9% and Rfree = 20.1% at 100 K. The asymmetric unit cell is comprised of two subunits. The resulting structures are very similar to that of human MnSOD and form a tetramer corresponding to a dimer of dimers. The subunit interface between dimers is comprised of two four-helix bundles that stabilize the biologically significant homotetramer. text/html Purification, crystallization and X-ray structures of the two manganese superoxide dismutases from Caenorhabditis elegans text 12 64 2008-12-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications 1744-3091 protein structure communications 1110 med@iucr.org 1114 1744-3091 http://scripts.iucr.org/cgi-bin/paper?hv5114 The mechanism of molecular oxygen entry into the buried active site of the copper amine oxidase family has been investigated in several family members using biochemical, structural and in silico methods. These studies have revealed a structurally conserved β-sandwich which acts as a hydrophobic reservoir from which molecular oxygen can take several species-specific preferred pathways to the active site. Escherichia coli copper amine oxidase (ECAO) possesses an extra N-terminal domain that lies close to one entrance to the β-sandwich. In order to investigate whether the presence of this domain alters molecular oxygen entry in this enzyme, xenon was used as a molecular oxygen binding-site probe. The resulting 2.5 Å resolution X-ray crystal structure reveals xenon bound in similar positions to those observed in xenon-derivative crystal structures of other family members, suggesting that the N-terminal domain does not affect oxygen entry and that the E. coli enzyme takes up oxygen in a similar manner to the rest of the copper amine oxidase family. http://creativecommons.org/licenses/by/2.0/uk urn:issn:1744-3091 Pirrat, P. Smith, M.A. Pearson, A.R. McPherson, M.J. Phillips, S.E.V. 2008-12-01 doi:10.1107/S1744309108036373 International Union of Crystallography The structure of a xenon derivative of E. coli copper amine oxidase confirms the pathway of oxygen entry to the buried active site proposed for this class of enzymes. en COPPER AMINE OXIDASE; XENON DERIVATIVES; OXYGEN ENTRY The mechanism of molecular oxygen entry into the buried active site of the copper amine oxidase family has been investigated in several family members using biochemical, structural and in silico methods. These studies have revealed a structurally conserved β-sandwich which acts as a hydrophobic reservoir from which molecular oxygen can take several species-specific preferred pathways to the active site. Escherichia coli copper amine oxidase (ECAO) possesses an extra N-terminal domain that lies close to one entrance to the β-sandwich. In order to investigate whether the presence of this domain alters molecular oxygen entry in this enzyme, xenon was used as a molecular oxygen binding-site probe. The resulting 2.5 Å resolution X-ray crystal structure reveals xenon bound in similar positions to those observed in xenon-derivative crystal structures of other family members, suggesting that the N-terminal domain does not affect oxygen entry and that the E. coli enzyme takes up oxygen in a similar manner to the rest of the copper amine oxidase family. text/html Structure of a xenon derivative of Escherichia coli copper amine oxidase: confirmation of the proposed oxygen-entry pathway text 12 64 2008-12-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications 1744-3091 protein structure communications 1105 med@iucr.org 1109 1744-3091 http://scripts.iucr.org/cgi-bin/paper?ll5164 Prostaglandin E2 is a major lipid mediator that regulates diverse biological processes. To elucidate how prostaglandin E2 is recognized specifically by its antibody, the Fab fragment of a monoclonal anti-prostaglandin E2 antibody was prepared and its complex with prostaglandin E2 was crystallized. The stable Fab–prostaglandin E2 complex was prepared by gel-filtration chromatography. Crystals were obtained by the microbatch method at 277 K using polyethylene glycol 4000 as a precipitant. A diffraction data set was collected to 2.2 Å resolution. The crystals belonged to space group P212121, with unit-cell parameters a = 70.3, b = 81.8, c = 82.2 Å. The asymmetric unit was suggested to contain one molecule of the Fab–prostaglandin E2 complex, with a corresponding crystal volume per protein weight of 2.75 Å3 Da−1. http://creativecommons.org/licenses/by/2.0/uk urn:issn:1744-3091 Kurahashi, Y. Sugahara, M. Ago, H. Aoyama, S. Takahashi, N. Takio, K. Katsukawa, M. Yamamoto, S. Miyano, M. 2008-11-01 doi:10.1107/S174430910803131X International Union of Crystallography The Fab fragment of a monoclonal anti-prostaglandin E2 antibody was prepared and its complex with prostaglandin E2 was crystallized. en PROSTAGLANDIN E2; FAB FRAGMENTS; ANTIBODIES Prostaglandin E2 is a major lipid mediator that regulates diverse biological processes. To elucidate how prostaglandin E2 is recognized specifically by its antibody, the Fab fragment of a monoclonal anti-prostaglandin E2 antibody was prepared and its complex with prostaglandin E2 was crystallized. The stable Fab–prostaglandin E2 complex was prepared by gel-filtration chromatography. Crystals were obtained by the microbatch method at 277 K using polyethylene glycol 4000 as a precipitant. A diffraction data set was collected to 2.2 Å resolution. The crystals belonged to space group P212121, with unit-cell parameters a = 70.3, b = 81.8, c = 82.2 Å. The asymmetric unit was suggested to contain one molecule of the Fab–prostaglandin E2 complex, with a corresponding crystal volume per protein weight of 2.75 Å3 Da−1. text/html Crystallization and preliminary diffraction studies of prostaglandin E2-specific monoclonal antibody Fab fragment in the ligand complex text 11 64 2008-11-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications 1744-3091 crystallization communications 1027 med@iucr.org 1030 1744-3091 http://scripts.iucr.org/cgi-bin/paper?hv5112 Streptococcus pneumoniae genomes encode three sialidases, NanA, NanB and NanC, which are key virulence factors that remove sialic acids from various glycoconjugates. The enzymes have potential as drug targets and also as vaccine candidates. The 115 kDa NanA is the largest of the three sialidases and is anchored to the bacterial membrane. Although recombinantly expressed full-length NanA was soluble, it failed to crystallize; therefore, a 56.5 kDa domain that retained full enzyme activity was subcloned. The purified enzyme was crystallized in 0.1 M MES pH 6.5, 30%(w/v) PEG 4000 using the sitting-drop vapour-diffusion method. Data were collected at 100 K to 2.5 Å resolution from a crystal grown in the presence of the inhibitor 2-deoxy-2,3-dehydro-N-acetyl neuraminic acid. The crystal belongs to space group P212121, with unit-cell parameters a = 49.2, b = 95.6, c = 226.6 Å. The structure was solved by molecular replacement and refined to final R and Rfree factors of 0.246 and 0.298, respectively. http://creativecommons.org/licenses/by/2.0/uk urn:issn:1744-3091 Xu, G. Li, X. Andrew, P.W. Taylor, G.L. 2008-09-01 doi:10.1107/S1744309108024044 International Union of Crystallography The structure of a catalytically active subdomain of the NanA sialidase from S. pneumoniae is reported to a resolution of 2.5 Å. The complex with the inhibitor Neu5Ac2en identifies the key catalytic residues and provides a platform for structure-based development of specific inhibitors. en NANA; SIALIDASES; STREPTOCOCCUS PNEUMONIAE Streptococcus pneumoniae genomes encode three sialidases, NanA, NanB and NanC, which are key virulence factors that remove sialic acids from various glycoconjugates. The enzymes have potential as drug targets and also as vaccine candidates. The 115 kDa NanA is the largest of the three sialidases and is anchored to the bacterial membrane. Although recombinantly expressed full-length NanA was soluble, it failed to crystallize; therefore, a 56.5 kDa domain that retained full enzyme activity was subcloned. The purified enzyme was crystallized in 0.1 M MES pH 6.5, 30%(w/v) PEG 4000 using the sitting-drop vapour-diffusion method. Data were collected at 100 K to 2.5 Å resolution from a crystal grown in the presence of the inhibitor 2-deoxy-2,3-dehydro-N-acetyl neuraminic acid. The crystal belongs to space group P212121, with unit-cell parameters a = 49.2, b = 95.6, c = 226.6 Å. The structure was solved by molecular replacement and refined to final R and Rfree factors of 0.246 and 0.298, respectively. text/html Structure of the catalytic domain of Streptococcus pneumoniae sialidase NanA text 9 64 2008-09-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications 1744-3091 protein structure communications 772 med@iucr.org 775 1744-3091 http://scripts.iucr.org/cgi-bin/paper?rp9017 A correction is made to the list of authors for Ito et al. [Acta Cryst. (2008). F64, 531–532]. Copyright (c) 2008 International Union of Crystallography urn:issn:1744-3091 Ito, L. Hidaka, Y. Okumura, M. Konishi, H. Adermann, K. Yamaguchi, H. 2008-08-01 doi:10.1107/S1744309108021477 International Union of Crystallography A corrigendum to the article by Ito et al. [Acta Cryst. (2008). F64, 531–532]. en PROUROGUANYLIN; PRECURSOR PROTEINS; PEPTIDE HORMONES; CORRIGENDUM A correction is made to the list of authors for Ito et al. [Acta Cryst. (2008). F64, 531–532]. text/html Crystallization and preliminary X-ray structural studies of human prouroguanylin. Corrigendum text 8 64 2008-08-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2008 International Union of Crystallography 1744-3091 addenda and errata 771 med@iucr.org 771 1744-3091 http://scripts.iucr.org/cgi-bin/paper?bw9226 A correction is made to the name one of the authors of Lafaye et al. [Acta Cryst. (2008). F64, 111–114]. Copyright (c) 2008 International Union of Crystallography urn:issn:1744-3091 Lafaye, C. Iwema, T. Ferrer, J.-L. Kroll, J.S. Griat, M. Serre, L. 2008-08-01 doi:10.1107/S1744309108020307 International Union of Crystallography A corrigendum to the paper by Lafaye et al. [Acta Cryst. (2008). F64, 111–114]. en NEISSERIA; VIRULENCE; OXIDOREDUCTASES; CORRIGENDUM A correction is made to the name one of the authors of Lafaye et al. [Acta Cryst. (2008). F64, 111–114]. text/html Preliminary crystallographic data of the three homologues of the thiol–disulfide oxidoreductase DsbA in Neisseria meningitidis. Corrigendum text 8 64 2008-08-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2008 International Union of Crystallography 1744-3091 addenda and errata 770 med@iucr.org 770 1744-3091 http://scripts.iucr.org/cgi-bin/paper?en5301 MotB is an essential component of the proton motive force-driven bacterial flagellar motor. It binds to the stress-bearing layer of peptidoglycan in the periplasm, anchoring the MotA/MotB stator unit to the cell wall. Proton flow through the channel formed by the transmembrane helices of MotA and MotB generates the turning force (torque) applied to the rotor. Crystals of recombinant Helicobacter pylori MotB have been obtained by the sitting-drop vapour-diffusion method using ammonium sulfate as a precipitant. These crystals belong to space group P41212 or its enantiomorph P43212, with unit-cell parameters a = 75.2, b = 75.2, c = 124.7 Å. The asymmetric unit appears to contain one subunit, corresponding to a packing density of 3.4 Å3 Da−1. The crystals diffract X-rays to at least 1.8 Å resolution on a synchrotron-radiation source. Copyright (c) 2008 International Union of Crystallography urn:issn:1744-3091 O'Neill, J. Roujeinikova, A. 2008-06-01 doi:10.1107/S1744309108012219 International Union of Crystallography The cloning, overexpression, purification, crystallization and preliminary X-ray diffraction analysis of H. pylori MotB, a peptidoglycan-binding component of the stator of the bacterial flagellar motor, are reported. en HELICOBACTER PYLORI; BACTERIAL FLAGELLAR MOTOR; PEPTIDOGLYCAN BINDING MotB is an essential component of the proton motive force-driven bacterial flagellar motor. It binds to the stress-bearing layer of peptidoglycan in the periplasm, anchoring the MotA/MotB stator unit to the cell wall. Proton flow through the channel formed by the transmembrane helices of MotA and MotB generates the turning force (torque) applied to the rotor. Crystals of recombinant Helicobacter pylori MotB have been obtained by the sitting-drop vapour-diffusion method using ammonium sulfate as a precipitant. These crystals belong to space group P41212 or its enantiomorph P43212, with unit-cell parameters a = 75.2, b = 75.2, c = 124.7 Å. The asymmetric unit appears to contain one subunit, corresponding to a packing density of 3.4 Å3 Da−1. The crystals diffract X-rays to at least 1.8 Å resolution on a synchrotron-radiation source. text/html Cloning, purification and crystallization of MotB, a stator component of the proton-driven bacterial flagellar motor text 6 64 2008-06-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2008 International Union of Crystallography 1744-3091 crystallization communications 561 med@iucr.org 563 1744-3091 http://scripts.iucr.org/cgi-bin/paper?hc5048 Hepatocyte nuclear factor 4α (HNF4α) is a member of the nuclear receptor superfamily that plays a central role in organ development and metabolic functions. Mutations on HNF4α cause maturity-onset diabetes of the young (MODY), a dominant monogenic cause of diabetes. In order to understand the molecular mechanism of promoter recognition and the molecular basis of disease-causing mutations, the recombinant HNF4α DNA-binding domain was prepared and used in a study of its binding properties and in crystallization with a 21-mer DNA fragment that contains the promoter element of another MODY gene, HNF1α. The HNF4α protein displays a cooperative and specific DNA-binding activity towards its target gene-recognition elements. Crystals of the complex diffract to 2.0 Å using a synchrotron-radiation source under cryogenic (100 K) conditions and belong to space group C2, with unit-cell parameters a = 121.63, b = 35.43, c = 70.99 Å, β = 119.36°. A molecular-replacement solution has been obtained and structure refinement is in progress. This structure and the binding studies will provide the groundwork for detailed functional and biochemical studies of the MODY mutants. Copyright (c) 2008 International Union of Crystallography urn:issn:1744-3091 Lu, P. Liu, J. Melikishvili, M. Fried, M.G. Chi, Y.-I. 2008-04-01 doi:10.1107/S1744309108007136 International Union of Crystallography Sample preparation, characterization, crystallization and preliminary X-ray analysis are reported for the HNF4α–DNA binary complex. en PROTEIN-DNA COMPLEX; GEL-SHIFT ASSAY; DIABETES; NUCLEAR RECEPTORS; ZINC-FINGER PROTEINS Hepatocyte nuclear factor 4α (HNF4α) is a member of the nuclear receptor superfamily that plays a central role in organ development and metabolic functions. Mutations on HNF4α cause maturity-onset diabetes of the young (MODY), a dominant monogenic cause of diabetes. In order to understand the molecular mechanism of promoter recognition and the molecular basis of disease-causing mutations, the recombinant HNF4α DNA-binding domain was prepared and used in a study of its binding properties and in crystallization with a 21-mer DNA fragment that contains the promoter element of another MODY gene, HNF1α. The HNF4α protein displays a cooperative and specific DNA-binding activity towards its target gene-recognition elements. Crystals of the complex diffract to 2.0 Å using a synchrotron-radiation source under cryogenic (100 K) conditions and belong to space group C2, with unit-cell parameters a = 121.63, b = 35.43, c = 70.99 Å, β = 119.36°. A molecular-replacement solution has been obtained and structure refinement is in progress. This structure and the binding studies will provide the groundwork for detailed functional and biochemical studies of the MODY mutants. text/html Crystallization of hepatocyte nuclear factor 4α (HNF4α) in complex with the HNF1α promoter element text 4 64 2008-04-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2008 International Union of Crystallography 1744-3091 crystallization communications 313 med@iucr.org 317 1744-3091 http://scripts.iucr.org/cgi-bin/paper?pu5213 The C-terminal domain of MotB (MotB-C) contains a putative peptidoglycan-binding motif and is believed to anchor the MotA/MotB stator unit of the bacterial flagellar motor to the cell wall. Crystals of Helicobacter pylori MotB-C (138 amino-acid residues) were obtained by the hanging-drop vapour-diffusion method using polyethylene glycol as a precipitant. These crystals belong to space group P21, with unit-cell parameters a = 50.8, b = 89.5, c = 66.3 Å, β = 112.5°. The crystals diffract X-rays to at least 1.6 Å resolution using a synchrotron-radiation source. Self-rotation function and Matthews coefficient calculations suggest that the asymmetric unit contains one tetramer with 222 point-group symmetry. The anomalous difference Patterson maps calculated for an ytterbium-derivative crystal using diffraction data at a wavelength of 1.38 Å showed significant peaks on the v = 1/2 Harker section, suggesting that ab initio phase information could be derived from the MAD data. Copyright (c) 2008 International Union of Crystallography urn:issn:1744-3091 Roujeinikova, A. 2008-04-01 doi:10.1107/S1744309108005277 International Union of Crystallography The cloning, overexpression, purification, crystallization and preliminary X-ray diffraction analysis of a putative peptidoglycan-binding domain of H. pylori MotB, a stator component of the bacterial flagellar motor, are reported. en HELICOBACTER PYLORI; BACTERIAL FLAGELLAR MOTOR; PEPTIDOGLYCAN BINDING The C-terminal domain of MotB (MotB-C) contains a putative peptidoglycan-binding motif and is believed to anchor the MotA/MotB stator unit of the bacterial flagellar motor to the cell wall. Crystals of Helicobacter pylori MotB-C (138 amino-acid residues) were obtained by the hanging-drop vapour-diffusion method using polyethylene glycol as a precipitant. These crystals belong to space group P21, with unit-cell parameters a = 50.8, b = 89.5, c = 66.3 Å, β = 112.5°. The crystals diffract X-rays to at least 1.6 Å resolution using a synchrotron-radiation source. Self-rotation function and Matthews coefficient calculations suggest that the asymmetric unit contains one tetramer with 222 point-group symmetry. The anomalous difference Patterson maps calculated for an ytterbium-derivative crystal using diffraction data at a wavelength of 1.38 Å showed significant peaks on the v = 1/2 Harker section, suggesting that ab initio phase information could be derived from the MAD data. text/html Cloning, purification and preliminary X-ray analysis of the C-terminal domain of Helicobacter pylori MotB text 4 64 2008-04-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2008 International Union of Crystallography 1744-3091 crystallization communications 277 med@iucr.org 280 1744-3091 http://scripts.iucr.org/cgi-bin/paper?fw5161 ADP-ribosylation is a reversible and covalent post-translational modification in which the attachment of ADP-ribose is catalyzed by ADP-ribosyltransferases and the removal of ADP-ribose is catalyzed by ADP-ribosylhydrolases. ADP-ribosylhydrolase 3 from mouse, consisting of 347 amino-acid residues, has been cloned, purified and crystallized. The three-dimensional structure has been resolved at a resolution of 1.8 Å. The structure constitutes a compact all-α-­helical protein with two Mg2+ ions located in the active-site crevice. A structural comparison of mouse ADP-ribosylhydrolase 3 with its human orthologue shows a high degree of structural similarity. Furthermore, four prokaryotic proteins deposited in the PDB could be identified as being structurally related. Copyright (c) 2008 International Union of Crystallography urn:issn:1744-3091 Mueller-Dieckmann, C. Kernstock, S. Mueller-Dieckmann, J. Weiss, M.S. Koch-Nolte, F. 2008-03-01 doi:10.1107/S1744309108001413 International Union of Crystallography The crystal structure of ADP-ribosylhydrolase 3 from M. musculus has been determined and refined to a resolution of 1.8 Å. A detailed comparison with the human orthologue at the protein-sequence level as well as of the three-dimensional architecture is presented. en ADP-RIBOSYLHYDROLASE 3 ADP-ribosylation is a reversible and covalent post-translational modification in which the attachment of ADP-ribose is catalyzed by ADP-ribosyltransferases and the removal of ADP-ribose is catalyzed by ADP-ribosylhydrolases. ADP-ribosylhydrolase 3 from mouse, consisting of 347 amino-acid residues, has been cloned, purified and crystallized. The three-dimensional structure has been resolved at a resolution of 1.8 Å. The structure constitutes a compact all-α-­helical protein with two Mg2+ ions located in the active-site crevice. A structural comparison of mouse ADP-ribosylhydrolase 3 with its human orthologue shows a high degree of structural similarity. Furthermore, four prokaryotic proteins deposited in the PDB could be identified as being structurally related. text/html Structure of mouse ADP-ribosylhydrolase 3 (mARH3) text 3 64 2008-03-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2008 International Union of Crystallography 1744-3091 protein structure communications 156 med@iucr.org 162 1744-3091 http://scripts.iucr.org/cgi-bin/paper?en5276 Tuberculosis is a major cause of death worldwide. Understanding of the pathogenicity of Mycobacterium tuberculosis has been advanced by gene analysis and has led to the identification of genes that are important for intracellular survival in macrophages. One of these genes encodes HsaD, a meta-cleavage product (MCP) hydrolase that catalyzes the hydrolytic cleavage of a carbon–carbon bond in cholesterol metabolism. This paper describes the production of HsaD as a recombinant protein and, following crystallization, the determination of its three-dimensional structure to 2.35 Å resolution by X-ray crystallography at the Diamond Light Source in Oxfordshire, England. To the authors' knowledge, this study constitutes the first report of a structure determined at the new synchrotron facility. The volume of the active-site cleft of the HsaD enzyme is more than double the corresponding active-site volumes of related MCP hydrolases involved in the catabolism of aromatic compounds, consistent with the specificity of HsaD for steroids such as cholesterol. Knowledge of the structure of the enzyme facilitates the design of inhibitors. Copyright (c) 2008 International Union of Crystallography urn:issn:1744-3091 Lack, N. Lowe, E.D. Liu, J. Eltis, L.D. Noble, M.E.M. Sim, E. Westwood, I.M. 2008-01-01 doi:10.1107/S1744309107065931 International Union of Crystallography The structure of HsaD, a carbon–carbon bond serine hydrolase involved in steroid catabolism that is critical for the survival of M. tuberculosis inside human macrophages, has been solved by X-ray crystallography. Data were collected at the Diamond Light Source in Oxfordshire, England: this paper describes one of the first structures determined at the new synchrotron. en HSAD; MCP HYDROLASES; C-C BOND HYDROLASES; CHOLESTEROL; TUBERCULOSIS; DIAMOND Tuberculosis is a major cause of death worldwide. Understanding of the pathogenicity of Mycobacterium tuberculosis has been advanced by gene analysis and has led to the identification of genes that are important for intracellular survival in macrophages. One of these genes encodes HsaD, a meta-cleavage product (MCP) hydrolase that catalyzes the hydrolytic cleavage of a carbon–carbon bond in cholesterol metabolism. This paper describes the production of HsaD as a recombinant protein and, following crystallization, the determination of its three-dimensional structure to 2.35 Å resolution by X-ray crystallography at the Diamond Light Source in Oxfordshire, England. To the authors' knowledge, this study constitutes the first report of a structure determined at the new synchrotron facility. The volume of the active-site cleft of the HsaD enzyme is more than double the corresponding active-site volumes of related MCP hydrolases involved in the catabolism of aromatic compounds, consistent with the specificity of HsaD for steroids such as cholesterol. Knowledge of the structure of the enzyme facilitates the design of inhibitors. text/html Structure of HsaD, a steroid-degrading hydrolase, from Mycobacterium tuberculosis text 1 64 2008-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2008 International Union of Crystallography 1744-3091 protein structure communications 2 med@iucr.org 7 1744-3091 http://scripts.iucr.org/cgi-bin/paper?hc9013 A correction is made to the Experimental methods section of the article by Kefala & Weiss [(2006), Acta Cryst. F62, 1116–1119]. Copyright (c) 2008 International Union of Crystallography urn:issn:1744-3091 Kefala, G. Weiss, M.S. 2008-01-01 doi:10.1107/S1744309107065566 International Union of Crystallography A correction is made to the article by Kefala & Weiss [(2006), Acta Cryst. F62, 1116–1119]. en DIHYDRODIPICOLINATE SYNTHASE; MYCOBACTERIUM TUBERCULOSIS; RV2753C A correction is made to the Experimental methods section of the article by Kefala & Weiss [(2006), Acta Cryst. F62, 1116–1119]. text/html Cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of DapA (Rv2753c) from Mycobacterium tuberculosis. Corrigendum text 1 64 2008-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2008 International Union of Crystallography 1744-3091 addenda and errata 62 med@iucr.org 62 1744-3091 http://scripts.iucr.org/cgi-bin/paper?pu9193 A correction is made to one of the affiliations of the authors and also to a table heading in Miyakawa et al. (2007), Acta Cryst. F63, 737–739. Copyright (c) 2007 International Union of Crystallography urn:issn:1744-3091 Miyakawa, T. Sawano, Y. Miyazono, K. Hatano, K. Tanokura, M. 2007-10-01 doi:10.1107/S1744309107044119 International Union of Crystallography An erratum to the paper by Miyakawa et al. [(2007), Acta Cryst. F63, 737–739]. en GINKBILOBIN-2; ANTIFUNGAL PROTEINS; GINKGO BILOBA; ERRATUM A correction is made to one of the affiliations of the authors and also to a table heading in Miyakawa et al. (2007), Acta Cryst. F63, 737–739. text/html Crystallization and preliminary X-ray analysis of ginkbilobin-2 from Ginkgo biloba seeds: a novel antifungal protein with homology to the extracellular domain of plant cysteine-rich receptor-like kinases. Erratum text 10 63 2007-10-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2007 International Union of Crystallography 1744-3091 addenda and errata 899 med@iucr.org 899 1744-3091 http://scripts.iucr.org/cgi-bin/paper?gj5024 Sialyltransferases transfer sialic acid from cytidine-5-monophospho-N-acetylneuraminic acid (CMP-NeuAc) to the nonreducing termini of the oligosaccharyl structures of various glycoproteins and glycolipids. The newly cloned α2,6-sialyltransferase from Photobacterium sp. JT-ISH-224 (from the Vibrionaceae family) is composed of two domains: an unknown N-terminal domain and a catalytic C-terminal domain which shares significant homology with the Pasteurella multocida multifunctional sialyltransferase. The putative mature form of JT-ISH-224 α2,6-sialyltransferase was overproduced in Escherichia coli, purified and crystallized using the hanging-drop vapour-diffusion method at 293 K. The crystal belonged to space group P3121 or P3221, with unit-cell parameters a = b = 90.29, c = 204.33 Å. X-ray diffraction data were collected to 2.5 Å resolution. Copyright (c) 2007 International Union of Crystallography urn:issn:1744-3091 Okino, N. Kakuta, Y. Kajiwara, H. Ichikawa, M. Takakura, Y. Ito, M. Yamamoto, T. 2007-08-01 doi:10.1107/S1744309107031363 International Union of Crystallography Crystallization of the α2,6-sialyltransferase from Photobacterium. en [ALPHA]2,6-SIALYLTRANSFERASE; PHOTOBACTERIUM SP. JT-ISH-224 Sialyltransferases transfer sialic acid from cytidine-5-monophospho-N-acetylneuraminic acid (CMP-NeuAc) to the nonreducing termini of the oligosaccharyl structures of various glycoproteins and glycolipids. The newly cloned α2,6-sialyltransferase from Photobacterium sp. JT-ISH-224 (from the Vibrionaceae family) is composed of two domains: an unknown N-terminal domain and a catalytic C-terminal domain which shares significant homology with the Pasteurella multocida multifunctional sialyltransferase. The putative mature form of JT-ISH-224 α2,6-sialyltransferase was overproduced in Escherichia coli, purified and crystallized using the hanging-drop vapour-diffusion method at 293 K. The crystal belonged to space group P3121 or P3221, with unit-cell parameters a = b = 90.29, c = 204.33 Å. X-ray diffraction data were collected to 2.5 Å resolution. text/html Purification, crystallization and preliminary crystallographic characterization of the α2,6-sialyltransferase from Photobacterium sp. JT-ISH-224 text 8 63 2007-08-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2007 International Union of Crystallography 1744-3091 crystallization communications 662 med@iucr.org 664 1744-3091 http://scripts.iucr.org/cgi-bin/paper?pu5183 The deoxyuridine triphosphate nucleotidohydrolase gene from Arabidopsis thaliana was expressed and the gene product was purified. Crystallization was performed by the hanging-drop vapour-diffusion method at 298 K using 2 M ammonium sulfate as the precipitant. X-ray diffraction data were collected to 2.2 Å resolution using Cu Kα radiation. The crystal belongs to the orthorhombic space group P212121, with unit-cell parameters a = 69.90, b = 70.86 Å, c = 75.55 Å. Assuming the presence of a trimer in the asymmetric unit, the solvent content was 30%, with a VM of 1.8 Å3 Da−1. Copyright (c) 2007 International Union of Crystallography urn:issn:1744-3091 Bajaj, M. Moriyama, H. 2007-05-01 doi:10.1107/S1744309107016004 International Union of Crystallography The first crystallization of deoxyuridine triphosphate nucleotidohydrolase from plant, Arabidopsis thaliana, has been performed. An additive, taurine, was effective in producing the single crystal. en DEOXYURIDINE TRIPHOSPHATE NUCLEOTIDOHYDROLASE; ARABIDOPSIS THALIANA The deoxyuridine triphosphate nucleotidohydrolase gene from Arabidopsis thaliana was expressed and the gene product was purified. Crystallization was performed by the hanging-drop vapour-diffusion method at 298 K using 2 M ammonium sulfate as the precipitant. X-ray diffraction data were collected to 2.2 Å resolution using Cu Kα radiation. The crystal belongs to the orthorhombic space group P212121, with unit-cell parameters a = 69.90, b = 70.86 Å, c = 75.55 Å. Assuming the presence of a trimer in the asymmetric unit, the solvent content was 30%, with a VM of 1.8 Å3 Da−1. text/html Purification, crystallization and preliminary crystallographic analysis of deoxyuridine triphosphate nucleotidohydrolase from Arabidopsis thaliana text 5 63 2007-05-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2007 International Union of Crystallography 1744-3091 crystallization communications 409 med@iucr.org 411 1744-3091 http://scripts.iucr.org/cgi-bin/paper?en5228 l-2-Keto-3-deoxyarabonate (l-KDA) dehydratase is a novel member of the dihydrodipicolinate synthase (DHDPS)/N-acetylneuraminate lyase (NAL) protein family and catalyzes the hydration of l-KDA to α-ketoglutaric semialdehyde. l-KDA dehydratase was overexpressed, purified and crystallized at 291 K using the hanging-drop vapour-diffusion method. The crystal diffracts to 2.0 Å resolution using synchrotron radiation and belongs to the trigonal space group P3121 or its enantiomorph P3221, with unit-cell parameters a = b = 78.91, c = 207.71 Å. Copyright (c) 2007 International Union of Crystallography urn:issn:1744-3091 Shimada, N. Mikami, B. Watanabe, S. Makino, K. 2007-05-01 doi:10.1107/S1744309107015102 International Union of Crystallography l-2-Keto-3-deoxyarabonate dehydratase was overexpressed, purified and crystallized at 291 K using the hanging-drop vapour-diffusion method. en L-2-KETO-3-DEOXYARABONATE DEHYDRATASE; DIHYDRODIPICOLINATE SYNTHASE/N-ACETYLNEURAMINATE LYASE PROTEIN FAMILY l-2-Keto-3-deoxyarabonate (l-KDA) dehydratase is a novel member of the dihydrodipicolinate synthase (DHDPS)/N-acetylneuraminate lyase (NAL) protein family and catalyzes the hydration of l-KDA to α-ketoglutaric semialdehyde. l-KDA dehydratase was overexpressed, purified and crystallized at 291 K using the hanging-drop vapour-diffusion method. The crystal diffracts to 2.0 Å resolution using synchrotron radiation and belongs to the trigonal space group P3121 or its enantiomorph P3221, with unit-cell parameters a = b = 78.91, c = 207.71 Å. text/html Preliminary crystallographic analysis of l-2-keto-3-deoxyarabonate dehydratase, an enzyme involved in an alternative bacterial pathway of l-­arabinose metabolism text 5 63 2007-05-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2007 International Union of Crystallography 1744-3091 crystallization communications 393 med@iucr.org 395 1744-3091 http://scripts.iucr.org/cgi-bin/paper?hc5021 1,3-Propanediol dehydrogenase (1,3-PD-DH), encoded by the dhaT gene, is a key enzyme in the dissimilation process for converting glycerol to 1,3-propanediol in the human pathogen Klebsiella pneumoniae. Single colourless crystals were obtained from a recombinant preparation of 1,3-propanediol dehydrogenase overexpressed in Escherichia coli. The crystals belong to space group P21, with unit-cell parameters a = 91.9, b = 226.6, c = 232.6 Å, β = 92.9°. The crystals probably contain two decamers in the asymmetric unit, with a VM value of 3.07 Å3 Da−1 and an estimated solvent content of 59%. Diffraction data were collected to 2.7 Å resolution using synchrotron radiation at the ID14-4 beamline of the European Synchrotron Radiation Facility. Copyright (c) 2007 International Union of Crystallography urn:issn:1744-3091 Marçal, D. Rego, A.T. Fogg, M.J. Wilson, K.S. Carrondo, M.A. Enguita, F.J. 2007-03-01 doi:10.1107/S1744309107008834 International Union of Crystallography 1,3-Propanediol dehydrogenase from K. pneumoniae has been overexpressed in E. coli, purified and crystallized. Diffraction data have been collected to 2.7 Å resolution. en 1,3-PROPANEDIOL DEHYDROGENASE; KLEBSIELLA PNEUMONIAE; GLYCEROL METABOLISM; KES; OPPORTUNISTIC PATHOGENS 1,3-Propanediol dehydrogenase (1,3-PD-DH), encoded by the dhaT gene, is a key enzyme in the dissimilation process for converting glycerol to 1,3-propanediol in the human pathogen Klebsiella pneumoniae. Single colourless crystals were obtained from a recombinant preparation of 1,3-propanediol dehydrogenase overexpressed in Escherichia coli. The crystals belong to space group P21, with unit-cell parameters a = 91.9, b = 226.6, c = 232.6 Å, β = 92.9°. The crystals probably contain two decamers in the asymmetric unit, with a VM value of 3.07 Å3 Da−1 and an estimated solvent content of 59%. Diffraction data were collected to 2.7 Å resolution using synchrotron radiation at the ID14-4 beamline of the European Synchrotron Radiation Facility. text/html Crystallization and preliminary X-ray characterization of 1,3-propanediol dehydrogenase from the human pathogen Klebsiella pneumoniae text 3 63 2007-03-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2007 International Union of Crystallography 1744-3091 crystallization communications 249 med@iucr.org 251 1744-3091 http://scripts.iucr.org/cgi-bin/paper?pu5181 Crystals of recombinant NovP (subunit MW = 29 967 Da; 262 amino acids), an S-adenosyl-l-methionine-dependent O-methyltransferase from Streptomyces spheroides, were grown by vapour diffusion. The protein crystallized in space group P2, with unit-cell parameters a = 51.81, b = 46.04, c = 61.22 Å, β = 104.97°. Native data to a maximum resolution of 1.4 Å were collected from a single crystal at the synchrotron. NovP is involved in the biosynthesis of the aminocoumarin antibiotic novobiocin that targets the essential bacterial enzyme DNA gyrase. Copyright (c) 2007 International Union of Crystallography urn:issn:1744-3091 Stevenson, C.E.M. Freel Meyers, C.L. Walsh, C.T. Lawson, D.M. 2007-03-01 doi:10.1107/S1744309107008287 International Union of Crystallography Monoclinic crystals of NovP, an O-methyltransferase from S. spheroides, were obtained and native X-ray data to 1.4 Å resolution were recorded. en NOVP; O-METHYLTRANSFERASE; STREPTOMYCES; NOVOBIOCIN; ANTIBIOTIC BIOSYNTHESIS Crystals of recombinant NovP (subunit MW = 29 967 Da; 262 amino acids), an S-adenosyl-l-methionine-dependent O-methyltransferase from Streptomyces spheroides, were grown by vapour diffusion. The protein crystallized in space group P2, with unit-cell parameters a = 51.81, b = 46.04, c = 61.22 Å, β = 104.97°. Native data to a maximum resolution of 1.4 Å were collected from a single crystal at the synchrotron. NovP is involved in the biosynthesis of the aminocoumarin antibiotic novobiocin that targets the essential bacterial enzyme DNA gyrase. text/html Crystallization and preliminary X-ray analysis of the O-methyltransferase NovP from the novobiocin-biosynthetic cluster of Streptomyces spheroides text 3 63 2007-03-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2007 International Union of Crystallography 1744-3091 crystallization communications 236 med@iucr.org 238 1744-3091 http://scripts.iucr.org/cgi-bin/paper?bw5192 Crystals of recombinant AbsC (subunit MW = 18 313 Da; 158 amino acids), a novel regulator of antibiotic production from Streptomyces coelicolor, were grown by vapour diffusion. The protein crystallizes in space group P212121, with unit-cell parameters a = 43.53, b = 121.30, c = 143.75 Å. Native data to a resolution of 2.25 Å were recorded at station PX 14.1 (Daresbury) from a single crystal. Preliminary analysis of these data suggests that the asymmetric unit contains four copies of the AbsC monomer, giving an estimated solvent content of 47.0%. AbsC belongs to the MarR family of proteins that mediate ligand-responsive transcriptional control. Copyright (c) 2007 International Union of Crystallography urn:issn:1744-3091 Stevenson, C.E.M. Kock, H. Mootien, S. Davies, S.C. Bibb, M.J. Lawson, D.M. 2007-03-01 doi:10.1107/S1744309107007944 International Union of Crystallography A novel regulator of antibiotic production in S. coelicolor, AbsC, has been crystallized in space group P212121. X-ray data to 2.25 Å resolution were collected on station PX 14.1 at Daresbury. en ABSC; STREPTOMYCES; ANTIBIOTIC PRODUCTION; MARR HOMOLOGUE; TRANSCRIPTIONAL REGULATION Crystals of recombinant AbsC (subunit MW = 18 313 Da; 158 amino acids), a novel regulator of antibiotic production from Streptomyces coelicolor, were grown by vapour diffusion. The protein crystallizes in space group P212121, with unit-cell parameters a = 43.53, b = 121.30, c = 143.75 Å. Native data to a resolution of 2.25 Å were recorded at station PX 14.1 (Daresbury) from a single crystal. Preliminary analysis of these data suggests that the asymmetric unit contains four copies of the AbsC monomer, giving an estimated solvent content of 47.0%. AbsC belongs to the MarR family of proteins that mediate ligand-responsive transcriptional control. text/html Crystallization and preliminary X-ray analysis of AbsC, a novel regulator of antibiotic production in Streptomyces coelicolor text 3 63 2007-03-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2007 International Union of Crystallography 1744-3091 crystallization communications 233 med@iucr.org 235 1744-3091 http://scripts.iucr.org/cgi-bin/paper?gx5109 Bacillus anthracis is a spore-forming bacterium and the causative agent of the disease anthrax. The Oxford Protein Production Facility has been targeting proteins from B. anthracis in order to develop high-throughput technologies within the Structural Proteomics in Europe project. As part of this work, the structure of 5-formyltetrahydrofolate cyclo-ligase (BA4489) has been determined by X-ray crystallography to 1.6 Å resolution. The structure, solved in complex with magnesium-ion-bound ADP and phosphate, gives a detailed picture of the proposed catalytic mechanism of the enzyme. Chemical differences from other cyclo-ligase structures close to the active site that could be exploited to design specific inhibitors are also highlighted. Copyright (c) 2007 International Union of Crystallography urn:issn:1744-3091 Meier, C. Carter, L.G. Winter, G. Owens, R.J. Stuart, D.I. Esnouf, R.M. 2007-03-01 doi:10.1107/S1744309107007221 International Union of Crystallography The structure of 5-formyltetrahydrofolate cyclo-ligase from B. anthracis determined by X-ray crystallography at a resolution of 1.6 Å is described. en 5,10-METHENYLTETRAHYDROFOLATE SYNTHETASE; MTHFS Bacillus anthracis is a spore-forming bacterium and the causative agent of the disease anthrax. The Oxford Protein Production Facility has been targeting proteins from B. anthracis in order to develop high-throughput technologies within the Structural Proteomics in Europe project. As part of this work, the structure of 5-formyltetrahydrofolate cyclo-ligase (BA4489) has been determined by X-ray crystallography to 1.6 Å resolution. The structure, solved in complex with magnesium-ion-bound ADP and phosphate, gives a detailed picture of the proposed catalytic mechanism of the enzyme. Chemical differences from other cyclo-ligase structures close to the active site that could be exploited to design specific inhibitors are also highlighted. text/html Structure of 5-formyltetrahydrofolate cyclo-ligase from Bacillus anthracis (BA4489) text 3 63 2007-03-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2007 International Union of Crystallography 1744-3091 protein structure communications 168 med@iucr.org 172 1744-3091 http://scripts.iucr.org/cgi-bin/paper?hv5079 The X-ray crystal structure of a single-chain monellin protein (MNEI) has been determined at 1.15 Å resolution. The model was refined to convergence employing anisotropic displacement parameters and riding H atoms to produce a final model with Rwork and Rfree values of 0.132 and 0.162, respectively. The crystal contains a single MNEI protein in the asymmetric unit and unusually lacks the dimer interface observed in all previous crystal structures of monellin and its single-chain derivatives. The high resolution allowed a more detailed view of MNEI than previously possible, with 38 of the 96 residues modelled with alternative side-chain conformations, including four core residues Thr12, Cys41, Leu62 and Ile75. Four stably bound negative ions were also located, providing new insight into potential electrostatic interactions of MNEI with the largely negatively charged surface of the sweet taste receptor T1R2–T1R3. Copyright (c) 2007 International Union of Crystallography urn:issn:1744-3091 Hobbs, J.R. Munger, S.D. Conn, G.L. 2007-03-01 doi:10.1107/S1744309107005271 International Union of Crystallography The crystal structure of the sweet protein MNEI at 1.15 Å resolution reveals networks of alternate conformations and stably bound negative ions. en MONELLIN; SWEET PROTEIN; SWEET TASTE; T1R2-T1R3 RECEPTOR The X-ray crystal structure of a single-chain monellin protein (MNEI) has been determined at 1.15 Å resolution. The model was refined to convergence employing anisotropic displacement parameters and riding H atoms to produce a final model with Rwork and Rfree values of 0.132 and 0.162, respectively. The crystal contains a single MNEI protein in the asymmetric unit and unusually lacks the dimer interface observed in all previous crystal structures of monellin and its single-chain derivatives. The high resolution allowed a more detailed view of MNEI than previously possible, with 38 of the 96 residues modelled with alternative side-chain conformations, including four core residues Thr12, Cys41, Leu62 and Ile75. Four stably bound negative ions were also located, providing new insight into potential electrostatic interactions of MNEI with the largely negatively charged surface of the sweet taste receptor T1R2–T1R3. text/html Monellin (MNEI) at 1.15 Å resolution text 3 63 2007-03-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2007 International Union of Crystallography 1744-3091 protein structure communications 162 med@iucr.org 167 1744-3091 http://scripts.iucr.org/cgi-bin/paper?be5077 Crystals of a single-point mutant (T109S) of Escherichia coli dihydroorotase (DHOase) with diminished activity grown in the presence of l-dihydroorotate (l-DHO) are tetragonal, with a monomer in the asymmetric unit. These crystals are extremely unstable and disintegrate shortly after formation, which is followed by the growth of orthorhombic crystals from the remnants of the tetragonal crystals or at new nucleation sites. Orthorhombic crystals, for which a structure has previously been reported [Thoden et al. (2001), Biochemistry, 40, 6989–6997; Lee et al. (2005), J. Mol. Biol. 348, 523–533], contain a dimer of DHOase in the asymmetric unit; the active site of one monomer contains the substrate N-carbamyl-l-aspartate (l-CA-asp) and the active site of the other monomer contains the product of the reaction, l-DHO. In the subunit with l-­DHO in the active site, a surface loop (residues 105–115) is `open'. In the other subunit, with l-CA-asp in the active site, the loop folds inwards, forming specific hydrogen bonds from the loop to the l-CA-asp. The tetragonal crystal form can be stabilized by crystallization in the presence of the inhibitor 5-fluoroorotate (FOA), a product (l-DHO) mimic. Crystals of the complex of T109S DHOase with FOA are tetragonal, space group P41212, with unit-cell parameters a = b = 72.6, c = 176.1 Å. The structure has been refined to R and Rfree values of 0.218 and 0.257, despite severe anisotropy of the diffraction. In this structure, the flexible loops are both in the `open' conformation, which is consistent with FOA, like l-­DHO, binding at both sites. The behaviour of the T109S mutant crystals of DHOase in the presence of l-DHO is explained by initial binding of l-DHO to both subunits, followed by slow conversion to l-CA-asp, with consequent movement of the flexible loop and dissolution of the crystals. Orthorhombic crystals are then able to grow in the presence of l-DHO and l-CA-asp. Copyright (c) 2007 International Union of Crystallography urn:issn:1744-3091 Lee, M. Maher, M.J. Guss, J.M. 2007-03-01 doi:10.1107/S1744309107004009 International Union of Crystallography A single-point mutant (T109S) of E. coli dihydroorotase initially crystallizes so that the two monomers of the dimer are related by a crystallographic twofold axis. In the presence of substrate, conversion to the previously observed asymmetric dimer with substrate bound in one subunit and product in the other is observed. en DIHYDROOROTASE; CONFORMATIONAL CHANGE; LOOP MOVEMENT; CATALYTIC STATE; CRYSTAL CONTACTS; CRYSTAL INSTABILITY Crystals of a single-point mutant (T109S) of Escherichia coli dihydroorotase (DHOase) with diminished activity grown in the presence of l-dihydroorotate (l-DHO) are tetragonal, with a monomer in the asymmetric unit. These crystals are extremely unstable and disintegrate shortly after formation, which is followed by the growth of orthorhombic crystals from the remnants of the tetragonal crystals or at new nucleation sites. Orthorhombic crystals, for which a structure has previously been reported [Thoden et al. (2001), Biochemistry, 40, 6989–6997; Lee et al. (2005), J. Mol. Biol. 348, 523–533], contain a dimer of DHOase in the asymmetric unit; the active site of one monomer contains the substrate N-carbamyl-l-aspartate (l-CA-asp) and the active site of the other monomer contains the product of the reaction, l-DHO. In the subunit with l-­DHO in the active site, a surface loop (residues 105–115) is `open'. In the other subunit, with l-CA-asp in the active site, the loop folds inwards, forming specific hydrogen bonds from the loop to the l-CA-asp. The tetragonal crystal form can be stabilized by crystallization in the presence of the inhibitor 5-fluoroorotate (FOA), a product (l-DHO) mimic. Crystals of the complex of T109S DHOase with FOA are tetragonal, space group P41212, with unit-cell parameters a = b = 72.6, c = 176.1 Å. The structure has been refined to R and Rfree values of 0.218 and 0.257, despite severe anisotropy of the diffraction. In this structure, the flexible loops are both in the `open' conformation, which is consistent with FOA, like l-­DHO, binding at both sites. The behaviour of the T109S mutant crystals of DHOase in the presence of l-DHO is explained by initial binding of l-DHO to both subunits, followed by slow conversion to l-CA-asp, with consequent movement of the flexible loop and dissolution of the crystals. Orthorhombic crystals are then able to grow in the presence of l-DHO and l-CA-asp. text/html Structure of the T109S mutant of Escherichia coli dihydroorotase complexed with the inhibitor 5-­fluoroorotate: catalytic activity is reflected by the crystal form text 3 63 2007-03-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2007 International Union of Crystallography 1744-3091 protein structure communications 154 med@iucr.org 161 1744-3091 http://scripts.iucr.org/cgi-bin/paper?bo9002 A correction is made to the names of two of the authors in Mastrangelo et al. (2006), Acta Cryst. F62, 768–770. Copyright (c) 2007 International Union of Crystallography urn:issn:1744-3091 Mastrangelo, E. Bollati, M. Milani, M. De Lamballerie, X. Brisbarre, N. Dalle, K. Lantez, V. Egloff, M.-P. Coutard, B. Canard, B. Gould, E. Forrester, N. Bolognesi, M. 2007-03-01 doi:10.1107/S1744309107009098 International Union of Crystallography A corrigendum to the paper by Mastrangelo et al. (2006), Acta Cryst. F62, 768–770. en (NUCLEOSIDE-2'-O-)-METHYLTRANSFERASES; FLAVIVIRUSES; MEABAN VIRUS; YOKOSE VIRUS; CORRIGENDUM A correction is made to the names of two of the authors in Mastrangelo et al. (2006), Acta Cryst. F62, 768–770. text/html Preliminary characterization of (nucleoside-2′-O-)-methyltransferase crystals from Meaban and Yokose flaviviruses. Corrigendum text 3 63 2007-03-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2007 International Union of Crystallography 1744-3091 addenda and errata 252 med@iucr.org 252 1744-3091 http://scripts.iucr.org/cgi-bin/paper?hc5020 The enzyme ω-transaminase catalyses the conversion of chiral ω-amines to ketones. The recombinant enzyme from Chromobacterium violaceum has been purified to homogeneity. The enzyme was crystallized from PEG 4000 using the microbatch method. Data were collected to 1.7 Å resolution from a crystal belonging to the triclinic space group P1, with unit-cell parameters a = 58.9, b = 61.9, c = 63.9 Å, α = 71.9, β = 87.0, γ = 74.6°. Data were also collected to 1.95 Å from a second triclinic crystal form. The structure has been solved using the molecular-replacement method. Copyright (c) 2007 International Union of Crystallography urn:issn:1744-3091 Sayer, C. Isupov, M.N. Littlechild, J.A. 2007-02-01 doi:10.1107/S1744309107000863 International Union of Crystallography An ω-amino acid:pyruvate transaminase from C. violaceum has been purified and crystallized in two crystal forms. The structure has been solved using molecular replacement. en [OMEGA]-TRANSAMINASE; PYRIDOXAL 5'-PHOSPHATE The enzyme ω-transaminase catalyses the conversion of chiral ω-amines to ketones. The recombinant enzyme from Chromobacterium violaceum has been purified to homogeneity. The enzyme was crystallized from PEG 4000 using the microbatch method. Data were collected to 1.7 Å resolution from a crystal belonging to the triclinic space group P1, with unit-cell parameters a = 58.9, b = 61.9, c = 63.9 Å, α = 71.9, β = 87.0, γ = 74.6°. Data were also collected to 1.95 Å from a second triclinic crystal form. The structure has been solved using the molecular-replacement method. text/html Crystallization and preliminary X-ray diffraction analysis of ω-amino acid:pyruvate transaminase from Chromobacterium violaceum text 2 63 2007-02-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2007 International Union of Crystallography 1744-3091 crystallization communications 117 med@iucr.org 119 1744-3091 http://scripts.iucr.org/cgi-bin/paper?hc5023 The DnaD protein is an essential component of the chromosome-replication machinery of the Gram-positive bacterium Bacillus subtilis and is part of the primosomal cascade that ultimately loads the replicative ring helicase DnaC onto DNA. Moreover, DnaD is a global regulator of DNA architecture, as it forms higher order nucleoprotein structures in order to open supercoiled DNA. Here, the crystallization and preliminary X-ray diffraction analysis of the two domains of DnaD from B. subtilis are reported. Crystals of the N-terminal domain are trigonal, with either P3121 or P3221 space-group symmetry, and diffracted X-­rays to 2.0 Å resolution; crystals of the C-terminal domain are hexagonal, with space group P61 or P65, and diffracted X-rays to 2.9 Å resolution in-house. Determination of the structure of the DnaD domains will provide insight into how remodelling of the nucleoid is associated with priming of replication in the model Gram-positive organism B. subtilis. Copyright (c) 2007 International Union of Crystallography urn:issn:1744-3091 Schneider, S. Carneiro, M.J.V.M. Ioannou, C. Soultanas, P. Paoli, M. 2007-02-01 doi:10.1107/S1744309107000474 International Union of Crystallography Crystallization and preliminary X-ray diffraction analysis of the two domains of DnaD from B. subtilis is reported. en DNA REPLICATION; DNAD; BACILLUS SUBTILIS; PRIMOSOMAL CASCADE The DnaD protein is an essential component of the chromosome-replication machinery of the Gram-positive bacterium Bacillus subtilis and is part of the primosomal cascade that ultimately loads the replicative ring helicase DnaC onto DNA. Moreover, DnaD is a global regulator of DNA architecture, as it forms higher order nucleoprotein structures in order to open supercoiled DNA. Here, the crystallization and preliminary X-ray diffraction analysis of the two domains of DnaD from B. subtilis are reported. Crystals of the N-terminal domain are trigonal, with either P3121 or P3221 space-group symmetry, and diffracted X-­rays to 2.0 Å resolution; crystals of the C-terminal domain are hexagonal, with space group P61 or P65, and diffracted X-rays to 2.9 Å resolution in-house. Determination of the structure of the DnaD domains will provide insight into how remodelling of the nucleoid is associated with priming of replication in the model Gram-positive organism B. subtilis. text/html Crystallization and X-ray diffraction analysis of the DNA-remodelling protein DnaD from Bacillus subtilis text 2 63 2007-02-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2007 International Union of Crystallography 1744-3091 crystallization communications 110 med@iucr.org 113 1744-3091 http://scripts.iucr.org/cgi-bin/paper?bw9180 A correction is made to a statement in the article by Carr et al. (2006), Acta Cryst. F62, 1164–1167. Copyright (c) 2007 International Union of Crystallography urn:issn:1744-3091 Carr, S.B. Makris, G. Phillips, S.E.V. Thomas, C.D. 2007-01-01 doi:10.1107/S1744309106054650 International Union of Crystallography A corrigendum to the paper by Carr et al. (2006), Acta Cryst. F62, 1164–1167. en TOPOISOMERASE IV; DNA CLEAVAGE; QUINOLONE BINDING; TRANSLATIONAL NCS A correction is made to a statement in the article by Carr et al. (2006), Acta Cryst. F62, 1164–1167. text/html Crystallization and preliminary X-ray diffraction analysis of two N-terminal fragments of the DNA-cleavage domain of topoisomerase IV from Staphylococcus aureus. Corrigendum text 1 63 2007-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2007 International Union of Crystallography 1744-3091 addenda and errata 59 med@iucr.org 59 1744-3091 http://scripts.iucr.org/cgi-bin/paper?fw5110 YcdB is a periplasmic haem-containing protein from Escherichia coli that has a potential role in iron transport. It is currently the only reported haem-containing Tat-secreted substrate. Here, the overexpression, purification, crystallization and structure determination at 2.0 Å resolution are reported for the apo form of the protein. The apo-YcdB structure resembles those of members of the haem-dependent peroxidase family and thus confirms that YcdB is also a member of this family. Haem-soaking experiments with preformed apo-YcdB crystals have been optimized to successfully generate haem-containing YcdB crystals that diffract to 2.9 Å. Completion of model building and structure refinement are under way. Copyright (c) 2007 International Union of Crystallography urn:issn:1744-3091 Cartron, M.L. Mitchell, S.A. Woodhall, M.R. Andrews, S.C. Watson, K.A. 2007-01-01 doi:10.1107/S174430910604509X International Union of Crystallography The crystallization and structure determination of the apo form of a novel haem-containing Tat substrate, YcdB from E. coli, has been solved to 2.0 Å resolution. The preliminary structure shows similarity to other haem-dependent peroxidases, despite low sequence homology. en YCDN; YCDO; YCDB; FTR1P; TAT; IRON TRANSPORT; PEROXIDASES; DYP; ESCHERICHIA COLI YcdB is a periplasmic haem-containing protein from Escherichia coli that has a potential role in iron transport. It is currently the only reported haem-containing Tat-secreted substrate. Here, the overexpression, purification, crystallization and structure determination at 2.0 Å resolution are reported for the apo form of the protein. The apo-YcdB structure resembles those of members of the haem-dependent peroxidase family and thus confirms that YcdB is also a member of this family. Haem-soaking experiments with preformed apo-YcdB crystals have been optimized to successfully generate haem-containing YcdB crystals that diffract to 2.9 Å. Completion of model building and structure refinement are under way. text/html Preliminary X-ray diffraction analysis of YcdB from Escherichia coli: a novel haem-containing and Tat-­secreted periplasmic protein with a potential role in iron transport text 1 63 2007-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2007 International Union of Crystallography 1744-3091 crystallization communications 37 med@iucr.org 41 1744-3091 http://scripts.iucr.org/cgi-bin/paper?bw5180 DNA topoisomerase IV removes undesirable topological features from DNA molecules in order to help maintain chromosome stability. Two constructs of 56 and 59 kDa spanning the DNA-cleavage domain of the A subunit of topoisomerase IV from Staphylococcus aureus (termed GrlA56 and GrlA59) have been crystallized. Crystals were grown at 291 K using the sitting-drop vapour-diffusion technique with PEG 3350 as a precipitant. Preliminary X-­ray analysis revealed that GrlA56 crystals belong to space group P21, diffract to a resolution of 2.9 Å and possess unit-cell parameters a = 83.6, b = 171.5, c = 87.8 Å, β = 90.1°, while crystals of GrlA59 belong to space group P21212, with unit-cell parameters a = 41.5, b = 171.89, c = 87.9 Å. These crystals diffract to a resolution of 2.8 Å. This is the first report of the crystallization and preliminary X-ray analysis of the DNA-cleavage domain of a topoisomerase IV from a Gram-positive organism. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Carr, S.B. Makris, G. Phillips, S.E.V. Thomas, C.D. 2006-11-01 doi:10.1107/S1744309106044150 International Union of Crystallography The crystallization and data collection of topoisomerase IV from S. aureus is described. Phasing by molecular replacement proved difficult owing to the presence of translational NCS and strategies used to overcome this are discussed. en TOPOISOMERASE IV; DNA CLEAVAGE; QUINOLONE BINDING; TRANSLATIONAL NCS DNA topoisomerase IV removes undesirable topological features from DNA molecules in order to help maintain chromosome stability. Two constructs of 56 and 59 kDa spanning the DNA-cleavage domain of the A subunit of topoisomerase IV from Staphylococcus aureus (termed GrlA56 and GrlA59) have been crystallized. Crystals were grown at 291 K using the sitting-drop vapour-diffusion technique with PEG 3350 as a precipitant. Preliminary X-­ray analysis revealed that GrlA56 crystals belong to space group P21, diffract to a resolution of 2.9 Å and possess unit-cell parameters a = 83.6, b = 171.5, c = 87.8 Å, β = 90.1°, while crystals of GrlA59 belong to space group P21212, with unit-cell parameters a = 41.5, b = 171.89, c = 87.9 Å. These crystals diffract to a resolution of 2.8 Å. This is the first report of the crystallization and preliminary X-ray analysis of the DNA-cleavage domain of a topoisomerase IV from a Gram-positive organism. text/html Crystallization and preliminary X-ray diffraction analysis of two N-terminal fragments of the DNA-cleavage domain of topoisomerase IV from Staphylococcus aureus text 11 62 2006-11-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 1164 med@iucr.org 1167 1744-3091 http://scripts.iucr.org/cgi-bin/paper?bw5178 Crystals of recombinant CloQ (subunit MW = 35 626 Da; 324 amino acids), an aromatic prenyltransferase from Streptomyces roseochromogenes, were grown by vapour diffusion. The protein crystallizes in space group I4122, with unit-cell parameters a = b = 135.19, c = 98.13 Å. Native data from a single crystal were recorded to a resolution of 2.2 Å in-house. Preliminary analysis of these data indicated that the asymmetric unit corresponds to a monomer, giving an estimated solvent content of 60.6%. CloQ is involved in the biosynthesis of the aminocoumarin antibiotic clorobiocin, which targets the essential bacterial enzyme DNA gyrase. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Keller, S. Pojer, F. Heide, L. Lawson, D.M. 2006-11-01 doi:10.1107/S1744309106042527 International Union of Crystallography An aromatic prenyltransferase (CloQ) from S. roseochromogenes that is implicated in clorobiocin biosynthesis has been crystallized in space group I4122. X-ray data to 2.2 Å resolution were collected in-house. en CLOQ; PRENYLTRANSFERASES; STREPTOMYCES; CLOROBIOCIN; ANTIBIOTIC BIOSYNTHESIS Crystals of recombinant CloQ (subunit MW = 35 626 Da; 324 amino acids), an aromatic prenyltransferase from Streptomyces roseochromogenes, were grown by vapour diffusion. The protein crystallizes in space group I4122, with unit-cell parameters a = b = 135.19, c = 98.13 Å. Native data from a single crystal were recorded to a resolution of 2.2 Å in-house. Preliminary analysis of these data indicated that the asymmetric unit corresponds to a monomer, giving an estimated solvent content of 60.6%. CloQ is involved in the biosynthesis of the aminocoumarin antibiotic clorobiocin, which targets the essential bacterial enzyme DNA gyrase. text/html Crystallization and preliminary X-ray analysis of the aromatic prenyltransferase CloQ from the clorobiocin biosynthetic cluster of Streptomyces roseochromogenes text 11 62 2006-11-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 1153 med@iucr.org 1155 1744-3091 http://scripts.iucr.org/cgi-bin/paper?sw5016 The structure of recombinant Aquifex aeolicus UDP-3-O-acyl-N-acetylglucosamine deacetylase (LpxC) in complex with UDP has been determined to a resolution of 2.2 Å. Previous studies have characterized the binding sites of the fatty-acid and sugar moieties of the substrate, UDP-(3-O-hydroxymyristoyl)-N-­acetylglucosamine, but not that of the nucleotide. The uracil-binding site is constructed from amino acids that are highly conserved across species. Hydrophobic associations with the Phe155 and Arg250 side chains in combination with hydrogen-bonding interactions with the main chain of Glu154 and the side chains of Tyr151 and Lys227 position the base. The phosphate and ribose groups are directed away from the active site and interact with Arg137, Lys156, Glu186 and Arg250. The orientation of the phosphate-ribose tail is not conducive to catalysis, perhaps owing to the position of an inhibitory Zn2+. However, based on the position of uracil revealed in this study and on the previously reported complex of LpxC with an inhibitor, a model is proposed for substrate binding. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Buetow, L. Dawson, A. Hunter, W.N. 2006-11-01 doi:10.1107/S1744309106041893 International Union of Crystallography The structure of UDP-3-O-acyl-N-acetylglucosamine deacetylase (LpxC) in complex with UDP is reported. The complex allows for a description of how the enzyme recognizes and binds a nucleotide moiety and enables the construction of an LpxC-substrate model. en LIPID A; AQUIFEX AEOLICUS; LPXC The structure of recombinant Aquifex aeolicus UDP-3-O-acyl-N-acetylglucosamine deacetylase (LpxC) in complex with UDP has been determined to a resolution of 2.2 Å. Previous studies have characterized the binding sites of the fatty-acid and sugar moieties of the substrate, UDP-(3-O-hydroxymyristoyl)-N-­acetylglucosamine, but not that of the nucleotide. The uracil-binding site is constructed from amino acids that are highly conserved across species. Hydrophobic associations with the Phe155 and Arg250 side chains in combination with hydrogen-bonding interactions with the main chain of Glu154 and the side chains of Tyr151 and Lys227 position the base. The phosphate and ribose groups are directed away from the active site and interact with Arg137, Lys156, Glu186 and Arg250. The orientation of the phosphate-ribose tail is not conducive to catalysis, perhaps owing to the position of an inhibitory Zn2+. However, based on the position of uracil revealed in this study and on the previously reported complex of LpxC with an inhibitor, a model is proposed for substrate binding. text/html The nucleotide-binding site of Aquifex aeolicus LpxC text 11 62 2006-11-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 protein structure communications 1082 med@iucr.org 1086 1744-3091 http://scripts.iucr.org/cgi-bin/paper?en5202 Pyrrolnitrin is the active ingredient of drugs for the treatment of superficial fungal infections and was used as a lead structure for the development of fludioxonil. It is an effective agent for plant diseases caused by the fungal pathogen Rhizoctonia solani. Pyrrolnitrin is made in four steps, the second of which, catalyzed by PrnB, is a novel chemical rearrangement of 7-chlorotryptophan. PrnB was overproduced in Pseudomonas fluorescens (BL915) and well diffracting crystals were obtained of a triple cysteine-to-serine mutant by sitting-drop vapour diffusion. Crystals grown in the presence of l-7-chlorotryptophan, d-­tryptophan and l-tryptophan are reported. Data sets for each are reported with high-resolution limits of 2.0, 1.75 and 1.75 Å, respectively. Two crystals (PrnB in the presence of d-tryptophan and l-7-chlorotryptophan) belong to space group C2 with similar unit-cell parameters (a = 68.6, b = 79.5, c = 92.7 Å, α = γ = 90.0, β = 103.8°). Crystals grown in the presence of l-­tryptophan belong to space group C2221 and have unit-cell parameters a = 67.7, b = 80.1, c = 129.5 Å. All crystals contain a monomer in the asymmetric unit. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 De Laurentis, W. Leang, K. Hahn, K. Podemski, B. Adam, A. Kroschwald, S. Carter, L.G. van Pee, K.-H. Naismith, J.H. 2006-11-01 doi:10.1107/S1744309106041649 International Union of Crystallography Crystals of PrnB, the second enzyme in pyrrolnitrin biosynthesis are reported. en PRNB; PYRROLNITRIN BIOSYNTHESIS Pyrrolnitrin is the active ingredient of drugs for the treatment of superficial fungal infections and was used as a lead structure for the development of fludioxonil. It is an effective agent for plant diseases caused by the fungal pathogen Rhizoctonia solani. Pyrrolnitrin is made in four steps, the second of which, catalyzed by PrnB, is a novel chemical rearrangement of 7-chlorotryptophan. PrnB was overproduced in Pseudomonas fluorescens (BL915) and well diffracting crystals were obtained of a triple cysteine-to-serine mutant by sitting-drop vapour diffusion. Crystals grown in the presence of l-7-chlorotryptophan, d-­tryptophan and l-tryptophan are reported. Data sets for each are reported with high-resolution limits of 2.0, 1.75 and 1.75 Å, respectively. Two crystals (PrnB in the presence of d-tryptophan and l-7-chlorotryptophan) belong to space group C2 with similar unit-cell parameters (a = 68.6, b = 79.5, c = 92.7 Å, α = γ = 90.0, β = 103.8°). Crystals grown in the presence of l-­tryptophan belong to space group C2221 and have unit-cell parameters a = 67.7, b = 80.1, c = 129.5 Å. All crystals contain a monomer in the asymmetric unit. text/html Preliminary crystallographic characterization of PrnB, the second enzyme in the pyrrolnitrin biosynthetic pathway text 11 62 2006-11-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 1134 med@iucr.org 1137 1744-3091 http://scripts.iucr.org/cgi-bin/paper?hc5016 Ranasmurfin, a previously uncharacterized ∼13 kDa blue protein found in the nests of the frog Polypedates leucomystax, has been purified and crystallized. The crystals are an intense blue colour and diffract to 1.51 Å with P21 symmetry and unit-cell parameters a = 40.9, b = 59.9, c = 45.0 Å, β = 93.3°. Self-rotation function analysis indicates the presence of a dimer in the asymmetric unit. Biochemical data suggest that the blue colour of the protein is related to dimer formation. Sequence data for the protein are incomplete, but thus far have identified no model for molecular replacement. A fluorescence scan shows a peak at 9.676 keV, indicating that the protein binds zinc and suggesting a route for structure solution. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 McMahon, S.A. Walsh, M.A. Ching, R.T.Y. Carter, L.G. Dorward, M. Johnson, K.A. Liu, H. Oke, M. Bloch, C. Kennedy, M.W. Latiff, A.A. Cooper, A. Taylor, G.L. White, M.F. Naismith, J.H. 2006-11-01 doi:10.1107/S1744309106040036 International Union of Crystallography A novel blue protein from frog nests has been crystallized. en RANASMURFIN Ranasmurfin, a previously uncharacterized ∼13 kDa blue protein found in the nests of the frog Polypedates leucomystax, has been purified and crystallized. The crystals are an intense blue colour and diffract to 1.51 Å with P21 symmetry and unit-cell parameters a = 40.9, b = 59.9, c = 45.0 Å, β = 93.3°. Self-rotation function analysis indicates the presence of a dimer in the asymmetric unit. Biochemical data suggest that the blue colour of the protein is related to dimer formation. Sequence data for the protein are incomplete, but thus far have identified no model for molecular replacement. A fluorescence scan shows a peak at 9.676 keV, indicating that the protein binds zinc and suggesting a route for structure solution. text/html Crystallization of Ranasmurfin, a blue-coloured protein from Polypedates leucomystax text 11 62 2006-11-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 1124 med@iucr.org 1126 1744-3091 http://scripts.iucr.org/cgi-bin/paper?fw5102 Escherichia coli bacterioferritin was serendipitously crystallized in a novel cubic crystal form and its structure could be determined to 2.5 Å resolution despite a high degree of merohedral twinning. This is the first report of crystallographic data on `as-isolated' E. coli bacterioferritin. The ferroxidase active site contains positive difference density consistent with two metal ions that had co-purified with the protein. X-ray fluorescence studies suggest that the metal composition is different from that of previous structures and is a mix of zinc and native iron ions. The ferroxidase-centre configuration displays a similar flexibility as previously noted for other bacterioferritins. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 van Eerde, A. Wolterink-van Loo, S. van der Oost, J. Dijkstra, B.W. 2006-11-01 doi:10.1107/S1744309106039583 International Union of Crystallography E. coli bacterioferritin was crystallized in a novel crystal form from different conditions and the structure was solved. The crystals belonged to space group P213 and diffracted to a resolution of 2.5 Å. en ESCHERICHIA COLI BACTERIOFERRITIN; IRON STORAGE AND HOMEOSTASIS; FERROXIDASE; MEROHEDRAL TWINNING Escherichia coli bacterioferritin was serendipitously crystallized in a novel cubic crystal form and its structure could be determined to 2.5 Å resolution despite a high degree of merohedral twinning. This is the first report of crystallographic data on `as-isolated' E. coli bacterioferritin. The ferroxidase active site contains positive difference density consistent with two metal ions that had co-purified with the protein. X-ray fluorescence studies suggest that the metal composition is different from that of previous structures and is a mix of zinc and native iron ions. The ferroxidase-centre configuration displays a similar flexibility as previously noted for other bacterioferritins. text/html Fortuitous structure determination of `as-isolated' Escherichia coli bacterioferritin in a novel crystal form text 11 62 2006-11-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 protein structure communications 1061 med@iucr.org 1066 1744-3091 http://scripts.iucr.org/cgi-bin/paper?pu5149 Laccases are members of the blue multi-copper oxidase family that oxidize substrate molecules by accepting electrons at a mononuclear copper centre and transferring them to a trinuclear centre. Dioxygen binds to the trinuclear centre and, following the transfer of four electrons, is reduced to two molecules of water. Crystals of the laccase from Cerrena maxima have been obtained and X-­ray data were collected to 1.9 Å resolution using synchrotron radiation. A preliminary analysis shows that the enzyme has the typical laccase structure and several carbohydrate sites have been identified. The carbohydrate chains appear to be involved in stabilization of the intermolecular contacts in the crystal structure, thus promoting the formation of well ordered crystals of the enzyme. Here, the results of an X-ray crystallographic study on the laccase from the fungus Cerrena maxima are reported. Crystals that diffract well to a resolution of at least 1.9 Å (R factor = 18.953%; Rfree = 23.835; r.m.s.d. bond lengths, 0.06 Å; r.m.s.d. bond angles, 1.07°) have been obtained despite the presence of glycan moieties. The overall spatial organization of C. maxima laccase and the structure of its copper-containing active centre have been determined by the molecular-replacement method using the laccase from Trametes versicolor (Piontek et al., 2002) as a structural template. In addition, four glycan-binding sites were identified and the 1.9 Å X-ray data were used to determine the previously unknown primary structure of this protein. The identity (calculated from sequence alignment) between the C. maxima laccase and the T. versicolor laccase is about 87%. Tyr196 and Tyr372 show significant extra density at the ortho positions and this has been interpreted in terms of NO2 substituents. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Lyashenko, A.V. Zhukhlistova, N.E. Gabdoulkhakov, A.G. Zhukova, Y.N. Voelter, W. Zaitsev, V.N. Bento, I. Stepanova, E.V. Kachalova, G.S. Koroleva, O.V. Cherkashyn, E.A. Tishkov, V.I. Lamzin, V.S. Schirwitz, K. Morgunova, E.Y. Betzel, C. Lindley, P.F. Mikhailov, A.M. 2006-10-01 doi:10.1107/S1744309106036578 International Union of Crystallography The crystallization and preliminary X-ray structure at 1.9 Å resolution of the fungal laccase from C. maxima are presented. en BLUE MULTI-COPPER ENZYMES; LACCASES; CERRENA MAXIMA Laccases are members of the blue multi-copper oxidase family that oxidize substrate molecules by accepting electrons at a mononuclear copper centre and transferring them to a trinuclear centre. Dioxygen binds to the trinuclear centre and, following the transfer of four electrons, is reduced to two molecules of water. Crystals of the laccase from Cerrena maxima have been obtained and X-­ray data were collected to 1.9 Å resolution using synchrotron radiation. A preliminary analysis shows that the enzyme has the typical laccase structure and several carbohydrate sites have been identified. The carbohydrate chains appear to be involved in stabilization of the intermolecular contacts in the crystal structure, thus promoting the formation of well ordered crystals of the enzyme. Here, the results of an X-ray crystallographic study on the laccase from the fungus Cerrena maxima are reported. Crystals that diffract well to a resolution of at least 1.9 Å (R factor = 18.953%; Rfree = 23.835; r.m.s.d. bond lengths, 0.06 Å; r.m.s.d. bond angles, 1.07°) have been obtained despite the presence of glycan moieties. The overall spatial organization of C. maxima laccase and the structure of its copper-containing active centre have been determined by the molecular-replacement method using the laccase from Trametes versicolor (Piontek et al., 2002) as a structural template. In addition, four glycan-binding sites were identified and the 1.9 Å X-ray data were used to determine the previously unknown primary structure of this protein. The identity (calculated from sequence alignment) between the C. maxima laccase and the T. versicolor laccase is about 87%. Tyr196 and Tyr372 show significant extra density at the ortho positions and this has been interpreted in terms of NO2 substituents. text/html Purification, crystallization and preliminary X-ray study of the fungal laccase from Cerrena maxima text 10 62 2006-10-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 protein structure communications 954 med@iucr.org 957 1744-3091 http://scripts.iucr.org/cgi-bin/paper?hv5065 Nucleotide monophosphate kinases (NMPKs) are potential antimicrobial drug targets owing to their role in supplying DNA and RNA precursors. The present work reports the crystal structure of Staphylococcus aureus guanylate monophosphate kinase (SaGMK) at 1.9 Å resolution. The structure shows that unlike most GMKs SaGMK is dimeric, confirming the role of the extended C-­terminus in dimer formation as first observed for Escherichia coli GMK (EcGMK). One of the two SaGMK dimers within the crystal asymmetric unit has two monomers in different conformations: an open form with a bound sulfate ion (mimicking the β-phosphate of ATP) and a closed form with bound GMP and sulfate ion. GMP-induced domain movements in SaGMK can thus be defined by comparison of these conformational states. Like other GMKs, the binding of GMP firstly triggers a partial closure of the enzyme, diminishing the distance between the GMP-binding and ATP-binding sites. In addition, the closed structure shows the presence of a potassium ion in contact with the guanine ring of GMP. The potassium ion appears to form an integral part of the GMP-binding site, as the Tyr36 side chain has significantly moved to form a metal ion–ligand coordination involving the lone pair of the side-chain O atom. The potassium-binding site might also be exploited in the design of novel inhibitors. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 El Omari, K. Dhaliwal, B. Lockyer, M. Charles, I. Hawkins, A.R. Stammers, D.K. 2006-10-01 doi:10.1107/S174430910603613X International Union of Crystallography The crystal structure of S. aureus guanylate monophosphate kinase has been determined to 1.9 Å resolution, revealing both open and closed forms within the asymmetric unit. These structures may be of use in anti-bacterial drug design. en GUANYLATE MONOPHOSPHATE KINASE; NUCLEOTIDE MONOPHOSPHATE KINASES; ANTIMICROBIALS Nucleotide monophosphate kinases (NMPKs) are potential antimicrobial drug targets owing to their role in supplying DNA and RNA precursors. The present work reports the crystal structure of Staphylococcus aureus guanylate monophosphate kinase (SaGMK) at 1.9 Å resolution. The structure shows that unlike most GMKs SaGMK is dimeric, confirming the role of the extended C-­terminus in dimer formation as first observed for Escherichia coli GMK (EcGMK). One of the two SaGMK dimers within the crystal asymmetric unit has two monomers in different conformations: an open form with a bound sulfate ion (mimicking the β-phosphate of ATP) and a closed form with bound GMP and sulfate ion. GMP-induced domain movements in SaGMK can thus be defined by comparison of these conformational states. Like other GMKs, the binding of GMP firstly triggers a partial closure of the enzyme, diminishing the distance between the GMP-binding and ATP-binding sites. In addition, the closed structure shows the presence of a potassium ion in contact with the guanine ring of GMP. The potassium ion appears to form an integral part of the GMP-binding site, as the Tyr36 side chain has significantly moved to form a metal ion–ligand coordination involving the lone pair of the side-chain O atom. The potassium-binding site might also be exploited in the design of novel inhibitors. text/html Structure of Staphylococcus aureus guanylate monophosphate kinase text 10 62 2006-10-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 protein structure communications 949 med@iucr.org 953 1744-3091 http://scripts.iucr.org/cgi-bin/paper?bw5160 The gene encoding 2-oxo-hept-3-ene-1,7-dioic acid (OHED) hydratase (HpcG) was cloned into the high-expression plasmid pET26b and overexpressed in Escherichia coli BL21(DE3). The enzyme was purified in three steps to greater than 95% purity prior to crystallization. Crystals were obtained by the hanging-drop vapour-diffusion method at 277 K in a number of screening conditions. Crystals measuring up to 1.5 mm in their longest dimension were grown from solutions containing polyethylene glycol 20 000. The crystals belonged to space group P41212 or P43212, with unit-cell parameters a = 136, b = 136, c = 192 Å. A complete data set was collected to 2.1 Å from a single cryocooled crystal at 100 K using synchrotron radiation. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Adachi, T. Izumi, A. Rea, D. Park, S.-Y. Tame, J.R.H. Roper, D.I. 2006-10-01 doi:10.1107/S1744309106035901 International Union of Crystallography The gene encoding HpcG from the homoprotocatechuate (4-hydroxyphenylacetic acid) degradative pathway of E. coli C has been cloned and expressed and the protein has been purified. Crystals obtained from the purified recombinant enzyme, belonging to a tetragonal space group, diffracted to a resolution of 2.1 Å. en HOMOPROTOCATECHUATE; 4-HYDROXYPHENYLACETIC ACID; HYDRATASES The gene encoding 2-oxo-hept-3-ene-1,7-dioic acid (OHED) hydratase (HpcG) was cloned into the high-expression plasmid pET26b and overexpressed in Escherichia coli BL21(DE3). The enzyme was purified in three steps to greater than 95% purity prior to crystallization. Crystals were obtained by the hanging-drop vapour-diffusion method at 277 K in a number of screening conditions. Crystals measuring up to 1.5 mm in their longest dimension were grown from solutions containing polyethylene glycol 20 000. The crystals belonged to space group P41212 or P43212, with unit-cell parameters a = 136, b = 136, c = 192 Å. A complete data set was collected to 2.1 Å from a single cryocooled crystal at 100 K using synchrotron radiation. text/html Expression, purification and crystallization of 2-­oxo-hept-4-ene-1,7-dioate hydratase (HpcG) from Escherichia coli C text 10 62 2006-10-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 1010 med@iucr.org 1012 1744-3091 http://scripts.iucr.org/cgi-bin/paper?pu5150 Pyridoxal kinases (PdxK) are able to catalyse the phosphorylation of three vitamin B6 precursors, pyridoxal, pyridoxine and pyridoxamine, to their 5′-­phosphates and play an important role in the vitamin B6 salvage pathway. Recently, the thiD gene of Bacillus subtilis was found to encode an enzyme which has the activity expected of a pyridoxal kinase despite its previous assignment as an HMPP kinase owing to higher sequence similarity. As such, this enzyme would appear to represent a new class of `HMPP kinase-like' pyridoxal kinases. B. subtilis thiD has been cloned and the protein has been overexpressed in Escherichia coli, purified and subsequently crystallized in a binary complex with ADP and Mg2+. X-ray diffraction data have been collected from crystals to 2.8 Å resolution at 100 K. The crystals belong to a primitive tetragonal system, point group 422, and analysis of the systematic absences suggest that they belong to one of the enantiomorphic pair of space groups P41212 or P43212. Consideration of the space-group symmetry and unit-cell parameters (a = b = 102.9, c = 252.6 Å, α = β = γ = 90°) suggest that the crystals contain between three and six molecules in the asymmetric unit. A full structure determination is under way to provide insights into aspects of the enzyme mechanism and substrate specificity. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Newman, J.A. Das, S.K. Sedelnikova, S.E. Rice, D.W. 2006-10-01 doi:10.1107/S1744309106035779 International Union of Crystallography A putative pyridoxal kinase from B. subtilis has been cloned, overexpressed, purified and crystallized and data have been collected to 2.8 Å resolution. en THID; PDXK; HMPP KINASE; PYRIDOXAL KINASE; RIBOKINASE SUPERFAMILY Pyridoxal kinases (PdxK) are able to catalyse the phosphorylation of three vitamin B6 precursors, pyridoxal, pyridoxine and pyridoxamine, to their 5′-­phosphates and play an important role in the vitamin B6 salvage pathway. Recently, the thiD gene of Bacillus subtilis was found to encode an enzyme which has the activity expected of a pyridoxal kinase despite its previous assignment as an HMPP kinase owing to higher sequence similarity. As such, this enzyme would appear to represent a new class of `HMPP kinase-like' pyridoxal kinases. B. subtilis thiD has been cloned and the protein has been overexpressed in Escherichia coli, purified and subsequently crystallized in a binary complex with ADP and Mg2+. X-ray diffraction data have been collected from crystals to 2.8 Å resolution at 100 K. The crystals belong to a primitive tetragonal system, point group 422, and analysis of the systematic absences suggest that they belong to one of the enantiomorphic pair of space groups P41212 or P43212. Consideration of the space-group symmetry and unit-cell parameters (a = b = 102.9, c = 252.6 Å, α = β = γ = 90°) suggest that the crystals contain between three and six molecules in the asymmetric unit. A full structure determination is under way to provide insights into aspects of the enzyme mechanism and substrate specificity. text/html Cloning, purification and preliminary crystallographic analysis of a putative pyridoxal kinase from Bacillus subtilis text 10 62 2006-10-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 1006 med@iucr.org 1009 1744-3091 http://scripts.iucr.org/cgi-bin/paper?en5189 PCNA is a ring-shaped protein that encircles DNA, providing a platform for the association of a wide variety of DNA-processing enzymes that utilize the PCNA sliding clamp to maintain proximity to their DNA substrates. PCNA is a homotrimer in eukaryotes, but a heterotrimer in crenarchaea such as Sulfolobus solfataricus. The three proteins are SsoPCNA1 (249 residues), SsoPCNA2 (245 residues) and SsoPCNA3 (259 residues). The heterotrimeric protein crystallizes in space group P21, with unit-cell parameters a = 44.8, b = 78.8, c = 125.6 Å, β = 100.5°. The crystal structure of this heterotrimeric PCNA molecule has been solved using molecular replacement. The resulting structure to 2.3 Å sheds light on the differential stabilities of the interactions observed between the three subunits and the specificity of individual subunits for partner proteins. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Williams, G.J. Johnson, K. Rudolf, J. McMahon, S.A. Carter, L. Oke, M. Liu, H. Taylor, G.L. White, M.F. Naismith, J.H. 2006-10-01 doi:10.1107/S1744309106034075 International Union of Crystallography The structure of the heterotrimeric PCNA complex from S. sulfataricus is reported to 2.3 Å. en PCNA; SULFOLOBUS SOLFATARICUS PCNA is a ring-shaped protein that encircles DNA, providing a platform for the association of a wide variety of DNA-processing enzymes that utilize the PCNA sliding clamp to maintain proximity to their DNA substrates. PCNA is a homotrimer in eukaryotes, but a heterotrimer in crenarchaea such as Sulfolobus solfataricus. The three proteins are SsoPCNA1 (249 residues), SsoPCNA2 (245 residues) and SsoPCNA3 (259 residues). The heterotrimeric protein crystallizes in space group P21, with unit-cell parameters a = 44.8, b = 78.8, c = 125.6 Å, β = 100.5°. The crystal structure of this heterotrimeric PCNA molecule has been solved using molecular replacement. The resulting structure to 2.3 Å sheds light on the differential stabilities of the interactions observed between the three subunits and the specificity of individual subunits for partner proteins. text/html Structure of the heterotrimeric PCNA from Sulfolobus solfataricus text 10 62 2006-10-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 protein structure communications 944 med@iucr.org 948 1744-3091 http://scripts.iucr.org/cgi-bin/paper?bo5004 Hsp70 is an important molecular chaperone involved in the regulation of protein folding. Crystals of the C-terminal 10 kDa helical lid domain (residues 542–640) from a Caenorhabditis elegans Hsp70 homologue have been produced that diffract X-rays to ∼3.4 Å. Crystals belong to space group I212121, with unit-cell parameters a = b = 197, c = 200 Å. The Matthews coefficient, self-rotation function and Patterson map indicate 24 monomers in the asymmetric unit, showing non-crystallographic 432 symmetry. Molecular-replacement studies using the corresponding domain from rat, the only eukaryotic homologue with a known structure, failed and a mercury derivative was obtained. Preliminary MAD phasing using SHELXD and SHARP for location and refinement of the heavy-atom substructure and SOLOMON for density modification produced interpretable maps with a clear protein–solvent boundary. Further density-modification, model-building and refinement are currently under way. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Worrall, L. Walkinshaw, M.D. 2006-09-01 doi:10.1107/S1744309106032064 International Union of Crystallography Crystals of the C-terminal 10 kDa lid subdomain from the C. elegans chaperone Hsp70 have been obtained that diffract X-rays to ∼3.5 Å and belong to space group I212121. Analysis of X-ray data and initial heavy-atom phasing reveals 24 monomers in the asymmetric unit related by 432 non-crystallographic symmetry. en HSP70; CHAPERONE; C. ELEGANS Hsp70 is an important molecular chaperone involved in the regulation of protein folding. Crystals of the C-terminal 10 kDa helical lid domain (residues 542–640) from a Caenorhabditis elegans Hsp70 homologue have been produced that diffract X-rays to ∼3.4 Å. Crystals belong to space group I212121, with unit-cell parameters a = b = 197, c = 200 Å. The Matthews coefficient, self-rotation function and Patterson map indicate 24 monomers in the asymmetric unit, showing non-crystallographic 432 symmetry. Molecular-replacement studies using the corresponding domain from rat, the only eukaryotic homologue with a known structure, failed and a mercury derivative was obtained. Preliminary MAD phasing using SHELXD and SHARP for location and refinement of the heavy-atom substructure and SOLOMON for density modification produced interpretable maps with a clear protein–solvent boundary. Further density-modification, model-building and refinement are currently under way. text/html Crystallization and X-ray data analysis of the 10 kDa C-terminal lid subdomain from Caenorhabditis elegans Hsp70 text 9 62 2006-09-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 938 med@iucr.org 943 1744-3091 http://scripts.iucr.org/cgi-bin/paper?hv5057 Recombinant Bacillus halmapalus α-amylase (BHA) was studied in two different crystal forms. The first crystal form was obtained by crystallization of BHA at room temperature in the presence of acarbose and maltose; data were collected at cryogenic temperature to a resolution of 1.9 Å. It was found that the crystal belonged to space group P212121, with unit-cell parameters a = 47.0, b = 73.5, c = 151.1 Å. A maltose molecule was observed and found to bind to BHA and previous reports of the binding of a nonasaccharide were confirmed. The second crystal form was obtained by pH-induced crystallization of BHA in a MES–HEPES–boric acid buffer (MHB buffer) at 303 K; the solubility of BHA in MHB has a retrograde temperature dependency and crystallization of BHA was only possible by raising the temperature to at least 298 K. Data were collected at cryogenic temperature to a resolution of 2.0 Å. The crystal belonged to space group P212121, with unit-cell parameters a = 38.6, b = 59.0, c = 209.8 Å. The structure was solved using molecular replacement. The maltose-binding site is described and the two structures are compared. No significant changes were seen in the structure upon binding of the substrates. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Lyhne-Iversen, L. Hobley, T.J. Kaasgaard, S.G. Harris, P. 2006-09-01 doi:10.1107/S174430910603096X International Union of Crystallography The structure of an uncomplexed form of α-amylase from B. halmapalus is compared with a form in which maltose, glucose and a nonasaccharide derived from acarbose and maltose are bound. en [ALPHA]-AMYLASE; MALTOSE-BINDING SITE; BACILLUS HALMAPALUS Recombinant Bacillus halmapalus α-amylase (BHA) was studied in two different crystal forms. The first crystal form was obtained by crystallization of BHA at room temperature in the presence of acarbose and maltose; data were collected at cryogenic temperature to a resolution of 1.9 Å. It was found that the crystal belonged to space group P212121, with unit-cell parameters a = 47.0, b = 73.5, c = 151.1 Å. A maltose molecule was observed and found to bind to BHA and previous reports of the binding of a nonasaccharide were confirmed. The second crystal form was obtained by pH-induced crystallization of BHA in a MES–HEPES–boric acid buffer (MHB buffer) at 303 K; the solubility of BHA in MHB has a retrograde temperature dependency and crystallization of BHA was only possible by raising the temperature to at least 298 K. Data were collected at cryogenic temperature to a resolution of 2.0 Å. The crystal belonged to space group P212121, with unit-cell parameters a = 38.6, b = 59.0, c = 209.8 Å. The structure was solved using molecular replacement. The maltose-binding site is described and the two structures are compared. No significant changes were seen in the structure upon binding of the substrates. text/html Structure of Bacillus halmapalus α-amylase crystallized with and without the substrate analogue acarbose and maltose text 9 62 2006-09-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 protein structure communications 849 med@iucr.org 854 1744-3091 http://scripts.iucr.org/cgi-bin/paper?gx5100 The structure of the NAD-dependent oxidoreductase UDP-galactose-4′-epimerase from Trypanosoma brucei in complex with cofactor and the substrate analogue UDP-4-deoxy-4-fluoro-α-d-galactose has been determined using diffraction data to 2.7 Å resolution. Despite the high level of sequence and structure conservation between the trypanosomatid enzyme and those from humans, yeast and bacteria, the binding of the 4-fluoro-α-d-galactose moiety is distinct from previously reported structures. Of particular note is the observation that when bound to the T. brucei enzyme, the galactose moiety of this fluoro-derivative is rotated approximately 180° with respect to the orientation of the hexose component of UDP-glucose when in complex with the human enzyme. The architecture of the catalytic centre is designed to effectively bind different orientations of the hexose, a finding that is consistent with a mechanism that requires the sugar to maintain a degree of flexibility within the active site. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Alphey, M.S. Burton, A. Urbaniak, M.D. Boons, G.-J. Ferguson, M.A.J. Hunter, W.N. 2006-09-01 doi:10.1107/S1744309106028740 International Union of Crystallography The structure of recombinant T. brucei UDP-galactose-4′-epimerase cocrystallized with NAD+ and the substrate analogue UDP-4-deoxy-4-fluoro-α-d-galactose has been determined at medium resolution. Comparisons with structures of human and E. coli UDP-galactose-4′-epimerase–ligand complexes reveal that the hexose moieties are able to adopt different orientations in the active site. en SHORT-CHAIN DEHYDROGENASE/REDUCTASES; TRYPANOSOMA BRUCEI; UDP-GALACTOSE-4'-EPIMERASE; UDP-4-DEOXY-4-FLUORO-[ALPHA]-D-GALACTOSE The structure of the NAD-dependent oxidoreductase UDP-galactose-4′-epimerase from Trypanosoma brucei in complex with cofactor and the substrate analogue UDP-4-deoxy-4-fluoro-α-d-galactose has been determined using diffraction data to 2.7 Å resolution. Despite the high level of sequence and structure conservation between the trypanosomatid enzyme and those from humans, yeast and bacteria, the binding of the 4-fluoro-α-d-galactose moiety is distinct from previously reported structures. Of particular note is the observation that when bound to the T. brucei enzyme, the galactose moiety of this fluoro-derivative is rotated approximately 180° with respect to the orientation of the hexose component of UDP-glucose when in complex with the human enzyme. The architecture of the catalytic centre is designed to effectively bind different orientations of the hexose, a finding that is consistent with a mechanism that requires the sugar to maintain a degree of flexibility within the active site. text/html Trypanosoma brucei UDP-galactose-4′-epimerase in ternary complex with NAD+ and the substrate analogue UDP-4-deoxy-4-fluoro-α-d-galactose text 9 62 2006-09-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 protein structure communications 829 med@iucr.org 834 1744-3091 http://scripts.iucr.org/cgi-bin/paper?bw5156 African sleeping sickness, also called trypanosomiasis, is a significant cause of morbidity and mortality in sub-Saharan Africa. Peptidases from Trypanosoma brucei, the causative agent, include the serine peptidase oligopeptidase B, a documented virulence factor and therapeutic target. Determination of the three-dimensional structure of oligopeptidase B is desirable to facilitate the development of novel inhibitors. Oligopeptidase B was overexpressed in Escherichia coli as an N-terminally hexahistidine-tagged fusion protein, purified using metal-affinity chromatography and crystallized using the hanging-drop vapour-diffusion technique in 7%(w/v) polyethylene glycol 6000, 1 M LiCl, 0.1 M bis-tris propane pH 7.5. Diffraction data to 2.7 Å resolution were collected using synchrotron radiation. The crystals belong to space group P3121 or P3221, with unit-cell parameters a = b = 124.5, c = 249.9 Å. A complete data set to 2.7 Å was collected using synchrotron radiation. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Rea, D. Hazell, C. Andrews, N.W. Morty, R.E. Fülöp, V. 2006-08-01 doi:10.1107/S1744309106027874 International Union of Crystallography Recombinant oligopeptidase B from T. brucei has been prepared and crystallized. Data were collected to 2.7 Å. Heavy-atom soaks and preparation of selenomethionine-substituted protein are in progress for structure determination by MAD or MIR. en OLIGOPEPTIDASE B; TRYPANOSOMA BRUCEI; AFRICAN TRYPANOSOMIASIS; SLEEPING SICKNESS African sleeping sickness, also called trypanosomiasis, is a significant cause of morbidity and mortality in sub-Saharan Africa. Peptidases from Trypanosoma brucei, the causative agent, include the serine peptidase oligopeptidase B, a documented virulence factor and therapeutic target. Determination of the three-dimensional structure of oligopeptidase B is desirable to facilitate the development of novel inhibitors. Oligopeptidase B was overexpressed in Escherichia coli as an N-terminally hexahistidine-tagged fusion protein, purified using metal-affinity chromatography and crystallized using the hanging-drop vapour-diffusion technique in 7%(w/v) polyethylene glycol 6000, 1 M LiCl, 0.1 M bis-tris propane pH 7.5. Diffraction data to 2.7 Å resolution were collected using synchrotron radiation. The crystals belong to space group P3121 or P3221, with unit-cell parameters a = b = 124.5, c = 249.9 Å. A complete data set to 2.7 Å was collected using synchrotron radiation. text/html Expression, purification and preliminary crystallographic analysis of oligopeptidase B from Trypanosoma brucei text 8 62 2006-08-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 808 med@iucr.org 810 1744-3091 http://scripts.iucr.org/cgi-bin/paper?fw5098 SA0961 is an unknown hypothetical protein from Staphylococcus aureus that can be identified in the Firmicutes division of Gram-positive bacteria. The gene for the homologue of SA0961 in Bacillus subtilis, ylaN, has been shown to be essential for cell survival, thus identifying the protein encoded by this gene as a potential target for the development of novel antibiotics. SA0961 was cloned and the protein was overexpressed in Escherichia coli, purified and subsequently crystallized. Crystals of selenomethionine-labelled SA0961 diffract to beyond 2.4 Å resolution and belong to the monoclinic space group P21, with unit-cell parameters a = 31.5, b = 42.7, c = 62.7 Å, β = 92.4° and two molecules in the asymmetric unit. A full structure determination is under way to provide insights into the function of this protein. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Xu, L. Sedelnikova, S.E. Baker, P.J. Rice, D.W. 2006-08-01 doi:10.1107/S1744309106027400 International Union of Crystallography SA0961 is an unknown hypothetical protein from Staphylococcus aureus that can be identified in the Firmicutes division of Gram-positive bacteria. SA0961 was cloned and the protein was overexpressed in Escherichia coli, purified and subsequently crystallized. en SA0961; YLAN; STAPHYLOCOCCUS AUREUS SA0961 is an unknown hypothetical protein from Staphylococcus aureus that can be identified in the Firmicutes division of Gram-positive bacteria. The gene for the homologue of SA0961 in Bacillus subtilis, ylaN, has been shown to be essential for cell survival, thus identifying the protein encoded by this gene as a potential target for the development of novel antibiotics. SA0961 was cloned and the protein was overexpressed in Escherichia coli, purified and subsequently crystallized. Crystals of selenomethionine-labelled SA0961 diffract to beyond 2.4 Å resolution and belong to the monoclinic space group P21, with unit-cell parameters a = 31.5, b = 42.7, c = 62.7 Å, β = 92.4° and two molecules in the asymmetric unit. A full structure determination is under way to provide insights into the function of this protein. text/html Cloning, purification and preliminary crystallographic analysis of a conserved hypothetical protein, SA0961 (YlaN), from Staphylococcus aureus text 8 62 2006-08-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 778 med@iucr.org 780 1744-3091 http://scripts.iucr.org/cgi-bin/paper?ll5068 A construct consisting of residues 10–310 of BipD, a component of the Burkholderia pseudomallei type III secretion system (T3SS), has been overexpressed as a GST fusion, cleaved from the GST tag and purified. Crystals were grown of native and selenomethionine-labelled BipD. The crystals grow in two different polymorphs from the same condition. The first polymorph belongs to space group C222, with unit-cell parameters a = 103.98, b = 122.79, c = 49.17 Å, a calculated Matthews coefficient of 2.4 Å3 Da−1 (47% solvent content) and one molecule per asymmetric unit. The second polymorph belongs to space group P21212, with unit-cell parameters a = 136.47, b = 89.84, c = 50.15 Å, and a calculated Matthews coefficient of 2.3 Å3 Da−1 (45% solvent content) for two molecules per asymmetric unit (analysis of the self-rotation function indicates the presence of a weak twofold non-crystallographic symmetry axis in this P21212 form). The native crystals of both forms give diffraction data to 2.7 Å resolution, while the SeMet-labelled P21212 crystals diffract to 3.3 Å resolution. A K2PtCl4 derivative of the P21212 form was also obtained and data were collected to 2.7 Å with radiation of wavelength λ = 0.933 Å. The Pt-derivative anomalous difference Patterson map revealed two self-peaks on the Harker sections. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Roversi, P. Johnson, S. Field, T. Deane, J.E. Galyov, E.E. Lea, S.M. 2006-09-01 doi:10.1107/S1744309106027035 International Union of Crystallography A construct consisting of residues 10–310 of mature BipD, a component of the B. pseudomallei type III secretion system, has been crystallized. Native BipD crystals and SeMet and K2PtCl4 derivative crystals have undergone preliminary crystallographic analysis. en BIPD; TYPE III SECRETION SYSTEM; BURKHOLDERIA PSEUDOMALLEI A construct consisting of residues 10–310 of BipD, a component of the Burkholderia pseudomallei type III secretion system (T3SS), has been overexpressed as a GST fusion, cleaved from the GST tag and purified. Crystals were grown of native and selenomethionine-labelled BipD. The crystals grow in two different polymorphs from the same condition. The first polymorph belongs to space group C222, with unit-cell parameters a = 103.98, b = 122.79, c = 49.17 Å, a calculated Matthews coefficient of 2.4 Å3 Da−1 (47% solvent content) and one molecule per asymmetric unit. The second polymorph belongs to space group P21212, with unit-cell parameters a = 136.47, b = 89.84, c = 50.15 Å, and a calculated Matthews coefficient of 2.3 Å3 Da−1 (45% solvent content) for two molecules per asymmetric unit (analysis of the self-rotation function indicates the presence of a weak twofold non-crystallographic symmetry axis in this P21212 form). The native crystals of both forms give diffraction data to 2.7 Å resolution, while the SeMet-labelled P21212 crystals diffract to 3.3 Å resolution. A K2PtCl4 derivative of the P21212 form was also obtained and data were collected to 2.7 Å with radiation of wavelength λ = 0.933 Å. The Pt-derivative anomalous difference Patterson map revealed two self-peaks on the Harker sections. text/html Expression, purification, crystallization and preliminary crystallographic analysis of BipD, a component of the Burkholderia pseudomallei type III secretion system text 9 62 2006-09-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 861 med@iucr.org 864 1744-3091 http://scripts.iucr.org/cgi-bin/paper?ll5069 IpaD, the putative needle-tip protein of the Shigella flexneri type III secretion system, has been overexpressed and purified. Crystals were grown of the native protein in space group P212121, with unit-cell parameters a = 55.9, b = 100.7, c = 112.0 Å, and data were collected to 2.9 Å resolution. Analysis of the native Patterson map revealed a peak at 50% of the origin on the Harker section v = 0.5, suggesting twofold non-crystallographic symmetry parallel to the b crystallographic axis. As attempts to derivatize or grow selenomethionine-labelled protein crystals failed, in-drop proteolysis was used to produce new crystal forms. A trace amount of subtilisin Carlsberg was added to IpaD before sparse-matrix screening, resulting in the production of several new crystal forms. This approach produced SeMet-labelled crystals and diffraction data were collected to 3.2 Å resolution. The SeMet crystals belong to space group C2, with unit-cell parameters a = 139.4, b = 45.0, c = 99.5 Å, β = 107.9°. An anomalous difference Patterson map revealed peaks on the Harker section v = 0, while the self-rotation function indicates the presence of a twofold noncrystallographic symmetry axis, which is consistent with two molecules per asymmetric unit. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Johnson, S. Roversi, P. Espina, M. Deane, J.E. Birket, S. Picking, W.D. Blocker, A. Picking, W.L. Lea, S.M. 2006-09-01 doi:10.1107/S1744309106027047 International Union of Crystallography IpaD, the putative needle-tip protein of the S. flexneri type III secretion system, has been crystallized in a variety of crystal forms using in-drop proteolysis. Native and selenomethionine-labelled data collection and preliminary analyses are reported. en IPAD; TYPE III SECRETION; SHIGELLA FLEXNERI IpaD, the putative needle-tip protein of the Shigella flexneri type III secretion system, has been overexpressed and purified. Crystals were grown of the native protein in space group P212121, with unit-cell parameters a = 55.9, b = 100.7, c = 112.0 Å, and data were collected to 2.9 Å resolution. Analysis of the native Patterson map revealed a peak at 50% of the origin on the Harker section v = 0.5, suggesting twofold non-crystallographic symmetry parallel to the b crystallographic axis. As attempts to derivatize or grow selenomethionine-labelled protein crystals failed, in-drop proteolysis was used to produce new crystal forms. A trace amount of subtilisin Carlsberg was added to IpaD before sparse-matrix screening, resulting in the production of several new crystal forms. This approach produced SeMet-labelled crystals and diffraction data were collected to 3.2 Å resolution. The SeMet crystals belong to space group C2, with unit-cell parameters a = 139.4, b = 45.0, c = 99.5 Å, β = 107.9°. An anomalous difference Patterson map revealed peaks on the Harker section v = 0, while the self-rotation function indicates the presence of a twofold noncrystallographic symmetry axis, which is consistent with two molecules per asymmetric unit. text/html Expression, limited proteolysis and preliminary crystallographic analysis of IpaD, a component of the Shigella flexneri type III secretion system text 9 62 2006-09-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 865 med@iucr.org 868 1744-3091 http://scripts.iucr.org/cgi-bin/paper?fw5094 Burkholderia pseudomallei, the causative agent of melioidosis, possesses a protein-secretion apparatus that is similar to those found in Salmonella and Shigella. A major function of these secretion systems is to secrete virulence-associated proteins into target cells of the host organism. The BipD gene of B. pseudomallei encodes a secreted virulence factor that is similar in sequence and most likely functionally analogous to IpaD from Shigella and SipD from Salmonella. Thus, the BipD protein is likely to be a component of a type III protein-secretion system (TTSS) in B. pseudomallei. Proteins in the same class as BipD, such as IpaD and SipD, are thought to act as extracellular chaperones to help the hydrophobic translocator proteins enter the target cell membrane, where they form a pore and might even link the translocon pore with the secretion needle. There is evidence that the translocator proteins also bind an integrin which stimulates actin-mediated insertion of the bacterium into the host-cell membrane. Native BipD has been crystallized in a monoclinic crystal form that diffracts X-rays to 2.5 Å resolution. BipD protein which incorporates selenomethionine (SeMet-BipD) has also been expressed and forms crystals which diffract to a higher resolution of 2.1 Å. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Knight, M.J. Ruaux, A. Mikolajek, H. Erskine, P.T. Gill, R. Wood, S.P. Wood, M. Cooper, J.B. 2006-08-01 doi:10.1107/S1744309106024857 International Union of Crystallography BipD is likely to be a component of a type-III protein secretion system (TTSS) in B. pseudomallei. Native and selenomethionyl-BipD proteins have been expressed and crystals have been obtained which diffract to 2.1 Å. en BIPD; BURKHOLDERIA PSEUDOMALLEI Burkholderia pseudomallei, the causative agent of melioidosis, possesses a protein-secretion apparatus that is similar to those found in Salmonella and Shigella. A major function of these secretion systems is to secrete virulence-associated proteins into target cells of the host organism. The BipD gene of B. pseudomallei encodes a secreted virulence factor that is similar in sequence and most likely functionally analogous to IpaD from Shigella and SipD from Salmonella. Thus, the BipD protein is likely to be a component of a type III protein-secretion system (TTSS) in B. pseudomallei. Proteins in the same class as BipD, such as IpaD and SipD, are thought to act as extracellular chaperones to help the hydrophobic translocator proteins enter the target cell membrane, where they form a pore and might even link the translocon pore with the secretion needle. There is evidence that the translocator proteins also bind an integrin which stimulates actin-mediated insertion of the bacterium into the host-cell membrane. Native BipD has been crystallized in a monoclinic crystal form that diffracts X-rays to 2.5 Å resolution. BipD protein which incorporates selenomethionine (SeMet-BipD) has also been expressed and forms crystals which diffract to a higher resolution of 2.1 Å. text/html Crystallization and preliminary X-ray diffraction analysis of BipD, a virulence factor from Burkholderia pseudomallei text 8 62 2006-08-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 761 med@iucr.org 764 1744-3091 http://scripts.iucr.org/cgi-bin/paper?sw5008 The crystal structure of Staphylococcus aureus cytidine monophosphate kinase (CMK) in complex with cytidine 5′-monophosphate (CMP) has been determined at 2.3 Å resolution. The active site reveals novel features when compared with two orthologues of known structure. Compared with the Streptococcus pneumoniae CMK solution structure of the enzyme alone, S. aureus CMK adopts a more closed conformation, with the NMP-binding domain rotating by ∼16° towards the central pocket of the molecule, thereby assembling the active site. Comparing Escherichia coli and S. aureus CMK–CMP complex structures reveals differences within the active site, including a previously unreported indirect interaction of CMP with Asp33, the replacement of a serine residue involved in the binding of CDP by Ala12 in S. aureus CMK and an additional sulfate ion in the E. coli CMK active site. The detailed understanding of the stereochemistry of CMP binding to CMK will assist in the design of novel inhibitors of the enzyme. Inhibitors are required to treat the widespread hospital infection methicillin-resistant S. aureus (MRSA), currently a major public health concern. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Dhaliwal, B. Ren, J. Lockyer, M. Charles, I. Hawkins, A.R. Stammers, D.K. 2006-08-01 doi:10.1107/S174430910602447X International Union of Crystallography The crystal structure of S. aureus cytidine monophosphate kinase in complex with cytidine 5′-monophosphate has been determined. en CYTIDINE MONOPHOSPHATE KINASE; STAPHYLOCOCCUS AUREUS; MRSA; CYTIDINE 5'-MONOPHOSPHATE The crystal structure of Staphylococcus aureus cytidine monophosphate kinase (CMK) in complex with cytidine 5′-monophosphate (CMP) has been determined at 2.3 Å resolution. The active site reveals novel features when compared with two orthologues of known structure. Compared with the Streptococcus pneumoniae CMK solution structure of the enzyme alone, S. aureus CMK adopts a more closed conformation, with the NMP-binding domain rotating by ∼16° towards the central pocket of the molecule, thereby assembling the active site. Comparing Escherichia coli and S. aureus CMK–CMP complex structures reveals differences within the active site, including a previously unreported indirect interaction of CMP with Asp33, the replacement of a serine residue involved in the binding of CDP by Ala12 in S. aureus CMK and an additional sulfate ion in the E. coli CMK active site. The detailed understanding of the stereochemistry of CMP binding to CMK will assist in the design of novel inhibitors of the enzyme. Inhibitors are required to treat the widespread hospital infection methicillin-resistant S. aureus (MRSA), currently a major public health concern. text/html Structure of Staphylococcus aureus cytidine monophosphate kinase in complex with cytidine 5′-monophosphate text 8 62 2006-08-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 protein structure communications 710 med@iucr.org 715 1744-3091 http://scripts.iucr.org/cgi-bin/paper?ll5070 DIR1, a putative LTP2 protein from Arabidopsis thaliana implicated in systemic acquired resistance in planta, has been crystallized in space group P212121 with one molecule per asymmetric unit. The crystals diffract to a resolution of 1.6 Å. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Lascombe, M.-B. Buhot, N. Bakan, B. Marion, D. Blein, J.P. Lamb, C.J. Prangé, T. 2006-07-01 doi:10.1107/S1744309106023748 International Union of Crystallography DIR1, a putative LTP2 protein from Arabidopsis thaliana implicated in systemic acquired resistance in planta, has been crystallized in space group P212121 with one molecule per asymmetric unit. en LTP2 PROTEINS; ARABIDOPSIS THALIANA; PLANT SYSTEMIC ACQUIRED RESISTANCE DIR1, a putative LTP2 protein from Arabidopsis thaliana implicated in systemic acquired resistance in planta, has been crystallized in space group P212121 with one molecule per asymmetric unit. The crystals diffract to a resolution of 1.6 Å. text/html Crystallization of DIR1, a LTP2-like resistance signalling protein from Arabidopsis thaliana text 7 62 2006-07-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 702 med@iucr.org 704 1744-3091 http://scripts.iucr.org/cgi-bin/paper?en5183 Streptococcus pneumoniae contains a large number of sugar-transport systems and the system responsible for raffinose uptake has recently been identified. The substrate-binding protein component of this system shares strong sequence homology with the multiple sugar metabolism substrate-binding protein MsmE from S. mutans and contains a lipoprotein-attachment site at cysteine residue 23. A truncated form (residues 24–419) of RafE from S. pneumoniae was cloned and overexpressed in Escherichia coli. Native and selenomethionine-labelled protein have been crystallized in the hexagonal space group P6122. Diffraction data have been successfully phased to 2.90 Å using Se SAD data and model building is in progress. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Paterson, N.G. Riboldi-Tunnicliffe, A. Mitchell, T.J. Isaacs, N.W. 2006-07-01 doi:10.1107/S1744309106021695 International Union of Crystallography The mature form of RafE has been expressed, purified and crystallized. X-ray diffraction data have been collected to 3.65 and 2.90 Å resolution from native and selenomethionine-derivative crystals, respectively. en SP1897; RAFE; RAFFINOSE; ABC TRANSPORTER; SELENOMETHIONINE; SAD Streptococcus pneumoniae contains a large number of sugar-transport systems and the system responsible for raffinose uptake has recently been identified. The substrate-binding protein component of this system shares strong sequence homology with the multiple sugar metabolism substrate-binding protein MsmE from S. mutans and contains a lipoprotein-attachment site at cysteine residue 23. A truncated form (residues 24–419) of RafE from S. pneumoniae was cloned and overexpressed in Escherichia coli. Native and selenomethionine-labelled protein have been crystallized in the hexagonal space group P6122. Diffraction data have been successfully phased to 2.90 Å using Se SAD data and model building is in progress. text/html Purification, crystallization and preliminary X-ray diffraction analysis of RafE, a sugar-binding lipoprotein from Streptococcus pneumoniae text 7 62 2006-07-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 676 med@iucr.org 679 1744-3091 http://scripts.iucr.org/cgi-bin/paper?hv5056 Human carbonic anhydrases (CAs) are well studied targets for the development of inhibitors for pharmaceutical applications. The crystal structure of human CA II has been determined in complex with two CA inhibitors (CAIs) containing conventional sulfonamide and thiadiazole moieties separated by a —CF2— or —­CHNH2— spacer group. The structures presented here reveal that these spacer groups allow novel binding modes for the thiadiazole moiety compared with conventional CAIs. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Fisher, S.Z. Govindasamy, L. Boyle, N. Agbandje-McKenna, M. Silverman, D.N. Blackburn, G.M. McKenna, R. 2006-07-01 doi:10.1107/S1744309106020446 International Union of Crystallography The crystal structure of human CA II has been determined in complex with two CA inhibitors (CAIs) containing conventional sulfonamide and thiadiazole moieties separated by a —CF2— or —­CHNH2— spacer group. en CARBONIC ANHYDRASE INHIBITORS; FLUORINE HYDROGEN BOND; SULFONAMIDE Human carbonic anhydrases (CAs) are well studied targets for the development of inhibitors for pharmaceutical applications. The crystal structure of human CA II has been determined in complex with two CA inhibitors (CAIs) containing conventional sulfonamide and thiadiazole moieties separated by a —CF2— or —­CHNH2— spacer group. The structures presented here reveal that these spacer groups allow novel binding modes for the thiadiazole moiety compared with conventional CAIs. text/html X-ray crystallographic studies reveal that the incorporation of spacer groups in carbonic anhydrase inhibitors causes alternate binding modes text 7 62 2006-07-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 protein structure communications 618 med@iucr.org 622 1744-3091 http://scripts.iucr.org/cgi-bin/paper?fw5083 The protozoan parasite Toxoplasma gondii is the causative agent of one of the most widespread parasitic infections of man and is a leading cause of congenital neurological birth defects and the third most common cause of food-borne deaths in the United States. Despite this, to date no drugs are available that provide a fully effective treatment. Recently, the antibacterial agent triclosan was shown to inhibit the fatty-acid biosynthesis pathway in T. gondii and to interact with the enoyl reductase (ENR). In order to analyse the potential of triclosan as a lead compound targeting T. gondii ENR and to explore unique features of the apicomplexan enzyme that could be exploited in future drug development, structural studies have been initiated on T. gondii ENR. Crystals of T. gondii ENR in complex with NAD+ and triclosan were grown using the hanging-drop vapour-diffusion method with PEG 8000 as precipitant. The crystals belong to space group P3221, with approximate unit-cell parameters a = 78.1, b = 78.1, c = 188.5 Å, α = β = 90, γ = 120° and a dimer in the asymmetric unit. Test data were collected to beyond 2.6 Å on cryocooled crystals (100 K) using a Rigaku MM007 rotating-anode X-ray source, revealing that the crystals are suitable for a full structural determination. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Muench, S.P. Prigge, S.T. Zhu, L. Kirisits, M.J. Roberts, C.W. Wernimont, S. McLeod, R. Rice, D.W. 2006-06-01 doi:10.1107/S1744309106018112 International Union of Crystallography Crystals of T. gondii ENR in complex with NAD+ and triclosan were grown using the hanging-drop vapour-diffusion method with PEG 8000 as precipitant. en ENOYL REDUCTASE; TRICLOSAN; T. GONDII The protozoan parasite Toxoplasma gondii is the causative agent of one of the most widespread parasitic infections of man and is a leading cause of congenital neurological birth defects and the third most common cause of food-borne deaths in the United States. Despite this, to date no drugs are available that provide a fully effective treatment. Recently, the antibacterial agent triclosan was shown to inhibit the fatty-acid biosynthesis pathway in T. gondii and to interact with the enoyl reductase (ENR). In order to analyse the potential of triclosan as a lead compound targeting T. gondii ENR and to explore unique features of the apicomplexan enzyme that could be exploited in future drug development, structural studies have been initiated on T. gondii ENR. Crystals of T. gondii ENR in complex with NAD+ and triclosan were grown using the hanging-drop vapour-diffusion method with PEG 8000 as precipitant. The crystals belong to space group P3221, with approximate unit-cell parameters a = 78.1, b = 78.1, c = 188.5 Å, α = β = 90, γ = 120° and a dimer in the asymmetric unit. Test data were collected to beyond 2.6 Å on cryocooled crystals (100 K) using a Rigaku MM007 rotating-anode X-ray source, revealing that the crystals are suitable for a full structural determination. text/html Expression, purification and preliminary crystallographic analysis of the Toxoplasma gondii enoyl reductase text 6 62 2006-06-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 604 med@iucr.org 606 1744-3091 http://scripts.iucr.org/cgi-bin/paper?en5174 The choline-binding protein CbpI from Streptococcus pneumoniae is a 23.4 kDa protein with no known function. The protein has been successfully purified initially using Ni–NTA chromatography and to homogeneity using Q-Sepharose ion-exchange resin as an affinity column. CbpI was crystallized using PEG 3350 as a precipitant and X-ray crystallographic analysis showed that the crystals belonged to the tetragonal space group P4, with unit-cell parameters a = b = 83.31, c = 80.29 Å, α = β = γ = 90°. The crystal contains two molecules in the asymmetric unit with a solvent content of 55.7% (VM = 2.77 Å3 Da−1) and shows a diffraction limit of 3.5 Å. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Paterson, N.G. Riboldi-Tunicliffe, A. Mitchell, T.J. Isaacs, N.W. 2006-07-01 doi:10.1107/S1744309106020616 International Union of Crystallography The choline-binding protein CbpI from S. pneumoniae has been purified and crystallized and diffraction data have been collected to 3.5 Å resolution. en SP0069; CBPI; CHOLINE-BINDING PROTEINS; STREPTOCOCCUS PNEUMONIAE The choline-binding protein CbpI from Streptococcus pneumoniae is a 23.4 kDa protein with no known function. The protein has been successfully purified initially using Ni–NTA chromatography and to homogeneity using Q-Sepharose ion-exchange resin as an affinity column. CbpI was crystallized using PEG 3350 as a precipitant and X-ray crystallographic analysis showed that the crystals belonged to the tetragonal space group P4, with unit-cell parameters a = b = 83.31, c = 80.29 Å, α = β = γ = 90°. The crystal contains two molecules in the asymmetric unit with a solvent content of 55.7% (VM = 2.77 Å3 Da−1) and shows a diffraction limit of 3.5 Å. text/html Overexpression, purification and crystallization of a choline-binding protein CbpI from Streptococcus pneumoniae text 7 62 2006-07-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 672 med@iucr.org 675 1744-3091 http://scripts.iucr.org/cgi-bin/paper?sw5006 The PII signal transduction proteins GlnB and GlnK are implicated in the regulation of nitrogen assimilation in Escherichia coli and other enteric bacteria. PII-like proteins are widely distributed in bacteria, archaea and plants. In contrast to other bacteria, Neisseria are limited to a single PII protein (NMB 1995), which shows a high level of sequence identity to GlnB and GlnK from Escherichia coli (73 and 62%, respectively). The structure of the PII protein from N. meningitidis (serotype B) has been solved by molecular replacement to a resolution of 1.85 Å. Comparison of the structure with those of other PII proteins shows that the overall fold is tightly conserved across the whole population of related proteins, in particular the positions of the residues implicated in ATP binding. It is proposed that the Neisseria PII protein shares functions with GlnB/GlnK of enteric bacteria. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Nichols, C.E. Sainsbury, S. Berrow, N.S. Alderton, D. Saunders, N.J. Stammers, D.K. Owens, R.J. 2006-06-01 doi:10.1107/S1744309106015430 International Union of Crystallography The structure of the PII signal transduction protein of N. meningitidis at 1.85 Å resolution is described. en PII SIGNAL TRANSDUCTION PROTEINS; NEISSERIAMENINGITIDIS The PII signal transduction proteins GlnB and GlnK are implicated in the regulation of nitrogen assimilation in Escherichia coli and other enteric bacteria. PII-like proteins are widely distributed in bacteria, archaea and plants. In contrast to other bacteria, Neisseria are limited to a single PII protein (NMB 1995), which shows a high level of sequence identity to GlnB and GlnK from Escherichia coli (73 and 62%, respectively). The structure of the PII protein from N. meningitidis (serotype B) has been solved by molecular replacement to a resolution of 1.85 Å. Comparison of the structure with those of other PII proteins shows that the overall fold is tightly conserved across the whole population of related proteins, in particular the positions of the residues implicated in ATP binding. It is proposed that the Neisseria PII protein shares functions with GlnB/GlnK of enteric bacteria. text/html Structure of the PII signal transduction protein of Neisseria meningitidis at 1.85 Å resolution text 6 62 2006-06-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 structural genomics communications 494 med@iucr.org 497 1744-3091 http://scripts.iucr.org/cgi-bin/paper?sw5009 Attempts to cocrystallize the cysteine protease papain derived from the latex of Carica papaya with an inhibitor of cysteine proteases (ICP) from Trypanosoma brucei were unsuccessful. However, crystals of papain that diffracted to higher resolution, 1.5 Å, than other crystals of this archetypal cysteine protease were obtained, so the analysis was continued. Surprisingly, the substrate-binding cleft was occupied by two short peptide fragments which have been assigned as remnants of ICP. Comparisons reveal that these peptides bind in the active site in a manner similar to that of the human cysteine protease inhibitor stefin B when it is complexed to papain. The assignment of the fragment sequences is consistent with the specificity of the protease. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Alphey, M.S. Hunter, W.N. 2006-06-01 doi:10.1107/S1744309106014849 International Union of Crystallography Attempts to crystallize a complex of papain (C. papaya) with a cysteine protease inhibitor from the parasitic pathogen T. brucei failed. However, over an extended period the mixture produced an ordered crystal of the protease carrying two peptide fragments in the active site. These correspond to dipeptides and tripeptides that are assigned as fragments of the inhibitor, which has presumably suffered proteolytic cleavage. en PAPAIN; CYSTEINE PROTEASE; INHIBITORS; TRYPANOSOMA BRUCEI Attempts to cocrystallize the cysteine protease papain derived from the latex of Carica papaya with an inhibitor of cysteine proteases (ICP) from Trypanosoma brucei were unsuccessful. However, crystals of papain that diffracted to higher resolution, 1.5 Å, than other crystals of this archetypal cysteine protease were obtained, so the analysis was continued. Surprisingly, the substrate-binding cleft was occupied by two short peptide fragments which have been assigned as remnants of ICP. Comparisons reveal that these peptides bind in the active site in a manner similar to that of the human cysteine protease inhibitor stefin B when it is complexed to papain. The assignment of the fragment sequences is consistent with the specificity of the protease. text/html High-resolution complex of papain with remnants of a cysteine protease inhibitor derived from Trypanosoma brucei text 6 62 2006-06-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 protein structure communications 504 med@iucr.org 508 1744-3091 http://scripts.iucr.org/cgi-bin/paper?pu5136 The soluble quinoprotein glucose dehydrogenase oxidizes glucose, maltose and a variety of other monosaccharides and disaccharides to the corresponding lactones. An efficient microseeding protocol is reported to produce crystals of three variants that display reduced activity towards maltose. Similar cross-seeding protocols to grow crystals of homologues from Escherichia coli and Streptomyces coelicolor are described. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Sanchez-Weatherby, J. Southall, S. Oubrie, A. 2006-06-01 doi:10.1107/S1744309106014862 International Union of Crystallography Three mutants of the soluble quinoprotein glucose dehydrogenase and two homologues from E. coli and S. coelicolor have been crystallized using an efficient microseeding method. en CROSS-SEEDING; GLUCOSE DEHYDROGENASE; MICROSEEDING; PQQ; QUINOPROTEIN The soluble quinoprotein glucose dehydrogenase oxidizes glucose, maltose and a variety of other monosaccharides and disaccharides to the corresponding lactones. An efficient microseeding protocol is reported to produce crystals of three variants that display reduced activity towards maltose. Similar cross-seeding protocols to grow crystals of homologues from Escherichia coli and Streptomyces coelicolor are described. text/html Crystallization of quinoprotein glucose dehydrogenase variants and homologues by microseeding text 6 62 2006-06-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 518 med@iucr.org 521 1744-3091 http://scripts.iucr.org/cgi-bin/paper?pu5123 Chorismate mutase catalyzes the conversion of chorismate to prephenate in the biosynthesis of the aromatic amino acids tyrosine and phenylalanine in bacteria, fungi and plants. Here, the crystallization of the unusual secreted chorismate mutase from Mycobacterium tuberculosis (encoded by Rv1885c), a 37.2 kDa dimeric protein belonging to the AroQγ subclass of mutases, is reported. Crystal optimization was non-trivial and is discussed in detail. To obtain crystals of sufficient quality, it was critical to initiate crystallization at higher precipitant concentration and then transfer the drops to lower precipitant concentrations within 5–15 min, in an adaptation of a previously described technique [Saridakis & Chayen (2000), Protein Sci. 9, 755–757]. As a result of the optimization, diffraction improved from 3.5 to 1.3 Å resolution. The crystals belong to space group P21, with unit-cell parameters a = 42.6, b = 72.6, c = 62.0 Å, β = 104.5°. The asymmetric unit contains one biological dimer, with 167 amino acids per protomer. A soak with a transition-state analogue is also described. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Krengel, U. Dey, R. Sasso, S. Ökvist, M. Ramakrishnan, C. Kast, P. 2006-05-01 doi:10.1107/S1744309106012036 International Union of Crystallography A method is presented that allowed the diffraction limit of crystals of the secreted chorismate mutase from M. tuberculosis to be improved from approximately 3.5 to 1.3 Å. To obtain large well diffracting crystals, it was critical to initiate crystallization at higher precipitant concentration and then transfer the drops to lower precipitant concentrations within 5–15 min. en AROQ FOLD; LIGAND COMPLEX; NUCLEATION; PATHOGENIC BACTERIUM; SECRETED CHORISMATE MUTASE; SHIKIMATE PATHWAY; TRANSITION-STATE ANALOGUE; EQUILIBRATION KINETICS; PROTEIN CRYSTALLIZATION Chorismate mutase catalyzes the conversion of chorismate to prephenate in the biosynthesis of the aromatic amino acids tyrosine and phenylalanine in bacteria, fungi and plants. Here, the crystallization of the unusual secreted chorismate mutase from Mycobacterium tuberculosis (encoded by Rv1885c), a 37.2 kDa dimeric protein belonging to the AroQγ subclass of mutases, is reported. Crystal optimization was non-trivial and is discussed in detail. To obtain crystals of sufficient quality, it was critical to initiate crystallization at higher precipitant concentration and then transfer the drops to lower precipitant concentrations within 5–15 min, in an adaptation of a previously described technique [Saridakis & Chayen (2000), Protein Sci. 9, 755–757]. As a result of the optimization, diffraction improved from 3.5 to 1.3 Å resolution. The crystals belong to space group P21, with unit-cell parameters a = 42.6, b = 72.6, c = 62.0 Å, β = 104.5°. The asymmetric unit contains one biological dimer, with 167 amino acids per protomer. A soak with a transition-state analogue is also described. text/html Preliminary X-ray crystallographic analysis of the secreted chorismate mutase from Mycobacterium tuberculosis: a tricky crystallization problem solved text 5 62 2006-05-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 441 med@iucr.org 445 1744-3091 http://scripts.iucr.org/cgi-bin/paper?en5169 The Bacillus subtilis YphC gene encodes an essential GTPase thought to be involved in ribosome binding and whose protein product may represent a target for the development of a novel antibacterial agent. Sequence analysis reveals that YphC belongs to the EngA family of GTPases, which uniquely contain two adjacent GTP-binding domains. Crystals of a selenomethionine-incorporated YphC–GDP complex have been grown using the hanging-drop vapour-diffusion method and polyethylene glycol as a precipitating agent. The crystals belong to space group P212121, with unit-cell parameters a = 62.71, b = 65.05, c = 110.61 Å, and have one molecule in the asymmetric unit. Data sets at three different wavelengths were collected on a single crystal to 2.5 Å resolution at the Daresbury SRS in order to solve the structure by MAD. Ultimately, analysis of YphC in complex with GDP may allow a greater understanding of the EngA family of essential GTPases. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Xu, L. Muench, S.P. Roujeinikova, A. Sedelnikova, S.E. Rice, D.W. 2006-05-01 doi:10.1107/S1744309106011456 International Union of Crystallography Crystals of a selenomethionine-incorporated YphC–GDP complex have been grown using the hanging-drop vapour-diffusion method and polyethylene glycol as a precipitating agent. en GTPASE; ENGA; YPHC The Bacillus subtilis YphC gene encodes an essential GTPase thought to be involved in ribosome binding and whose protein product may represent a target for the development of a novel antibacterial agent. Sequence analysis reveals that YphC belongs to the EngA family of GTPases, which uniquely contain two adjacent GTP-binding domains. Crystals of a selenomethionine-incorporated YphC–GDP complex have been grown using the hanging-drop vapour-diffusion method and polyethylene glycol as a precipitating agent. The crystals belong to space group P212121, with unit-cell parameters a = 62.71, b = 65.05, c = 110.61 Å, and have one molecule in the asymmetric unit. Data sets at three different wavelengths were collected on a single crystal to 2.5 Å resolution at the Daresbury SRS in order to solve the structure by MAD. Ultimately, analysis of YphC in complex with GDP may allow a greater understanding of the EngA family of essential GTPases. text/html Cloning, purification and preliminary crystallographic analysis of the Bacillus subtilis GTPase YphC–GDP complex text 5 62 2006-05-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 435 med@iucr.org 437 1744-3091 http://scripts.iucr.org/cgi-bin/paper?pu5133 N-acetylglucosamine-phosphate mutase (AGM1) is an essential enzyme in the synthesis of UDP-N-acetylglucosamine (UDP-GlcNAc) in eukaryotes and belongs to the α-d-phosphohexomutase superfamily. AGM1 from Candida albicans (CaAGM1) was purified and crystallized by the sitting-drop vapour-diffusion method. The crystals obtained belong to the primitive monoclinic space group P21, with unit-cell parameters a = 60.2, b = 130.2, c = 78.0 Å, β = 106.7°. The crystals diffract X-rays to beyond 1.8 Å resolution using synchrotron radiation. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Nishitani, Y. Maruyama, D. Nonaka, T. Kita, A. Fukami, T.A. Mio, T. Yamada-Okabe, H. Yamada-Okabe, T. Miki, K. 2006-04-01 doi:10.1107/S1744309106010177 International Union of Crystallography Preliminary X-ray diffraction studies on N-acetylglucosamine-phosphate mutase from C. albicans are reported. en N-ACETYLGLUCOSAMINE-PHOSPHATE MUTASE; CANDIDA ALBICANS N-acetylglucosamine-phosphate mutase (AGM1) is an essential enzyme in the synthesis of UDP-N-acetylglucosamine (UDP-GlcNAc) in eukaryotes and belongs to the α-d-phosphohexomutase superfamily. AGM1 from Candida albicans (CaAGM1) was purified and crystallized by the sitting-drop vapour-diffusion method. The crystals obtained belong to the primitive monoclinic space group P21, with unit-cell parameters a = 60.2, b = 130.2, c = 78.0 Å, β = 106.7°. The crystals diffract X-rays to beyond 1.8 Å resolution using synchrotron radiation. text/html Purification, crystallization and preliminary X-ray diffraction studies of N-acetylglucosamine-phosphate mutase from Candida albicans text 4 62 2006-04-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 419 med@iucr.org 421 1744-3091 http://scripts.iucr.org/cgi-bin/paper?tb5003 As a part of a structural genomics program, the 2.2 Å resolution crystal structure of the PurC gene product from Thermotoga maritima has been solved. This 26.2 kDa protein belongs to the phophoribosylaminoimidazole-succinocarboxamide or SAICAR synthase family of enzymes, the members of which are involved in de novo purine biosynthesis. SAICAR synthase can be divided into three subdomains: two α+β regions exhibiting structural homology with ATP-binding proteins and a carboxy-terminal subdomain of two α-helices. The asymmetric unit contains two copies of the protein which are covalently linked by a disulfide bond between Cys126(A) and Cys126(B). This 230-amino-acid protein exhibits high structural homology with SAICAR synthase from baker's yeast. The protein structure is described and compared with that of the ATP–SAICAR synthase complex from yeast. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Zhang, R. Skarina, T. Evdokimova, E. Edwards, A. Savchenko, A. Laskowski, R. Cuff, M.E. Joachimiak, A. 2006-04-01 doi:10.1107/S1744309106009651 International Union of Crystallography The crystal structure of phophoribosylaminoimidazole-succinocarboxamide or SAICAR synthase from T. maritima at 2.2 Å revealed an unusual covalent dimer. en SAICAR SYNTHASE; PURINE BIOSYNTHESIS; THERMOTOGA MARITIMA As a part of a structural genomics program, the 2.2 Å resolution crystal structure of the PurC gene product from Thermotoga maritima has been solved. This 26.2 kDa protein belongs to the phophoribosylaminoimidazole-succinocarboxamide or SAICAR synthase family of enzymes, the members of which are involved in de novo purine biosynthesis. SAICAR synthase can be divided into three subdomains: two α+β regions exhibiting structural homology with ATP-binding proteins and a carboxy-terminal subdomain of two α-helices. The asymmetric unit contains two copies of the protein which are covalently linked by a disulfide bond between Cys126(A) and Cys126(B). This 230-amino-acid protein exhibits high structural homology with SAICAR synthase from baker's yeast. The protein structure is described and compared with that of the ATP–SAICAR synthase complex from yeast. text/html Structure of SAICAR synthase from Thermotoga maritima at 2.2 Å reveals an unusual covalent dimer text 4 62 2006-04-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 structural genomics communications 335 med@iucr.org 339 1744-3091 http://scripts.iucr.org/cgi-bin/paper?ll5051 A monodisperse truncation mutant of MxiH, the subunit of the needle from the Shigella flexneri type III secretion system (TTSS), has been overexpressed and purified. Crystals were grown of native and selenomethionine-labelled MxiHCΔ5 and diffraction data were collected to 1.9 Å resolution. The crystals belong to space group C2, with unit-cell parameters a = 183.4, b = 28.1, c = 27.8 Å, β = 96.5°. An anomalous difference Patterson map calculated with the data from the SeMet-labelled crystals revealed a single peak on the Harker section v = 0. Inspection of a uranyl derivative also revealed one peak in the isomorphous difference Patterson map on the Harker section v = 0. Analysis of the self-rotation function indicates the presence of a twofold non-crystallographic symmetry axis approximately along a. The calculated Matthews coefficient is 1.9 Å3 Da−1 for two molecules per asymmetric unit, corresponding to a solvent content of 33%. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Deane, J.E. Cordes, F.S. Roversi, P. Johnson, S. Kenjale, R. Picking, W.D. Picking, W.L. Lea, S.M. Blocker, A. 2006-03-01 doi:10.1107/S1744309106006555 International Union of Crystallography A monodisperse truncation mutant of MxiH, the subunit of the S. flexneri type III secretion system needle, has been crystallized. SeMet derivatives and a uranyl derivative have undergone preliminary crystallographic analysis. en MXIH; TYPE III SECRETION SYSTEM; SHIGELLA FLEXNERI A monodisperse truncation mutant of MxiH, the subunit of the needle from the Shigella flexneri type III secretion system (TTSS), has been overexpressed and purified. Crystals were grown of native and selenomethionine-labelled MxiHCΔ5 and diffraction data were collected to 1.9 Å resolution. The crystals belong to space group C2, with unit-cell parameters a = 183.4, b = 28.1, c = 27.8 Å, β = 96.5°. An anomalous difference Patterson map calculated with the data from the SeMet-labelled crystals revealed a single peak on the Harker section v = 0. Inspection of a uranyl derivative also revealed one peak in the isomorphous difference Patterson map on the Harker section v = 0. Analysis of the self-rotation function indicates the presence of a twofold non-crystallographic symmetry axis approximately along a. The calculated Matthews coefficient is 1.9 Å3 Da−1 for two molecules per asymmetric unit, corresponding to a solvent content of 33%. text/html Expression, purification, crystallization and preliminary crystallographic analysis of MxiH, a subunit of the Shigella flexneri type III secretion system needle text 3 62 2006-03-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 302 med@iucr.org 305 1744-3091 http://scripts.iucr.org/cgi-bin/paper?ll5046 Sso6206, a 10.5 kDa protein from Sulfolobus solfataricus, has been overexpressed, purified and crystallized. The protein crystallizes in space group P61/522, with unit-cell parameters a = b = 157.8, c = 307.3 Å. The crystals are hexagonal bipyramids and a data set has been collected to 2.4 Å resolution. Molecular replacement cannot be attempted as no convincing model can be identified. Crystals of selenomethionine-variant protein have not yet been obtained. Interestingly, crystal packing, gel filtration and mass spectrometry all suggest the native protein forms a multi-subunit oligomer consisting of >9 subunits. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 McEwan, A.R. Liu, H. Oke, M. Carter, L. Powers, H. Dorward, M. McMahon, S.A. White, M.F. Naismith, J.H. 2006-03-01 doi:10.1107/S1744309106003654 International Union of Crystallography The novel protein Sso6206 has been crystallized; interestingly, the protein may form large multi-subunit oligomers. en SSO6206; SULFOLOBUS SOLFATARICUS Sso6206, a 10.5 kDa protein from Sulfolobus solfataricus, has been overexpressed, purified and crystallized. The protein crystallizes in space group P61/522, with unit-cell parameters a = b = 157.8, c = 307.3 Å. The crystals are hexagonal bipyramids and a data set has been collected to 2.4 Å resolution. Molecular replacement cannot be attempted as no convincing model can be identified. Crystals of selenomethionine-variant protein have not yet been obtained. Interestingly, crystal packing, gel filtration and mass spectrometry all suggest the native protein forms a multi-subunit oligomer consisting of >9 subunits. text/html Overexpression, purification, crystallization and data collection of Sulfolobus solfataricus Sso6206, a novel highly conserved protein text 3 62 2006-03-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 228 med@iucr.org 230 1744-3091 http://scripts.iucr.org/cgi-bin/paper?mp5005 Hsp70s are essential chaperones with roles in a variety of cellular processes and representatives in all kingdoms of life. They are comprised of a nucleotide-binding domain (NBD) and a protein substrate-binding domain (SBD). Structures of isolated NBDs and SBDs have been reported but, until recently, a functionally intact Hsp70 containing both the NBD and SBD has resisted structure determination. Here, it is reported that preparation of diffraction-quality crystals of functionally intact bovine Hsc70 required (i) deletion of part of the protein to reduce oligomerization, (ii) point mutations in the interface between the SBD and NBD and (iii) use of high concentrations of the structure-stabilizing agents glycerol and trimethylamine oxide (TMAO). The introduction of point mutations in interdomain interfaces and the use of the potent structure stabilizer TMAO may be generally useful in crystallization of multidomain proteins that exhibit interdomain motions. Copyright (c) 2006 International Union of Crystallography urn:issn:1744-3091 Jiang, J. Lafer, E.M. Sousa, R. 2006-01-01 doi:10.1107/S1744309105040303 International Union of Crystallography Success in crystallization of a functionally intact Hsp70 chaperone required genetic engineering to minimize polydispersity and modulate interdomain interactions, as well as high concentrations of the potent structure stabilizer TMAO. These approaches may be generally useful in crystallization of conformationally flexible proteins that exhibit interdomain motions. en HSP70; HSC70; CHAPERONES Hsp70s are essential chaperones with roles in a variety of cellular processes and representatives in all kingdoms of life. They are comprised of a nucleotide-binding domain (NBD) and a protein substrate-binding domain (SBD). Structures of isolated NBDs and SBDs have been reported but, until recently, a functionally intact Hsp70 containing both the NBD and SBD has resisted structure determination. Here, it is reported that preparation of diffraction-quality crystals of functionally intact bovine Hsc70 required (i) deletion of part of the protein to reduce oligomerization, (ii) point mutations in the interface between the SBD and NBD and (iii) use of high concentrations of the structure-stabilizing agents glycerol and trimethylamine oxide (TMAO). The introduction of point mutations in interdomain interfaces and the use of the potent structure stabilizer TMAO may be generally useful in crystallization of multidomain proteins that exhibit interdomain motions. text/html Crystallization of a functionally intact Hsc70 chaperone text 1 62 2006-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2006 International Union of Crystallography 1744-3091 crystallization communications 39 med@iucr.org 43 1744-3091 http://scripts.iucr.org/cgi-bin/paper?ll5044 The human C-type lectin-like protein CLEC-2 has recently been shown to be expressed on the surface of platelets and to function as a receptor for the snake-venom protein rhodocytin. The C-type lectin-like domain (CTLD) of CLEC-2 was expressed in Escherichia coli, refolded and purified. Crystals of this recombinant CLEC-2 were grown by sitting-drop vapour diffusion using polyethylene glycol (PEG) 6000 as a precipitant. After optimization, crystals were grown which diffracted to 2.0 Å using in-house radiation (λ = 1.5418 Å). These crystals belonged to the orthorhombic space group P212121, with unit-cell parameters a = 35.407, b = 55.143, c = 56.078 Å. The presence of one molecule per asymmetric unit is consistent with a crystal volume per unit weight (VM) of 1.82 Å3 Da−1 and a solvent content of 32.6%. These results suggest that crystals producing diffraction of this quality will be suitable for the structural determination of human CLEC-2. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Watson, A.A. O'Callaghan, C.A. 2005-12-01 doi:10.1107/S1744309105037991 International Union of Crystallography Recombinant human CLEC-2 was crystallized in the orthorhombic space group P212121 and X-ray diffraction data were collected to 2.0 Å. en CLEC-2; CLEC1B; RHODOCYTIN; AGGRETIN; C-TYPE LECTINS; PLATELETS; THROMBOSIS The human C-type lectin-like protein CLEC-2 has recently been shown to be expressed on the surface of platelets and to function as a receptor for the snake-venom protein rhodocytin. The C-type lectin-like domain (CTLD) of CLEC-2 was expressed in Escherichia coli, refolded and purified. Crystals of this recombinant CLEC-2 were grown by sitting-drop vapour diffusion using polyethylene glycol (PEG) 6000 as a precipitant. After optimization, crystals were grown which diffracted to 2.0 Å using in-house radiation (λ = 1.5418 Å). These crystals belonged to the orthorhombic space group P212121, with unit-cell parameters a = 35.407, b = 55.143, c = 56.078 Å. The presence of one molecule per asymmetric unit is consistent with a crystal volume per unit weight (VM) of 1.82 Å3 Da−1 and a solvent content of 32.6%. These results suggest that crystals producing diffraction of this quality will be suitable for the structural determination of human CLEC-2. text/html Crystallization and X-ray diffraction analysis of human CLEC-2 text 12 61 2005-12-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 1094 med@iucr.org 1096 1744-3091 http://scripts.iucr.org/cgi-bin/paper?ll5034 Acylaminoacyl peptidase (also known as acylamino-acid-releasing enzyme or acylpeptide hydrolase; EC 3.4.19.1) is an unusual member of the prolyl oligopeptidase family catalysing the hydrolysis of an N-acylated peptide to an acylamino acid and a peptide with a free N-terminus. Acylaminoacyl peptidase purified from porcine liver has been crystallized in mother liquor containing 0.1 M Tris–HCl pH 7.0, 10%(w/v) polyethylene glycol 8000, 50 mM MgCl2 and 1%(w/v) CHAPS using the hanging-drop vapour-diffusion technique. A full data set to 3.4 Å resolution was collected at ESRF beamline ID14-4 and space group C222 was assigned, with unit-cell parameters a = 84.8, b = 421.1, c = 212.0 Å and four molecules in the asymmetric unit. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Wright, H. Kiss, A.L. Szeltner, Z. Polgár, L. Fülöp, V. 2005-10-01 doi:10.1107/S1744309105029222 International Union of Crystallography Acylaminoacyl peptidase from porcine liver has been crystallized. Data were collected to 3.4 Å from native crystals and a search for heavy-atom derivatives is in progress. en ACYLAMINOACYL PEPTIDASE; PORCINE LIVER; CROSS-LINKING Acylaminoacyl peptidase (also known as acylamino-acid-releasing enzyme or acylpeptide hydrolase; EC 3.4.19.1) is an unusual member of the prolyl oligopeptidase family catalysing the hydrolysis of an N-acylated peptide to an acylamino acid and a peptide with a free N-terminus. Acylaminoacyl peptidase purified from porcine liver has been crystallized in mother liquor containing 0.1 M Tris–HCl pH 7.0, 10%(w/v) polyethylene glycol 8000, 50 mM MgCl2 and 1%(w/v) CHAPS using the hanging-drop vapour-diffusion technique. A full data set to 3.4 Å resolution was collected at ESRF beamline ID14-4 and space group C222 was assigned, with unit-cell parameters a = 84.8, b = 421.1, c = 212.0 Å and four molecules in the asymmetric unit. text/html Crystallization and preliminary crystallographic analysis of porcine acylaminoacyl peptidase text 10 61 2005-10-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 942 med@iucr.org 944 1744-3091 http://scripts.iucr.org/cgi-bin/paper?pu5101 Crystals of the RADIALIS protein from Antirrhinum majus were grown by vapour diffusion after limited proteolysis. Mass spectrometry indicated that an 8 kDa fragment had been crystallized corresponding to the predicted MYB DNA-binding domain. X-ray data collected at room temperature were consistent with tetragonal symmetry, whereas data collected at 100 K using crystals cryoprotected by supplementing the mother liquor with ethylene glycol conformed to orthorhombic symmetry. It was subsequently shown that crystals soaked in cryoprotectants that were `osmolality-matched' to the mother liquor retained tetragonal symmetry. Using these crystals, X-ray data were collected in-house to a maximum resolution of 2 Å. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Stevenson, C.E.M. Burton, N. Costa, M. Nath, U. Dixon, R.A. Coen, E.S. Lawson, D.M. 2005-10-01 doi:10.1107/S1744309105027168 International Union of Crystallography An 8 kDa proteolytic fragment of the A. majus RADIALIS protein was crystallized and X-ray data were collected to 2 Å resolution. en RADIALIS; LIMITED PROTEOLYSIS; OSMOLALITY Crystals of the RADIALIS protein from Antirrhinum majus were grown by vapour diffusion after limited proteolysis. Mass spectrometry indicated that an 8 kDa fragment had been crystallized corresponding to the predicted MYB DNA-binding domain. X-ray data collected at room temperature were consistent with tetragonal symmetry, whereas data collected at 100 K using crystals cryoprotected by supplementing the mother liquor with ethylene glycol conformed to orthorhombic symmetry. It was subsequently shown that crystals soaked in cryoprotectants that were `osmolality-matched' to the mother liquor retained tetragonal symmetry. Using these crystals, X-ray data were collected in-house to a maximum resolution of 2 Å. text/html Crystallization and preliminary X-ray analysis of the RAD protein from Antirrhinum majus text 10 61 2005-10-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 885 med@iucr.org 888 1744-3091 http://scripts.iucr.org/cgi-bin/paper?fw5045 The neuronal protein calexcitin from the long-finned squid Loligo pealei has been expressed in Escherichia coli and purified to homogeneity. Calexcitin is a 22 kDa calcium-binding protein that becomes up-regulated in invertebrates following Pavlovian conditioning and is likely to be involved in signal transduction events associated with learning and memory. Recombinant squid calexcitin has been crystallized using the hanging-drop vapour-diffusion technique in the orthorhombic space group P212121. The unit-cell parameters of a = 46.6, b = 69.2, c = 134.8 Å suggest that the crystals contain two monomers per asymmetric unit and have a solvent content of 49%. This crystal form diffracts X-rays to at least 1.8 Å resolution and yields data of high quality using synchrotron radiation. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Beaven, G.D.E. Erskine, P.T. Wright, J.N. Mohammed, F. Gill, R. Wood, S.P. Vernon, J. Giese, K.P. Cooper, J.B. 2005-10-01 doi:10.1107/S1744309105026758 International Union of Crystallography Recombinant squid calexcitin has been crystallized using the hanging-drop vapour-diffusion technique in the orthorhombic space group P212121. en CALEXCITIN; CALCIUM-BINDING PROTEINS The neuronal protein calexcitin from the long-finned squid Loligo pealei has been expressed in Escherichia coli and purified to homogeneity. Calexcitin is a 22 kDa calcium-binding protein that becomes up-regulated in invertebrates following Pavlovian conditioning and is likely to be involved in signal transduction events associated with learning and memory. Recombinant squid calexcitin has been crystallized using the hanging-drop vapour-diffusion technique in the orthorhombic space group P212121. The unit-cell parameters of a = 46.6, b = 69.2, c = 134.8 Å suggest that the crystals contain two monomers per asymmetric unit and have a solvent content of 49%. This crystal form diffracts X-rays to at least 1.8 Å resolution and yields data of high quality using synchrotron radiation. text/html Crystallization and preliminary X-ray diffraction analysis of calexcitin from Loligo pealei: a neuronal protein implicated in learning and memory text 10 61 2005-10-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 879 med@iucr.org 881 1744-3091 http://scripts.iucr.org/cgi-bin/paper?za5112 Bacteria have evolved strategies to acquire iron from their environment. Pathogenic microbes rely on specialized proteins to `steal' haem from their host and use it as an iron source. HemS is the ultimate recipient of a molecular-relay system for haem uptake in Gram-negative species, functioning as the cytosolic carrier of haem. Soluble expression and high-quality diffraction crystals were obtained for HemS from Yersinia enterocolitica. Crystals belong to the orthorhombic space group I222, with unit-cell parameters a = 74.86, b = 77.45, c = 114.09 Å, and diffract X-rays to 2.6 Å spacing in-house. Determination of the structure of the haem–HemS complex will reveal the molecular basis of haem binding. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Schneider, S. Paoli, M. 2005-08-01 doi:10.1107/S1744309105023523 International Union of Crystallography The haem binding protein HemS from Y. enterocolitica has been crystallized in complex with its ligand. The crystals diffracted X-rays to 2.6 Å in-house. en HAEM PROTEINS; HAEM UPTAKE; LIGAND BINDING Bacteria have evolved strategies to acquire iron from their environment. Pathogenic microbes rely on specialized proteins to `steal' haem from their host and use it as an iron source. HemS is the ultimate recipient of a molecular-relay system for haem uptake in Gram-negative species, functioning as the cytosolic carrier of haem. Soluble expression and high-quality diffraction crystals were obtained for HemS from Yersinia enterocolitica. Crystals belong to the orthorhombic space group I222, with unit-cell parameters a = 74.86, b = 77.45, c = 114.09 Å, and diffract X-rays to 2.6 Å spacing in-house. Determination of the structure of the haem–HemS complex will reveal the molecular basis of haem binding. text/html Crystallization and preliminary X-ray diffraction analysis of the haem-binding protein HemS from Yersinia enterocolitica text 8 61 2005-08-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 802 med@iucr.org 805 1744-3091 http://scripts.iucr.org/cgi-bin/paper?bw5101 The gene encoding 2,4-dihydroxy-hepta-2-ene-1,7-dioate (HHED) aldolase (HpcH; EC 4.1.2) from Escherichia coli C was cloned into the high-expression plasmid pProEx-HTa and overexpressed in E. coli BL21 (DE3). The 28 kDa enzyme was purified using immobilized metal-affinity and size-exclusion chromatography prior to crystallization. Crystals were obtained by the hanging-drop vapour-diffusion method at 277 K from a number of screening conditions. Type I crystals grown in a solution containing 0.4 M ammonium dihydrogen phosphate belong to space group R32, with unit-cell parameters a = b = 128.92, c = 175.30 Å. Type II crystals grown in a solution containing 0.5 M sodium chloride, 0.1 M sodium citrate pH 5.5 belong to space group I222, with unit-cell parameters a = 133.39, b = 155.39, c = 168.80 Å. Complete data sets were collected to 1.6 and 2.0 Å from type I and type II crystals, respectively, using synchrotron radiation. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Rea, D. Fülöp, V. Bugg, T.D.H. Roper, D.I. 2005-09-01 doi:10.1107/S1744309105023079 International Union of Crystallography 2,4-Dihydroxy-hepta-2-ene-1,7-dioate aldolase from E. coli C has been purified and crystallized. Diffraction data were collected to 1.6 Å and structure determination by molecular replacement is in progress. en ALDOLASE; ESCHERICHIA COLI; AROMATIC DEGRADATIVE PATHWAY; 4-HYDROXYPHENYLACETATE The gene encoding 2,4-dihydroxy-hepta-2-ene-1,7-dioate (HHED) aldolase (HpcH; EC 4.1.2) from Escherichia coli C was cloned into the high-expression plasmid pProEx-HTa and overexpressed in E. coli BL21 (DE3). The 28 kDa enzyme was purified using immobilized metal-affinity and size-exclusion chromatography prior to crystallization. Crystals were obtained by the hanging-drop vapour-diffusion method at 277 K from a number of screening conditions. Type I crystals grown in a solution containing 0.4 M ammonium dihydrogen phosphate belong to space group R32, with unit-cell parameters a = b = 128.92, c = 175.30 Å. Type II crystals grown in a solution containing 0.5 M sodium chloride, 0.1 M sodium citrate pH 5.5 belong to space group I222, with unit-cell parameters a = 133.39, b = 155.39, c = 168.80 Å. Complete data sets were collected to 1.6 and 2.0 Å from type I and type II crystals, respectively, using synchrotron radiation. text/html Expression, purification and preliminary crystallographic analysis of 2,4-dihydroxy-hepta-2-ene-1,7-dioate aldolase (HpcH) from Escherichia coli C text 9 61 2005-09-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 821 med@iucr.org 824 1744-3091 http://scripts.iucr.org/cgi-bin/paper?za5106 Imidazoleglycerol-phosphate dehydratase catalyses the sixth step of the histidine-biosynthesis pathway in plants and microorganisms and has been identified as a possible target for the development of novel herbicides. Arabidopsis thaliana IGPD has been cloned and overexpressed in Escherichia coli, purified and subsequently crystallized in the presence of manganese. Under these conditions, the inactive trimeric form of the metal-free enzyme is assembled into a fully active species consisting of a 24-mer exhibiting 432 symmetry. X-ray diffraction data have been collected to 3.0 Å resolution from a single crystal at 293 K. The crystal belongs to space group R3, with approximate unit-cell parameters a = b = 157.9, c = 480.0 Å, α = β = 90, γ = 120° and with either 16 or 24 subunits in the asymmetric unit. A full structure determination is under way in order to provide insights into the mode of subunit assembly and to initiate a programme of rational herbicide design. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Glynn, S.E. Baker, P.J. Sedelnikova, S.E. Levy, C.W. Rodgers, H.F. Blank, J. Hawkes, T.R. Rice, D.W. 2005-08-01 doi:10.1107/S1744309105022451 International Union of Crystallography Imidazoleglycerol-phosphate dehydratase from A. thaliana has been overexpressed, purified and crystallized and data have been collected to 3 Å resolution. en IGPD; HISTIDINE BIOSYNTHESIS; MANGANESE; HERBICIDES Imidazoleglycerol-phosphate dehydratase catalyses the sixth step of the histidine-biosynthesis pathway in plants and microorganisms and has been identified as a possible target for the development of novel herbicides. Arabidopsis thaliana IGPD has been cloned and overexpressed in Escherichia coli, purified and subsequently crystallized in the presence of manganese. Under these conditions, the inactive trimeric form of the metal-free enzyme is assembled into a fully active species consisting of a 24-mer exhibiting 432 symmetry. X-ray diffraction data have been collected to 3.0 Å resolution from a single crystal at 293 K. The crystal belongs to space group R3, with approximate unit-cell parameters a = b = 157.9, c = 480.0 Å, α = β = 90, γ = 120° and with either 16 or 24 subunits in the asymmetric unit. A full structure determination is under way in order to provide insights into the mode of subunit assembly and to initiate a programme of rational herbicide design. text/html Purification, crystallization and preliminary crystallographic analysis of Arabidopsis thaliana imidazoleglycerol-phosphate dehydratase text 8 61 2005-08-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 776 med@iucr.org 778 1744-3091 http://scripts.iucr.org/cgi-bin/paper?za5102 Crystals of the complex formed between the outer membrane protein OmpC from Escherichia coli and the eukaryotic antibacterial protein lactoferrin from Camelus dromedarius (camel) have been obtained using a detergent environment. Initial data processing suggests that the crystals belong to the hexagonal space group P6, with unit-cell parameters a = b = 116.3, c = 152.4 Å, α = β = 90, γ = 120°. This indicated a Matthews coefficient (VM) of 3.3 Å3 Da−1, corresponding to a possible molecular complex involving four molecules of lactoferrin and two porin trimers in the unit cell (4832 amino acids; 533.8 kDa) with 63% solvent content. A complete set of diffraction data was collected to 3 Å resolution at 100 K. Structure determination by molecular replacement is in progress. Structural study of this first surface-exposed membrane-protein complex with an antibacterial protein will provide insights into the mechanism of action of OmpC as well as lactoferrin. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Sundara Baalaji, N. Acharya, K.R. Singh, T.P. Krishnaswamy, S. 2005-08-01 doi:10.1107/S1744309105022086 International Union of Crystallography Crystals of the complex formed between the bacterial membrane protein OmpC and the antibacterial protein lactoferrin suitable for high-resolution structure determination have been obtained. The crystals belong to the hexagonal space group P6, with unit-cell parameters a = b = 116.3, c = 152.4 Å. en OMPC; LACTOFERRIN; DETERGENTS; ANTIBACTERIAL PROTEINS; MEMBRANE-PROTEIN COMPLEXES Crystals of the complex formed between the outer membrane protein OmpC from Escherichia coli and the eukaryotic antibacterial protein lactoferrin from Camelus dromedarius (camel) have been obtained using a detergent environment. Initial data processing suggests that the crystals belong to the hexagonal space group P6, with unit-cell parameters a = b = 116.3, c = 152.4 Å, α = β = 90, γ = 120°. This indicated a Matthews coefficient (VM) of 3.3 Å3 Da−1, corresponding to a possible molecular complex involving four molecules of lactoferrin and two porin trimers in the unit cell (4832 amino acids; 533.8 kDa) with 63% solvent content. A complete set of diffraction data was collected to 3 Å resolution at 100 K. Structure determination by molecular replacement is in progress. Structural study of this first surface-exposed membrane-protein complex with an antibacterial protein will provide insights into the mechanism of action of OmpC as well as lactoferrin. text/html High-resolution diffraction from crystals of a membrane-protein complex: bacterial outer membrane protein OmpC complexed with the antibacterial eukaryotic protein lactoferrin text 8 61 2005-08-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 773 med@iucr.org 775 1744-3091 http://scripts.iucr.org/cgi-bin/paper?fw5041 Glyoxalase I (GLO1) is a putative drug target for trypanosomatids, which are pathogenic protozoa that include the causative agents of leishmaniasis. Significant sequence and functional differences between Leishmania major and human GLO1 suggest that it may make a suitable template for rational inhibitor design. L. major GLO1 was crystallized in two forms: the first is extremely disordered and does not diffract, while the second, an orthorhombic form, produces diffraction to 2.0 Å. Molecular-replacement calculations indicate that there are three GLO1 dimers in the asymmetric unit, which take up a helical arrangement with their molecular dyads arranged approximately perpendicular to the c axis. Further analysis of these data are under way. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Ariza, A. Vickers, T.J. Greig, N. Fairlamb, A.H. Bond, C.S. 2005-08-01 doi:10.1107/S174430910502169X International Union of Crystallography The detoxification enzyme glyoxalase I from L. major has been crystallized. Preliminary molecular-replacement calculations indicate the presence of three glyoxalase I dimers in the asymmetric unit. en GLYOXALASE I; LEISHMANIASIS Glyoxalase I (GLO1) is a putative drug target for trypanosomatids, which are pathogenic protozoa that include the causative agents of leishmaniasis. Significant sequence and functional differences between Leishmania major and human GLO1 suggest that it may make a suitable template for rational inhibitor design. L. major GLO1 was crystallized in two forms: the first is extremely disordered and does not diffract, while the second, an orthorhombic form, produces diffraction to 2.0 Å. Molecular-replacement calculations indicate that there are three GLO1 dimers in the asymmetric unit, which take up a helical arrangement with their molecular dyads arranged approximately perpendicular to the c axis. Further analysis of these data are under way. text/html Crystallization and preliminary X-ray analysis of Leishmania major glyoxalase I text 8 61 2005-08-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 769 med@iucr.org 772 1744-3091 http://scripts.iucr.org/cgi-bin/paper?en5116 PM2 (Corticoviridae) is a dsDNA bacteriophage which contains a lipid membrane beneath its icosahedral capsid. In this respect it resembles bacteriophage PRD1 (Tectiviridae), although it is not known whether the similarity extends to the detailed molecular architecture of the virus, for instance the fold of the major coat protein P2. Structural analysis of PM2 has been initiated and virus-derived P2 has been crystallized by sitting-nanodrop vapour diffusion. Crystals of P2 have been obtained in space group P21212, with two trimers in the asymmetric unit and unit-cell parameters a = 171.1, b = 78.7, c = 130.1 Å. The crystals diffract to 4 Å resolution at the ESRF BM14 beamline (Grenoble, France) and the orientation of the non-crystallographic threefold axes, the spatial relationship between the two trimers and the packing of the trimers within the unit cell have been determined. The trimers form tightly packed layers consistent with the crystal morphology, possibly recapitulating aspects of the arrangement of subunits in the virus. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Abrescia, N.G.A. Kivelä, H.M. Grimes, J.M. Bamford, J.K.H. Bamford, D.H. Stuart, D.I. 2005-08-01 doi:10.1107/S174430910502141X International Union of Crystallography The viral capsid protein P2 of bacteriophage PM2 has been crystallized. Preliminary X-ray analysis demonstrates the position and orientation of the two trimers in the asymmetric unit. en VIRUS CRYSTALLOGRAPHY; LIPID-CONTAINING BACTERIOPHAGES; PRD1-ADENOVIRAL LINEAGE PM2 (Corticoviridae) is a dsDNA bacteriophage which contains a lipid membrane beneath its icosahedral capsid. In this respect it resembles bacteriophage PRD1 (Tectiviridae), although it is not known whether the similarity extends to the detailed molecular architecture of the virus, for instance the fold of the major coat protein P2. Structural analysis of PM2 has been initiated and virus-derived P2 has been crystallized by sitting-nanodrop vapour diffusion. Crystals of P2 have been obtained in space group P21212, with two trimers in the asymmetric unit and unit-cell parameters a = 171.1, b = 78.7, c = 130.1 Å. The crystals diffract to 4 Å resolution at the ESRF BM14 beamline (Grenoble, France) and the orientation of the non-crystallographic threefold axes, the spatial relationship between the two trimers and the packing of the trimers within the unit cell have been determined. The trimers form tightly packed layers consistent with the crystal morphology, possibly recapitulating aspects of the arrangement of subunits in the virus. text/html Preliminary crystallographic analysis of the major capsid protein P2 of the lipid-containing bacteriophage PM2 text 8 61 2005-08-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 762 med@iucr.org 765 1744-3091 http://scripts.iucr.org/cgi-bin/paper?vr5037 A flavoprotein amine dehydrogenase/oxidase with subunit molecular weights of 54.8 kDa (α-subunit) and 42.4 kDa (β-subunit) and specificity for l-proline was cloned from the genomic DNA of the hyperthermophilic marine archaeon Pyrococcus furiosus DSM 3638. The enzyme was overexpressed in Escherichia coli and purified to homogeneity. The enzyme was crystallized using the sitting-drop vapour-diffusion technique. Diffraction data from two different crystal forms were collected to 3.3 and 3.6 Å, respectively, using synchrotron radiation. Both crystals belonged to space group P1, with unit-cell parameters a = 91.3, b = 136.3, c = 203.8 Å, α = 94.5, β = 99.4, γ = 102.7° and a = 93.7, b = 116.3, c = 126.9 Å, α = 97.3, β = 99.9, γ = 104.6°. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Monaghan, P.J. Leys, D. Scrutton, N.S. 2005-08-01 doi:10.1107/S1744309105020737 International Union of Crystallography This report describes the crystallization of a recombinant flavoprotein amine dehydrogenase/oxidase with specificity for l-proline from the hyperthermophile P. furiosus DSM 3638 and X-ray diffraction data collection. Crystals belonged to space group P1 and diffracted to a resolution of 3.3 Å. en PYROCOCCUS FURIOSUS DSM 3638; FLAVOPROTEIN AMINE DEHYDROGENASE; FLAVIN ADENINE DINUCLEOTIDE A flavoprotein amine dehydrogenase/oxidase with subunit molecular weights of 54.8 kDa (α-subunit) and 42.4 kDa (β-subunit) and specificity for l-proline was cloned from the genomic DNA of the hyperthermophilic marine archaeon Pyrococcus furiosus DSM 3638. The enzyme was overexpressed in Escherichia coli and purified to homogeneity. The enzyme was crystallized using the sitting-drop vapour-diffusion technique. Diffraction data from two different crystal forms were collected to 3.3 and 3.6 Å, respectively, using synchrotron radiation. Both crystals belonged to space group P1, with unit-cell parameters a = 91.3, b = 136.3, c = 203.8 Å, α = 94.5, β = 99.4, γ = 102.7° and a = 93.7, b = 116.3, c = 126.9 Å, α = 97.3, β = 99.9, γ = 104.6°. text/html Crystallization and preliminary X-ray diffraction analysis of a flavoenzyme amine dehydrogenase/oxidase from Pyrococcus furiosus DSM 3638 text 8 61 2005-08-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 756 med@iucr.org 758 1744-3091 http://scripts.iucr.org/cgi-bin/paper?za5107 Haloferax mediterranei glucose dehydrogenase (EC 1.1.1.47) belongs to the medium-chain alcohol dehydrogenase superfamily and requires zinc for catalysis. In the majority of these family members, the catalytic zinc is tetrahedrally coordinated by the side chains of a cysteine, a histidine, a cysteine or glutamate and a water molecule. In H. mediterranei glucose dehydrogenase, sequence analysis indicates that the zinc coordination is different, with the invariant cysteine replaced by an aspartate residue. In order to analyse the significance of this replacement and to contribute to an understanding of the role of the metal ion in catalysis, a range of binary and ternary complexes of the wild-type and a D38C mutant protein have been crystallized. For most of the complexes, crystals belonging to space group I222 were obtained using sodium/potassium citrate as a precipitant. However, for the binary and non-productive ternary complexes with NADPH/Zn, it was necessary to replace the citrate with 2-methyl-2,4-pentanediol. Despite the radical change in conditions, the crystals thus formed were isomorphous. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Esclapez, J. Britton, K.L. Baker, P.J. Fisher, M. Pire, C. Ferrer, J. Bonete, M.J. Rice, D.W. 2005-08-01 doi:10.1107/S1744309105019949 International Union of Crystallography Single crystals of binary and ternary complexes of wild-type and D38C mutant H. mediterranei glucose dehydrogenase have been obtained by the hanging-drop vapour-diffusion method. en GLUCOSE DEHYDROGENASE; SITE-DIRECTED MUTAGENESIS; MDR SUPERFAMILY; ARCHAEA Haloferax mediterranei glucose dehydrogenase (EC 1.1.1.47) belongs to the medium-chain alcohol dehydrogenase superfamily and requires zinc for catalysis. In the majority of these family members, the catalytic zinc is tetrahedrally coordinated by the side chains of a cysteine, a histidine, a cysteine or glutamate and a water molecule. In H. mediterranei glucose dehydrogenase, sequence analysis indicates that the zinc coordination is different, with the invariant cysteine replaced by an aspartate residue. In order to analyse the significance of this replacement and to contribute to an understanding of the role of the metal ion in catalysis, a range of binary and ternary complexes of the wild-type and a D38C mutant protein have been crystallized. For most of the complexes, crystals belonging to space group I222 were obtained using sodium/potassium citrate as a precipitant. However, for the binary and non-productive ternary complexes with NADPH/Zn, it was necessary to replace the citrate with 2-methyl-2,4-pentanediol. Despite the radical change in conditions, the crystals thus formed were isomorphous. text/html Crystallization and preliminary X-ray analysis of binary and ternary complexes of Haloferax mediterranei glucose dehydrogenase text 8 61 2005-08-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 743 med@iucr.org 746 1744-3091 http://scripts.iucr.org/cgi-bin/paper?sw5001 The essential enzyme 2C-methyl-d-erythritol-2,4-cyclodiphosphate (MECP) synthase, found in most eubacteria and the apicomplexan parasites, participates in isoprenoid-precursor biosynthesis and is a validated target for the development of broad-spectrum antimicrobial drugs. The structure and mechanism of the enzyme have been elucidated and the recent exciting finding that the enzyme actually binds diphosphate-containing isoprenoids at the interface formed by the three subunits that constitute the active protein suggests the possibility of feedback regulation of MECP synthase. To investigate such a possibility, a form of the enzyme was sought that did not bind these ligands but which would retain the quaternary structure necessary to create the active site. Two amino acids, Arg142 and Glu144, in Escherichia coli MECP synthase were identified as contributing to ligand binding. Glu144 interacts directly with Arg142 and positions the basic residue to form two hydrogen bonds with the terminal phosphate group of the isoprenoid diphosphate ligand. This association occurs at the trimer interface and three of these arginines interact with the ligand phosphate group. A dual mutation was designed (Arg142 to methionine and Glu144 to leucine) to disrupt the electrostatic attractions between the enzyme and the phosphate group to investigate whether an enzyme without isoprenoid diphosphate could be obtained. A low-resolution crystal structure of the mutated MECP synthase Met142/Leu144 revealed that geranyl diphosphate was retained despite the removal of six hydrogen bonds normally formed with the enzyme. This indicates that these two hydrophilic residues on the surface of the enzyme are not major determinants of isoprenoid binding at the trimer interface but rather that hydrophobic interactions between the hydrocarbon tail and the core of the enzyme trimer dominate ligand binding. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Sgraja, T. Kemp, L.E. Ramsden, N. Hunter, W.N. 2005-07-01 doi:10.1107/S1744309105018762 International Union of Crystallography A double mutation designed to disrupt binding of isoprenoid diphosphate to an enzyme involved in isoprenoid biosynthesis was made and the structure determined. Despite the removal of six hydrogen-bonding interactions, the ligand, acquired during production in E. coli, remains bound. The reasons for this are discussed. en MECP SYNTHASE; SITE-DIRECTED MUTAGENESIS; ISOPRENOID BIOSYNTHESIS The essential enzyme 2C-methyl-d-erythritol-2,4-cyclodiphosphate (MECP) synthase, found in most eubacteria and the apicomplexan parasites, participates in isoprenoid-precursor biosynthesis and is a validated target for the development of broad-spectrum antimicrobial drugs. The structure and mechanism of the enzyme have been elucidated and the recent exciting finding that the enzyme actually binds diphosphate-containing isoprenoids at the interface formed by the three subunits that constitute the active protein suggests the possibility of feedback regulation of MECP synthase. To investigate such a possibility, a form of the enzyme was sought that did not bind these ligands but which would retain the quaternary structure necessary to create the active site. Two amino acids, Arg142 and Glu144, in Escherichia coli MECP synthase were identified as contributing to ligand binding. Glu144 interacts directly with Arg142 and positions the basic residue to form two hydrogen bonds with the terminal phosphate group of the isoprenoid diphosphate ligand. This association occurs at the trimer interface and three of these arginines interact with the ligand phosphate group. A dual mutation was designed (Arg142 to methionine and Glu144 to leucine) to disrupt the electrostatic attractions between the enzyme and the phosphate group to investigate whether an enzyme without isoprenoid diphosphate could be obtained. A low-resolution crystal structure of the mutated MECP synthase Met142/Leu144 revealed that geranyl diphosphate was retained despite the removal of six hydrogen bonds normally formed with the enzyme. This indicates that these two hydrophilic residues on the surface of the enzyme are not major determinants of isoprenoid binding at the trimer interface but rather that hydrophobic interactions between the hydrocarbon tail and the core of the enzyme trimer dominate ligand binding. text/html A double mutation of Escherichia coli 2C-methyl-d-erythritol-2,4-cyclodiphosphate synthase disrupts six hydrogen bonds with, yet fails to prevent binding of, an isoprenoid diphosphate text 7 61 2005-07-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 protein structure communications 625 med@iucr.org 629 1744-3091 http://scripts.iucr.org/cgi-bin/paper?pu5093 Penicillin acylase proteins are amidohydrolase enzymes that cleave penicillins at the amide bond connecting the side chain to their β-lactam nucleus. An unannotated protein from Bacillus subtilis has been expressed in Escherichia coli, purified and confirmed to possess penicillin V acylase activity. The protein was crystallized using the hanging-drop vapour-diffusion method from a solution containing 4 M sodium formate in 100 mM Tris–HCl buffer pH 8.2. Diffraction data were collected under cryogenic conditions to a spacing of 2.5 Å. The crystals belonged to the orthorhombic space group C2221, with unit-cell parameters a = 111.0, b = 308.0, c = 56.0 Å. The estimated Matthews coefficient was 3.23 Å3 Da−1, corresponding to 62% solvent content. The structure has been solved using molecular-replacement methods with B. sphaericus penicillin V acylase (PDB code 2pva) as the search model. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Rathinaswamy, P. Pundle, A.V. Prabhune, A.A. SivaRaman, H. Brannigan, J.A. Dodson, G.G. Suresh, C.G. 2005-07-01 doi:10.1107/S1744309105017987 International Union of Crystallography An unannotated protein reported from B. subtilis has been expressed in E. coli and identified as possessing penicillin V acylase activity. The crystallization and preliminary crystallographic analysis of this penicillin V acylase is presented. en NTN HYDROLASE; PENICILLIN V ACYLASE; CONJUGATED BILE-SALT HYDROLASE Penicillin acylase proteins are amidohydrolase enzymes that cleave penicillins at the amide bond connecting the side chain to their β-lactam nucleus. An unannotated protein from Bacillus subtilis has been expressed in Escherichia coli, purified and confirmed to possess penicillin V acylase activity. The protein was crystallized using the hanging-drop vapour-diffusion method from a solution containing 4 M sodium formate in 100 mM Tris–HCl buffer pH 8.2. Diffraction data were collected under cryogenic conditions to a spacing of 2.5 Å. The crystals belonged to the orthorhombic space group C2221, with unit-cell parameters a = 111.0, b = 308.0, c = 56.0 Å. The estimated Matthews coefficient was 3.23 Å3 Da−1, corresponding to 62% solvent content. The structure has been solved using molecular-replacement methods with B. sphaericus penicillin V acylase (PDB code 2pva) as the search model. text/html Cloning, purification, crystallization and preliminary structural studies of penicillin V acylase from Bacillus subtilis text 7 61 2005-07-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 680 med@iucr.org 683 1744-3091 http://scripts.iucr.org/cgi-bin/paper?gx5059 The BA4499 and BA5696 genes of Bacillus anthracis encode proteins homologous to manganese superoxide dismutase, suggesting that this organism has an expanded repertoire of antioxidant proteins. Differences in metal specificity and quaternary structure between the dismutases of prokaryotes and higher eukaryotes may be exploited in the development of therapeutic antibacterial compounds. Here, the crystal structure of two Mn superoxide dismutases from B. anthracis solved to high resolution are reported. Comparison of their structures reveals that a highly conserved residue near the active centre is substituted in one of the proteins and that this is a characteristic feature of superoxide dismutases from the B. cereus/B. anthracis/B. thuringiensis group of organisms. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Boucher, I.W. Kalliomaa, A.K. Levdikov, V.M. Blagova, E.V. Fogg, M.J. Brannigan, J.A. Wilson, K.S. Wilkinson, A.J. 2005-07-01 doi:10.1107/S1744309105017380 International Union of Crystallography The crystal structures of two manganese superoxide dismutases from B. anthracis were solved by X-ray crystallography using molecular replacement. en SUPEROXIDE DISMUTASES The BA4499 and BA5696 genes of Bacillus anthracis encode proteins homologous to manganese superoxide dismutase, suggesting that this organism has an expanded repertoire of antioxidant proteins. Differences in metal specificity and quaternary structure between the dismutases of prokaryotes and higher eukaryotes may be exploited in the development of therapeutic antibacterial compounds. Here, the crystal structure of two Mn superoxide dismutases from B. anthracis solved to high resolution are reported. Comparison of their structures reveals that a highly conserved residue near the active centre is substituted in one of the proteins and that this is a characteristic feature of superoxide dismutases from the B. cereus/B. anthracis/B. thuringiensis group of organisms. text/html Structures of two superoxide dismutases from Bacillus anthracis reveal a novel active centre text 7 61 2005-07-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 protein structure communications 621 med@iucr.org 624 1744-3091 http://scripts.iucr.org/cgi-bin/paper?bw5087 Simian immunodeficiency virus (SIV) infection in rhesus macaques has been used as the best model for the study of human immunodeficiency virus (HIV) infection in humans, especially in the cytotoxic T-lymphocyte (CTL) response. However, the structure of rhesus macaque (or any other monkey model) major histocompatibility complex class I (MHC I) presenting a specific peptide (the ligand for CTL) has not yet been elucidated. Here, using in vitro refolding, the preparation of the complex of the rhesus macaque MHC I allele (Mamu-A*01) with human β2m and an immunodominant peptide, CTPYDINQM (Gag_CM9), derived from SIV Gag protein is reported. The complex (45 kDa) was crystallized; the crystal belongs to space group I422, with unit-cell parameters a = b = 183.8, c = 155.2 Å. The crystal contains two molecules in the asymmetric unit and diffracts X-rays to 2.8 Å resolution. The structure is being solved by molecular replacement and this is the first attempt to determined the crystal structure of a peptide–nonhuman primate MHC complex Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Chu, F. Lou, Z. Gao, B. Bell, J.I. Rao, Z. Gao, G.F. 2005-06-01 doi:10.1107/S1744309105016453 International Union of Crystallography Crystallization of the first rhesus macaque MHC class I complex. en MAMU-A*01 COMPLEX; SIMIAN IMMUNODEFICIENCY VIRUS; MAJOR HISTOCOMPATIBILITY COMPLEX Simian immunodeficiency virus (SIV) infection in rhesus macaques has been used as the best model for the study of human immunodeficiency virus (HIV) infection in humans, especially in the cytotoxic T-lymphocyte (CTL) response. However, the structure of rhesus macaque (or any other monkey model) major histocompatibility complex class I (MHC I) presenting a specific peptide (the ligand for CTL) has not yet been elucidated. Here, using in vitro refolding, the preparation of the complex of the rhesus macaque MHC I allele (Mamu-A*01) with human β2m and an immunodominant peptide, CTPYDINQM (Gag_CM9), derived from SIV Gag protein is reported. The complex (45 kDa) was crystallized; the crystal belongs to space group I422, with unit-cell parameters a = b = 183.8, c = 155.2 Å. The crystal contains two molecules in the asymmetric unit and diffracts X-rays to 2.8 Å resolution. The structure is being solved by molecular replacement and this is the first attempt to determined the crystal structure of a peptide–nonhuman primate MHC complex text/html Complex assembly, crystallization and preliminary X-ray crystallographic studies of rhesus macaque MHC Mamu-A*01 complexed with an immunodominant SIV-Gag nonapeptide text 6 61 2005-06-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 614 med@iucr.org 616 1744-3091 http://scripts.iucr.org/cgi-bin/paper?bw5088 Mevalonate diphosphate decarboxylase catalyses the last and least well characterized step in the mevalonate pathway for the biosynthesis of isopentenyl pyrophosphate, an isoprenoid precursor. A gene predicted to encode the enzyme from Trypanosoma brucei has been cloned, a highly efficient expression system established and a purification protocol determined. The enzyme gives monoclinic crystals in space group P21, with unit-cell parameters a = 51.5, b = 168.7, c = 54.9 Å, β = 118.8°. A Matthews coefficient VM of 2.5 Å3 Da−1 corresponds to two monomers, each approximately 42 kDa (385 residues), in the asymmetric unit with 50% solvent content. These crystals are well ordered and data to high resolution have been recorded using synchrotron radiation. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Byres, E. Martin, D.M.A. Hunter, W.N. 2005-06-01 doi:10.1107/S1744309105014594 International Union of Crystallography The gene encoding the putative mevalonate diphosphate decarboxylase, an enzyme from the mevalonate pathway of isoprenoid precursor biosynthesis, has been cloned from T. brucei. Recombinant protein has been expressed, purified and highly ordered crystals obtained and characterized to aid the structure–function analysis of this enzyme. en DECARBOXYLASES; MEVALONATE BIOSYNTHESIS; ISOPRENOIDS; TRYPANOSOMA Mevalonate diphosphate decarboxylase catalyses the last and least well characterized step in the mevalonate pathway for the biosynthesis of isopentenyl pyrophosphate, an isoprenoid precursor. A gene predicted to encode the enzyme from Trypanosoma brucei has been cloned, a highly efficient expression system established and a purification protocol determined. The enzyme gives monoclinic crystals in space group P21, with unit-cell parameters a = 51.5, b = 168.7, c = 54.9 Å, β = 118.8°. A Matthews coefficient VM of 2.5 Å3 Da−1 corresponds to two monomers, each approximately 42 kDa (385 residues), in the asymmetric unit with 50% solvent content. These crystals are well ordered and data to high resolution have been recorded using synchrotron radiation. text/html A preliminary crystallographic analysis of the putative mevalonate diphosphate decarboxylase from Trypanosoma brucei text 6 61 2005-06-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 581 med@iucr.org 584 1744-3091 http://scripts.iucr.org/cgi-bin/paper?en5115 Crystals of native histone octamers (H2A–H2B)–(H4–H3)–(H3′–H4′)–(H2B′–­H2A′) from chick erythrocytes in 2 M KCl, 1.35 M potassium phosphate pH 6.9 diffract X-rays to 1.90 Å resolution, yielding a structure with an Rwork value of 18.7% and an Rfree of 22.2%. The crystal space group is P65, the asymmetric unit of which contains one complete octamer. This high-resolution model of the histone-core octamer allows further insight into intermolecular interactions, including water molecules, that dock the histone dimers to the tetramer in the nucleosome-core particle and have relevance to nucleosome remodelling. The three key areas analysed are the H2A′–H3–H4 molecular cluster (also H2A–H3′–H4′), the H4–H2B′ interaction (also H4′–H2B) and the H2A′–H4 β-sheet interaction (also H2A–H4′). The latter of these three regions is important to nucleosome remodelling by RNA polymerase II, as it is shown to be a likely core-histone binding site, and its disruption creates an instability in the nucleosome-core particle. A majority of the water molecules in the high-resolution octamer have positions that correlate to similar positions in the high-­resolution nucleosome-core particle structure, suggesting that the high-resolution octamer model can be used for comparative studies with the high-resolution nucleosome-core particle. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Wood, C.M. Nicholson, J.M. Lambert, S.J. Chantalat, L. Reynolds, C.D. Baldwin, J.P. 2005-06-01 doi:10.1107/S1744309105013813 International Union of Crystallography The high-resolution (1.90 Å) model of the native histone octamer allows structural comparisons to be made with the nucleosome-core particle, along with an identification of a likely core-histone binding site. en HISTONE OCTAMER Crystals of native histone octamers (H2A–H2B)–(H4–H3)–(H3′–H4′)–(H2B′–­H2A′) from chick erythrocytes in 2 M KCl, 1.35 M potassium phosphate pH 6.9 diffract X-rays to 1.90 Å resolution, yielding a structure with an Rwork value of 18.7% and an Rfree of 22.2%. The crystal space group is P65, the asymmetric unit of which contains one complete octamer. This high-resolution model of the histone-core octamer allows further insight into intermolecular interactions, including water molecules, that dock the histone dimers to the tetramer in the nucleosome-core particle and have relevance to nucleosome remodelling. The three key areas analysed are the H2A′–H3–H4 molecular cluster (also H2A–H3′–H4′), the H4–H2B′ interaction (also H4′–H2B) and the H2A′–H4 β-sheet interaction (also H2A–H4′). The latter of these three regions is important to nucleosome remodelling by RNA polymerase II, as it is shown to be a likely core-histone binding site, and its disruption creates an instability in the nucleosome-core particle. A majority of the water molecules in the high-resolution octamer have positions that correlate to similar positions in the high-­resolution nucleosome-core particle structure, suggesting that the high-resolution octamer model can be used for comparative studies with the high-resolution nucleosome-core particle. text/html High-resolution structure of the native histone octamer text 6 61 2005-06-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 protein structure communications 541 med@iucr.org 545 1744-3091 http://scripts.iucr.org/cgi-bin/paper?bw5085 The oxirane (1R,2S)-1,2-epoxypropylphosphonic acid (fosfomycin) is a natural product antibiotic produced in Streptomyces wedmorensis by the metal-ion-dependent (S)-2-hydroxypropylphosphonic acid epoxidase. This epoxidase is highly unusual since it has no requirement for a haem prosthetic group. The gene encoding the enzyme, fom4, has been cloned and a highly efficient recombinant source of the enzyme established. Two different crystal forms, tetragonal and hexagonal, have been obtained. The hexagonal form displays symmetry consistent with space group P61/522 and unit-cell parameters a = 86.44, c = 221.56 Å, γ = 120°. The Matthews coefficient, VM, of 2.7 Å3 Da−1 corresponds to two subunits, each of approximate weight 21.4 kDa, in the asymmetric unit with 55% solvent content. These crystals diffract to high resolution and experimental phases are being sought to determine the structure. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Cameron, S. McLuskey, K. Chamberlayne, R. Hallyburton, I. Hunter, W.N. 2005-05-01 doi:10.1107/S1744309105012376 International Union of Crystallography The gene encoding the unusual metal-ion-dependent epoxidase involved in fosfomycin biosynthesis, S. wedmorensis (S)-2-hydroxypropylphosphonic acid epoxidase, has been cloned and the protein expressed, purified and crystallized. Two crystal forms have been obtained, one of which diffracts to high resolution. en ANTIBIOTICS; EPOXIDASES; METALLOENZYMES; STREPTOMYCES The oxirane (1R,2S)-1,2-epoxypropylphosphonic acid (fosfomycin) is a natural product antibiotic produced in Streptomyces wedmorensis by the metal-ion-dependent (S)-2-hydroxypropylphosphonic acid epoxidase. This epoxidase is highly unusual since it has no requirement for a haem prosthetic group. The gene encoding the enzyme, fom4, has been cloned and a highly efficient recombinant source of the enzyme established. Two different crystal forms, tetragonal and hexagonal, have been obtained. The hexagonal form displays symmetry consistent with space group P61/522 and unit-cell parameters a = 86.44, c = 221.56 Å, γ = 120°. The Matthews coefficient, VM, of 2.7 Å3 Da−1 corresponds to two subunits, each of approximate weight 21.4 kDa, in the asymmetric unit with 55% solvent content. These crystals diffract to high resolution and experimental phases are being sought to determine the structure. text/html Initiating a crystallographic analysis of recombinant (S)-2-hydroxypropylphosphonic acid epoxidase from Streptomyces wedmorensis text 5 61 2005-05-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 534 med@iucr.org 536 1744-3091 http://scripts.iucr.org/cgi-bin/paper?gx5050 Protein structures from the causative agent of anthrax (Bacillus anthracis) are being determined as part of a structural genomics programme. Amongst initial candidates for crystallographic analysis are enzymes involved in nucleotide biosynthesis, since these are recognized as potential targets in antibacterial therapy. Purine nucleoside phosphorylase is a key enzyme in the purine-salvage pathway. The crystal structure of purine nucleoside phosphorylase (DeoD) from B. anthracis has been solved by molecular replacement at 2.24 Å resolution and refined to an R factor of 18.4%. This is the first report of a DeoD structure from a Gram-positive bacterium. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Grenha, R. Levdikov, V.M. Fogg, M.J. Blagova, E.V. Brannigan, J.A. Wilkinson, A.J. Wilson, K.S. 2005-05-01 doi:10.1107/S174430910501095X International Union of Crystallography The crystal structure of purine nucleoside phosphorylase (DeoD) from B. anthracis was solved by X-ray crystallography using molecular replacement and refined at a resolution of 2.24 Å. en PURINE NUCLEOSIDE PHOSPHORYLASE; PURINE SALVAGE Protein structures from the causative agent of anthrax (Bacillus anthracis) are being determined as part of a structural genomics programme. Amongst initial candidates for crystallographic analysis are enzymes involved in nucleotide biosynthesis, since these are recognized as potential targets in antibacterial therapy. Purine nucleoside phosphorylase is a key enzyme in the purine-salvage pathway. The crystal structure of purine nucleoside phosphorylase (DeoD) from B. anthracis has been solved by molecular replacement at 2.24 Å resolution and refined to an R factor of 18.4%. This is the first report of a DeoD structure from a Gram-positive bacterium. text/html Structure of purine nucleoside phosphorylase (DeoD) from Bacillus anthracis text 5 61 2005-05-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 structural genomics communications 459 med@iucr.org 462 1744-3091 http://scripts.iucr.org/cgi-bin/paper?pu5087 Domain III of the West Nile virus envelope protein, the putative receptor-binding domain, is a major virion-surface determinant for virulence. This protein was reported to be intrinsically unstable and has defied previous crystallization attempts. It has now been purified from inclusion bodies by protein refolding and was crystallized using the hanging-drop vapour-diffusion method at 291 K. The crystals belong to space group P2221, with unit-cell parameters a = 52.6, b = 59.7, c = 95.0 Å. A complete data set was collected to 2.8 Å at 100 K with Cu Kα X-rays from a rotating-anode generator. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Yuan, F. Lou, Z. Li, X. Chen, Y.W. Bell, J.I. Rao, Z. Gao, G.F. 2005-04-01 doi:10.1107/S1744309105008195 International Union of Crystallography Domain III of the West Nile virus envelope protein, the putative receptor-binding domain, was reported to be intrinsically unstable and has defied previous crystallization attempts. It has now been purified from inclusion bodies by protein refolding and was crystallized using the hanging-drop vapour-diffusion method at 291 K. en WEST NILE VIRUS; FLAVIVIRUSES; ENVELOPE PROTEIN DOMAIN III; PROTEIN REFOLDING Domain III of the West Nile virus envelope protein, the putative receptor-binding domain, is a major virion-surface determinant for virulence. This protein was reported to be intrinsically unstable and has defied previous crystallization attempts. It has now been purified from inclusion bodies by protein refolding and was crystallized using the hanging-drop vapour-diffusion method at 291 K. The crystals belong to space group P2221, with unit-cell parameters a = 52.6, b = 59.7, c = 95.0 Å. A complete data set was collected to 2.8 Å at 100 K with Cu Kα X-rays from a rotating-anode generator. text/html Refolding, crystallization and preliminary X-ray structural studies of the West Nile virus envelope (E) protein domain III text 4 61 2005-04-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 421 med@iucr.org 423 1744-3091 http://scripts.iucr.org/cgi-bin/paper?za5094 CobE, a protein implicated in vitamin B12 biosynthesis, from Pseudomonas aeruginosa has been overexpressed in Escherichia coli, purified and crystallized using hanging-drop vapour diffusion. The crystals belong to the primitive orthorhombic space group P212121, with unit-cell parameters a = 31.86, b = 41.07, c = 87.41 Å. The diffraction extends to a resolution of 1.9 Å. There is one molecule per asymmetric unit and the estimated solvent content is 35%. SeMet-labelled CobE has been prepared and crystallizes under the same conditions as the native protein with diffraction to 1.7 Å. The anomalous measurements will be used for phasing. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Vévodová, J. Graham, R.M. Raux, E. Warren, M.J. Wilson, K.S. 2005-04-01 doi:10.1107/S1744309105006731 International Union of Crystallography P. aeruginosa CobE, a protein implicated in vitamin B12 biosynthesis, has been crystallized and data on the native and SeMet forms recorded to resolutions of 1.9 and 1.7 Å, respectively. The anomalous measurements will be used for phasing. en COBE; VITAMIN B12 BIOSYNTHESIS CobE, a protein implicated in vitamin B12 biosynthesis, from Pseudomonas aeruginosa has been overexpressed in Escherichia coli, purified and crystallized using hanging-drop vapour diffusion. The crystals belong to the primitive orthorhombic space group P212121, with unit-cell parameters a = 31.86, b = 41.07, c = 87.41 Å. The diffraction extends to a resolution of 1.9 Å. There is one molecule per asymmetric unit and the estimated solvent content is 35%. SeMet-labelled CobE has been prepared and crystallizes under the same conditions as the native protein with diffraction to 1.7 Å. The anomalous measurements will be used for phasing. text/html Crystallization and preliminary structure analysis of CobE, an essential protein of cobalamin (vitamin B12) biosynthesis text 4 61 2005-04-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 442 med@iucr.org 444 1744-3091 http://scripts.iucr.org/cgi-bin/paper?pu5080 An open reading frame designated b2910 (ygfE) in the Escherichia coli K12-MG1655 genome sequence, identified as a possible homologue to the cell-division protein ZapA, was cloned into the high-expression plasmid pETDuet-1 and overexpressed in E. coli BL21 (DE3)-AI. The protein was purified in three steps to 99% purity. Crystals were obtained by the hanging-drop vapour-diffusion method at 291 K from a wide range of screened conditions, but principally from solutions containing 0.1 M HEPES pH 7.0, 18% PEG 6000, 5 mM CaCl2. Diffraction data to 1.8 Å were collected at the European Synchrotron Radiation Facility (ESRF). The crystals belong to space group P6122 or P6522, with unit-cell parameters a = 53.8, b = 53.8, c = 329.7 Å, α = β = 90, γ = 120°. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Addinall, S.G. Johnson, K.A. Dafforn, T. Smith, C. Rodger, A. Gomez, R.P. Sloan, K. Blewett, A. Scott, D.J. Roper, D.I. 2005-03-01 doi:10.1107/S1744309105003945 International Union of Crystallography An open reading frame from E. coli MG1655 has been cloned, expressed and purified. Crystals obtained from the purified recombinant protein have been obtained in a variety of different forms diffracting to 1.8 Å resolution. en CELL-DIVISION PROTEINS; YGFE An open reading frame designated b2910 (ygfE) in the Escherichia coli K12-MG1655 genome sequence, identified as a possible homologue to the cell-division protein ZapA, was cloned into the high-expression plasmid pETDuet-1 and overexpressed in E. coli BL21 (DE3)-AI. The protein was purified in three steps to 99% purity. Crystals were obtained by the hanging-drop vapour-diffusion method at 291 K from a wide range of screened conditions, but principally from solutions containing 0.1 M HEPES pH 7.0, 18% PEG 6000, 5 mM CaCl2. Diffraction data to 1.8 Å were collected at the European Synchrotron Radiation Facility (ESRF). The crystals belong to space group P6122 or P6522, with unit-cell parameters a = 53.8, b = 53.8, c = 329.7 Å, α = β = 90, γ = 120°. text/html Expression, purification and crystallization of the cell-division protein YgfE from Escherichia coli text 3 61 2005-03-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 305 med@iucr.org 307 1744-3091 http://scripts.iucr.org/cgi-bin/paper?pu5081 The class I CD8 positive T-cell response is involved in a number of conditions in which artificial down-regulation and control would be therapeutically beneficial. Such conditions include a number of autoimmune diseases and graft rejection in transplant patients. Although the CD8 T-cell response is dominated by the TCR–pMHC interaction, activation of T cells is in most cases also dependent on a number of associated signalling molecules. Previous work has demonstrated the ability of one such molecule (CD8) to act as an antagonist to T-cell activation if added in soluble form. Therefore, a high-affinity mutant CD8 (haCD8) has been developed with the aim of developing a therapeutic immunosuppressor. In order to fully understand the nature of the haCD8 interaction, this protein was crystallized using the sitting-drop vapour-diffusion method. Single haCD8 crystals were cryocooled and used for data collection. These crystals belonged to space group P6422 (assumed by similarity to the wild type), with unit-cell parameters a = 101.08, c = 56.54 Å. VM calculations indicated one molecule per asymmetric unit. A 2 Å data set was collected and the structure is currently being determined using molecular replacement. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Cole, D.K. Rizkallah, P.J. Sami, M. Lissin, N.M. Gao, F. Bell, J.I. Boulter, J.M. Glick, M. Vuidepot, A. Jakobsen, B.K. Gao, G.F. 2005-03-01 doi:10.1107/S1744309105002988 International Union of Crystallography A high-affinity mutant CD8 (haCD8) has been developed with the aim of developing a therapeutic immunosuppressor. In order to fully understand the nature of the haCD8 interaction, this protein was crystallized using the sitting-drop vapour-diffusion method. en CD8[ALPHA][ALPHA]; HIGH AFFINITY; STRUCTURE-BASED DESIGN; IMMUNOSUPPRESSORS The class I CD8 positive T-cell response is involved in a number of conditions in which artificial down-regulation and control would be therapeutically beneficial. Such conditions include a number of autoimmune diseases and graft rejection in transplant patients. Although the CD8 T-cell response is dominated by the TCR–pMHC interaction, activation of T cells is in most cases also dependent on a number of associated signalling molecules. Previous work has demonstrated the ability of one such molecule (CD8) to act as an antagonist to T-cell activation if added in soluble form. Therefore, a high-affinity mutant CD8 (haCD8) has been developed with the aim of developing a therapeutic immunosuppressor. In order to fully understand the nature of the haCD8 interaction, this protein was crystallized using the sitting-drop vapour-diffusion method. Single haCD8 crystals were cryocooled and used for data collection. These crystals belonged to space group P6422 (assumed by similarity to the wild type), with unit-cell parameters a = 101.08, c = 56.54 Å. VM calculations indicated one molecule per asymmetric unit. A 2 Å data set was collected and the structure is currently being determined using molecular replacement. text/html Crystallization and preliminary X-ray structural studies of a high-affinity CD8αα co-receptor to pMHC text 3 61 2005-03-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 285 med@iucr.org 287 1744-3091 http://scripts.iucr.org/cgi-bin/paper?en5079 The constitutive androstane receptor (CAR) is a member of the nuclear receptor superfamily. In contrast to classical nuclear receptors, which possess small-molecule ligand-inducible activity, CAR exhibits constitutive transcriptional activity in the apparent absence of ligand. CAR is among the most important transcription factors; it coordinately regulates the expression of microsomal cytochrome P450 genes and other drug-metabolizing enzymes. The murine CAR ligand-binding domain (LBD) was coexpressed with the steroid receptor coactivator protein (SRC-1) receptor-interacting domain (RID) in Escherichia coli. The mCAR LBD subunit was purified away from SRC-1 by affinity, anion-exchange and size-exclusion chromatography, crystallized with androstenol and the structure of the complex determined by molecular replacement. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Vincent, J. Shan, L. Fan, M. Brunzelle, J.S. Forman, B.M. Fernandez, E.J. 2005-01-01 doi:10.1107/S1744309104032762 International Union of Crystallography The purification and structure determination of the murine constitutive androstane receptor bound to its inverse agonist/antagonist androstenol is described. en NUCLEAR RECEPTOR; CONSTITUTIVE ANDROSTANE RECEPTOR; CAR; ANDROSTENOL The constitutive androstane receptor (CAR) is a member of the nuclear receptor superfamily. In contrast to classical nuclear receptors, which possess small-molecule ligand-inducible activity, CAR exhibits constitutive transcriptional activity in the apparent absence of ligand. CAR is among the most important transcription factors; it coordinately regulates the expression of microsomal cytochrome P450 genes and other drug-metabolizing enzymes. The murine CAR ligand-binding domain (LBD) was coexpressed with the steroid receptor coactivator protein (SRC-1) receptor-interacting domain (RID) in Escherichia coli. The mCAR LBD subunit was purified away from SRC-1 by affinity, anion-exchange and size-exclusion chromatography, crystallized with androstenol and the structure of the complex determined by molecular replacement. text/html Crystallographic analysis of murine constitutive androstane receptor ligand-binding domain complexed with 5α-androst-16-en-3α-ol text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 156 med@iucr.org 159 1744-3091 http://scripts.iucr.org/cgi-bin/paper?gx5036 The crystal structure of APE2540, the putative trans-editing enzyme ProX from Aeropyrum pernix K1, was determined in a high-throughput manner. The crystal belongs to the monoclinic space group P21, with unit-cell parameters a = 47.4, b = 58.9, c = 53.6 Å, β = 106.8°. The structure was solved by the multiwavelength anomalous dispersion method at 1.7 Å and refined to an R factor of 16.8% (Rfree = 20.5%). The crystal structure includes two protein molecules in the asymmetric unit. Each monomer consists of eight β-strands and seven α-helices. A structure-homology search revealed similarity between the trans-editing enzyme YbaK (or cysteinyl-tRNAPro deacylase) from Haemophilus influenzae (HI1434; 22% sequence identity) and putative ProX proteins from Caulobacter crescentus (16%) and Agrobacterium tumefaciens (21%). Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Murayama, K. Kato-Murayama, M. Katsura, K. Uchikubo-Kamo, T. Yamaguchi-Hirafuji, M. Kawazoe, M. Akasaka, R. Hanawa-Suetsugu, K. Hori-Takemoto, C. Terada, T. Shirouzu, M. Yokoyama, S. 2005-01-01 doi:10.1107/S1744309104032555 International Union of Crystallography The three-dimensional structure of the APE2540 protein from A. pernix K1 has been determined by the multiple anomalous dispersion method at 1.7 Å resolution. The structure includes two monomers in the asymmetric unit and shares structural similarity with the YbaK protein or cysteinyl-tRNAPro deacylase from H. influenzae. en TRANS-EDITING ENZYMES; APE2540 The crystal structure of APE2540, the putative trans-editing enzyme ProX from Aeropyrum pernix K1, was determined in a high-throughput manner. The crystal belongs to the monoclinic space group P21, with unit-cell parameters a = 47.4, b = 58.9, c = 53.6 Å, β = 106.8°. The structure was solved by the multiwavelength anomalous dispersion method at 1.7 Å and refined to an R factor of 16.8% (Rfree = 20.5%). The crystal structure includes two protein molecules in the asymmetric unit. Each monomer consists of eight β-strands and seven α-helices. A structure-homology search revealed similarity between the trans-editing enzyme YbaK (or cysteinyl-tRNAPro deacylase) from Haemophilus influenzae (HI1434; 22% sequence identity) and putative ProX proteins from Caulobacter crescentus (16%) and Agrobacterium tumefaciens (21%). text/html Structure of a putative trans-editing enzyme for prolyl-tRNA synthetase from Aeropyrum pernix K1 at 1.7 Å resolution text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 structural genomics communications 26 med@iucr.org 29 1744-3091 http://scripts.iucr.org/cgi-bin/paper?pu5066 Coagulation factor IX-binding protein isolated from Trimeresurus flavoviridis (IX-bp) is a C-type lectin-like protein. It is an anticoagulant protein consisting of homologous subunits A and B. The subunits both contain a Ca2+-binding site with differing affinity (Kd values of 14 and 130 µM at pH 7.5). These binding characteristics are pH-dependent; under acidic conditions, the affinity of the low-affinity site was reduced considerably. In order to identify which site has high affinity and also to investigate the Ca2+-releasing mechanism, IX-bp was crystallized at pH 6.5 and 4.6. The crystals at pH 6.5 and 4.6 diffracted to 1.72 and 2.29 Å resolution, respectively; the former crystals belong to the monoclinic space group P21, with unit-cell parameters a = 60.7, b = 63.5, c = 66.9 Å, β = 117.0°, while the latter belong to the monoclinic space group C2, with a = 134.1, b = 37.8, c = 55.8 Å, β = 110.4°. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Suzuki, N. Shikamoto, Y. Fujimoto, Z. Morita, T. Mizuno, H. 2005-01-01 doi:10.1107/S1744309104032439 International Union of Crystallography Crystals of habu coagulation factor IX-binding protein have been obtained at pH 6.5 and 4.6 and characterized by X-ray diffraction. en COAGULATION FACTOR BINDING PROTEIN; CALCIUM BINDING; SNAKE VENOMS; PH-DEPENDENT BINDING Coagulation factor IX-binding protein isolated from Trimeresurus flavoviridis (IX-bp) is a C-type lectin-like protein. It is an anticoagulant protein consisting of homologous subunits A and B. The subunits both contain a Ca2+-binding site with differing affinity (Kd values of 14 and 130 µM at pH 7.5). These binding characteristics are pH-dependent; under acidic conditions, the affinity of the low-affinity site was reduced considerably. In order to identify which site has high affinity and also to investigate the Ca2+-releasing mechanism, IX-bp was crystallized at pH 6.5 and 4.6. The crystals at pH 6.5 and 4.6 diffracted to 1.72 and 2.29 Å resolution, respectively; the former crystals belong to the monoclinic space group P21, with unit-cell parameters a = 60.7, b = 63.5, c = 66.9 Å, β = 117.0°, while the latter belong to the monoclinic space group C2, with a = 134.1, b = 37.8, c = 55.8 Å, β = 110.4°. text/html Crystallization and preliminary X-ray analysis of coagulation factor IX-binding protein from habu snake venom at pH 6.5 and 4.6 text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 147 med@iucr.org 149 1744-3091 http://scripts.iucr.org/cgi-bin/paper?en5074 In bacteria, protein expression initiates with an N-formyl group and this needs to be removed in order to ensure proper bacterial growth. These formylation and deformylation processes are unique to eubacteria; therefore, inhibition of these would provide a novel antibacterial therapy. Deformylation is carried out by peptide deformylase (PDF). PDF from Bacillus cereus, one of the major pathogenic bacteria, was cloned into expression plasmid pET-28a (Novagen), overexpressed in Escherichia coli BL21 (DE3) and purified to high quality. Crystals have been obtained of both ligand-free PDF and PDF to which actinonin, a highly potent naturally occurring inhibitor, is bound. Both crystals belong to space group P212121, with unit-cell parameters a = 42.72, b = 44.04, c = 85.19 Å and a = 41.31, b = 44.56, c = 84.47 Å, respectively. Diffraction data were collected to 1.7 Å resolution for the inhibitor-free crystals and to 2.0 Å resolution for the actinonin-bound crystals. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Park, J.K. Moon, J.H. Kim, J.-H. Kim, E.E. 2005-01-01 doi:10.1107/S1744309104032440 International Union of Crystallography Peptide deformylase (PDF) from B. cereus has been overexpressed, purified and crystallized in ligand-free and actinonin-bound forms. Diffraction data have been collected from these crystals to 1.7 and 2.0 Å resolution, respectively. en PROTEIN SYNTHESIS; PDF; PEPTIDE DEFORMYLASE; ANTIBACTERIAL DRUG TARGETS; BACILLUS CEREUS In bacteria, protein expression initiates with an N-formyl group and this needs to be removed in order to ensure proper bacterial growth. These formylation and deformylation processes are unique to eubacteria; therefore, inhibition of these would provide a novel antibacterial therapy. Deformylation is carried out by peptide deformylase (PDF). PDF from Bacillus cereus, one of the major pathogenic bacteria, was cloned into expression plasmid pET-28a (Novagen), overexpressed in Escherichia coli BL21 (DE3) and purified to high quality. Crystals have been obtained of both ligand-free PDF and PDF to which actinonin, a highly potent naturally occurring inhibitor, is bound. Both crystals belong to space group P212121, with unit-cell parameters a = 42.72, b = 44.04, c = 85.19 Å and a = 41.31, b = 44.56, c = 84.47 Å, respectively. Diffraction data were collected to 1.7 Å resolution for the inhibitor-free crystals and to 2.0 Å resolution for the actinonin-bound crystals. text/html Crystallization and preliminary X-ray crystallographic analysis of peptide deformylase (PDF) from Bacillus cereus in ligand-free and actinonin-bound forms text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 150 med@iucr.org 152 1744-3091 http://scripts.iucr.org/cgi-bin/paper?gx5031 Keap1 (Kelch-like ECH-associating protein 1) is a negative regulator of the Nrf2 transcription factor in the cytoplasm. The Kelch/DGR (double-glycine repeat) domain of Keap1 associates with Nrf2 as well as with actin filaments. A recombinant protein containing both the Kelch/DGR domain and the C-­terminal region of mouse Keap1 was expressed in Escherichia coli, purified to near-homogeneity and crystallized by the sitting-drop vapour-diffusion method. The crystal belongs to space group P61 or P65, with unit-cell parameters a = b = 102.95, c = 55.21 Å, and contains one molecule in the asymmetric unit. A complete diffraction data was collected to 2.25 Å resolution using an R-AXIS IV++ imaging plate mounted on an RA-Micro7 Cu Kα rotating-anode X-ray generator. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Padmanabhan, B. Scharlock, M. Tong, K.I. Nakamura, Y. Kang, M.-I. Kobayashi, A. Matsumoto, T. Tanaka, A. Yamamoto, M. Yokoyama, S. 2005-01-01 doi:10.1107/S1744309104032506 International Union of Crystallography Keap1-DC (Kelch/double-glycine repeat and C-terminal region) of mouse Keap1 has been overexpressed in E. coli, purified and crystallized using the vapour-diffusion method. en NRF2; TRANSCRIPTION FACTOR; KEAP1; ACTIN BINDING; ANTIOXIDANTS Keap1 (Kelch-like ECH-associating protein 1) is a negative regulator of the Nrf2 transcription factor in the cytoplasm. The Kelch/DGR (double-glycine repeat) domain of Keap1 associates with Nrf2 as well as with actin filaments. A recombinant protein containing both the Kelch/DGR domain and the C-­terminal region of mouse Keap1 was expressed in Escherichia coli, purified to near-homogeneity and crystallized by the sitting-drop vapour-diffusion method. The crystal belongs to space group P61 or P65, with unit-cell parameters a = b = 102.95, c = 55.21 Å, and contains one molecule in the asymmetric unit. A complete diffraction data was collected to 2.25 Å resolution using an R-AXIS IV++ imaging plate mounted on an RA-Micro7 Cu Kα rotating-anode X-ray generator. text/html Purification, crystallization and preliminary X-ray diffraction analysis of the Kelch-like motif region of mouse Keap1 text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 153 med@iucr.org 155 1744-3091 http://scripts.iucr.org/cgi-bin/paper?za5080 Acylphosphatase is a ubiquitous small enzyme that was first characterized in mammals. It is involved in the hydrolysis of carboxyl-phosphate bonds in several acylphosphate substrates, such as carbamoylphosphate and 1,3-biphosphoglycerate; however, a consensus on acylphosphatase action in vivo has not yet been reached. Recent investigations have focused on acylphosphatases from lower phyla, such as Drosophila melanogaster and Escherichia coli, in view of the application of these small proteins as models in the study of folding, misfolding and aggregation processes. An acylphosphatase from the hyperthermophilic archaeon Sulfolobus solfataricus has been cloned, expressed and purified. Here, the growth and characterization of a triclinic and a monoclinic crystal form of the hyperthermophilic enzyme are reported; X-ray diffraction data have been collected to 1.27 and 1.90 Å resolution, respectively. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Zuccotti, S. Rosano, C. Bemporad, F. Stefani, M. Bolognesi, M. 2005-01-01 doi:10.1107/S1744309104032336 International Union of Crystallography S. solfataricus acylphosphatase has been expressed, purified and crystallized in two different crystal forms. Preliminary characterization of a triclinic and a monoclinic crystal form is reported and data were collected to 1.27 and 1.90 Å, respectively. en ACYLPHOSPHATASE; EXTREMOPHILIC PROTEIN; HYPERTHERMOPHILIC PROTEIN Acylphosphatase is a ubiquitous small enzyme that was first characterized in mammals. It is involved in the hydrolysis of carboxyl-phosphate bonds in several acylphosphate substrates, such as carbamoylphosphate and 1,3-biphosphoglycerate; however, a consensus on acylphosphatase action in vivo has not yet been reached. Recent investigations have focused on acylphosphatases from lower phyla, such as Drosophila melanogaster and Escherichia coli, in view of the application of these small proteins as models in the study of folding, misfolding and aggregation processes. An acylphosphatase from the hyperthermophilic archaeon Sulfolobus solfataricus has been cloned, expressed and purified. Here, the growth and characterization of a triclinic and a monoclinic crystal form of the hyperthermophilic enzyme are reported; X-ray diffraction data have been collected to 1.27 and 1.90 Å resolution, respectively. text/html Preliminary characterization of two different crystal forms of acylphosphatase from the hyperthermophile archaeon Sulfolobus solfataricus text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 144 med@iucr.org 146 1744-3091 http://scripts.iucr.org/cgi-bin/paper?en5072 The lectin isolated from mature seeds of Cicer arietinum (CAL) agglutinates pronase-treated rabbit and human erythrocytes and its haemagglutination activity is inhibited by fetuin and desialated fetuin but not by simple monosaccharides or oligosaccharides. The purified lectin is a dimer of molecular weight 43 000 Da composed of two identical subunits (MW 21 500), as confirmed by SDS–PAGE. The lectin has been crystallized using the hanging-drop vapour-diffusion method at 295 K over a well solution containing 0.2 M sodium acetate, 0.1 M sodium phosphate buffer pH 6.5 and 14%(w/v) polyethylene glycol 8000. The triangular prism-shaped crystals belong to space group R3 and have unit-cell parameters a = b = 81.2, c = 69.4 Å. The diffraction data are 93.8% complete to 2.3 Å Bragg spacing with an Rmerge of 0.103. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Katre, U.V. Gaikwad, S.M. Bhagyawant, S.S. Deshpande, U.D. Khan, M.I. Suresh, C.G. 2005-01-01 doi:10.1107/S1744309104032166 International Union of Crystallography The crystallization and characterization of a lectin isolated and purified from C. arietinum and possessing complex sugar specificity is reported. en COMPLEX SUGAR SPECIFICITY; LEGUME LECTIN; SEED ALBUMINS The lectin isolated from mature seeds of Cicer arietinum (CAL) agglutinates pronase-treated rabbit and human erythrocytes and its haemagglutination activity is inhibited by fetuin and desialated fetuin but not by simple monosaccharides or oligosaccharides. The purified lectin is a dimer of molecular weight 43 000 Da composed of two identical subunits (MW 21 500), as confirmed by SDS–PAGE. The lectin has been crystallized using the hanging-drop vapour-diffusion method at 295 K over a well solution containing 0.2 M sodium acetate, 0.1 M sodium phosphate buffer pH 6.5 and 14%(w/v) polyethylene glycol 8000. The triangular prism-shaped crystals belong to space group R3 and have unit-cell parameters a = b = 81.2, c = 69.4 Å. The diffraction data are 93.8% complete to 2.3 Å Bragg spacing with an Rmerge of 0.103. text/html Crystallization and preliminary X-ray characterization of a lectin from Cicer arietinum (chickpea) text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 141 med@iucr.org 143 1744-3091 http://scripts.iucr.org/cgi-bin/paper?gx5034 Isoquinoline 1-oxidoreductase (IOR) from Brevundimonas diminuta is a mononuclear molybdoenzyme of the xanthine-dehydrogenase family of proteins and catalyzes the conversion of isoquinoline to isoquinoline-1-one. Its primary sequence and behaviour, specifically in its substrate specificity and lipophilicity, differ from other members of the family. A crystal structure of the enzyme is expected to provide an explanation for these differences. This paper describes the crystallization and preliminary X-ray diffraction experiments as well as an optimized purification protocol for IOR. Crystallization of IOR was achieved using two different crystallization buffers. Streak-seeding and cross-linking were essential to obtain well diffracting crystals. Suitable cryo-conditions were found and a structure solution was obtained by molecular replacement. However, phases need to be improved in order to obtain a more interpretable electron-density map. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Boer, D.R. Müller, A. Fetzner, S. Lowe, D.J. Romão, M.J. 2005-01-01 doi:10.1107/S1744309104032105 International Union of Crystallography Crystallization of isoquinoline 1-oxidoreductase from B. diminuta was achieved using two different crystallization buffers. Streak-seeding and cross-linking were essential to obtain well diffracting crystals. Suitable cryo-conditions were found and a structure solution was obtained by molecular replacement. en ISOQUINOLINE 1-OXIDOREDUCTASE; XANTHINE OXIDASE/XANTHINE DEHYDROGENASE; OXIDOREDUCTASES; MOLYBDENUM ENZYMES; MOLYBDOPTERIN; BREVUNDIMONAS DIMINUTA Isoquinoline 1-oxidoreductase (IOR) from Brevundimonas diminuta is a mononuclear molybdoenzyme of the xanthine-dehydrogenase family of proteins and catalyzes the conversion of isoquinoline to isoquinoline-1-one. Its primary sequence and behaviour, specifically in its substrate specificity and lipophilicity, differ from other members of the family. A crystal structure of the enzyme is expected to provide an explanation for these differences. This paper describes the crystallization and preliminary X-ray diffraction experiments as well as an optimized purification protocol for IOR. Crystallization of IOR was achieved using two different crystallization buffers. Streak-seeding and cross-linking were essential to obtain well diffracting crystals. Suitable cryo-conditions were found and a structure solution was obtained by molecular replacement. However, phases need to be improved in order to obtain a more interpretable electron-density map. text/html On the purification and preliminary crystallographic analysis of isoquinoline 1-oxidoreductase from Brevundimonas diminuta 7 text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 137 med@iucr.org 140 1744-3091 http://scripts.iucr.org/cgi-bin/paper?bw5071 Crystals of the drug-discharge outer membrane protein OprM (MW = 50.9 kDa) of the MexAB-OprM multidrug transporter of Pseudomonas aeruginosa have been grown at 293 K in the presence of 2-methyl-2,4-propanediol and a combination of surfactants. The crystal belonged to space group R32, with unit-cell parameters a = b = 85.43, c = 1044.3 Å. Diffraction data for OprM were obtained using the undulator synchrotron-radiation beamline at SPring-8 (BL44XU, Osaka University), which allowed an extra-long specimen-to-detector distance with a wide detector area. The crystal diffracted to 2.56 Å resolution using 0.9 Å X-rays from the synchrotron-radiation source. A heavy-atom derivative for isomorphous replacement phasing was obtained using iridium chloride. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Akama, H. Kanemaki, M. Tsukihara, T. Nakagawa, A. Nakae, T. 2005-01-01 doi:10.1107/S1744309104031914 International Union of Crystallography The OprM subunit of the MexAB-OprM efflux pump in P. aeruginosa is an outer membrane-anchored lipoprotein. OprM crystals have been grown at 293 K in the presence of 2-methyl-2,4-propanediol and a combination of surfactants and diffracted to 2.56 Å resolution. en OPRM; LIPOPROTEINS; OUTER MEMBRANE PROTEINS Crystals of the drug-discharge outer membrane protein OprM (MW = 50.9 kDa) of the MexAB-OprM multidrug transporter of Pseudomonas aeruginosa have been grown at 293 K in the presence of 2-methyl-2,4-propanediol and a combination of surfactants. The crystal belonged to space group R32, with unit-cell parameters a = b = 85.43, c = 1044.3 Å. Diffraction data for OprM were obtained using the undulator synchrotron-radiation beamline at SPring-8 (BL44XU, Osaka University), which allowed an extra-long specimen-to-detector distance with a wide detector area. The crystal diffracted to 2.56 Å resolution using 0.9 Å X-rays from the synchrotron-radiation source. A heavy-atom derivative for isomorphous replacement phasing was obtained using iridium chloride. text/html Preliminary crystallographic analysis of the antibiotic discharge outer membrane lipoprotein OprM of Pseudomonas aeruginosa with an exceptionally long unit cell and complex lattice structure text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 131 med@iucr.org 133 1744-3091 http://scripts.iucr.org/cgi-bin/paper?en5082 Growth and differentiation factor 5 (GDF-5) belongs to the large TGF-β superfamily of secreted signalling proteins and plays a pivotal role in skeletal development during embryogenesis. The gene for human GDF-5 was cloned, expressed in Escherichia coli and purified to homogeneity. Crystals were obtained that diffracted to 2.2 Å resolution. A native data set was acquired, showing that the crystals belong to a trigonal space group, i.e. P3121 or P3221, with unit-cell parameters a = b = 97.1, c = 48.3 Å. Initial analysis suggest the presence of only one monomer in the asymmetric unit, resulting in a high solvent content of 72% in the crystal. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Mueller, T.D. Gottermeier, M. Sebald, W. Nickel, J. 2005-01-01 doi:10.1107/S1744309104031963 International Union of Crystallography Crystals of human growth and differentiation factor 5 are trigonal, belonging to space group P3121 or its enantiomer, with one molecule per asymmetric unit and diffract to 2.2 Å resolution. en HUMAN GROWTH AND DIFFERENTIATION FACTOR 5; SIGNALLING PROTEINS Growth and differentiation factor 5 (GDF-5) belongs to the large TGF-β superfamily of secreted signalling proteins and plays a pivotal role in skeletal development during embryogenesis. The gene for human GDF-5 was cloned, expressed in Escherichia coli and purified to homogeneity. Crystals were obtained that diffracted to 2.2 Å resolution. A native data set was acquired, showing that the crystals belong to a trigonal space group, i.e. P3121 or P3221, with unit-cell parameters a = b = 97.1, c = 48.3 Å. Initial analysis suggest the presence of only one monomer in the asymmetric unit, resulting in a high solvent content of 72% in the crystal. text/html Crystallization and preliminary X-ray diffraction analysis of human growth and differentiation factor 5 (GDF-5) text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 134 med@iucr.org 136 1744-3091 http://scripts.iucr.org/cgi-bin/paper?en5078 Osteoclast-stimulating factor increases osteoclast formation and bone resorption through a cellular signal transduction cascade, possibly by its interaction with c-Src or related family members. Crystals of human osteoclast-stimulating factor were obtained by the hanging-drop vapour-diffusion method using ammonium sulfate as a precipitant. The crystals are primitive orthorhombic and belong to P222 or a related space group, with unit-cell parameters a = 38.1, b = 54.9, c = 64.7 Å. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Li, M. Meng, Z. Xu, Y. Lou, Z. Rao, Z. 2005-01-01 doi:10.1107/S1744309104031653 International Union of Crystallography Crystals of human osteoclast-stimulating factor were obtained by the hanging-drop vapour-diffusion method using ammonium sulfate as a precipitant. The crystals are primitive orthorhombic and belong to P222 or a related space group, with unit-cell parameters a = 38.1, b = 54.9, c = 64.7 Å. en OSTEOCLAST-STIMULATING FACTOR Osteoclast-stimulating factor increases osteoclast formation and bone resorption through a cellular signal transduction cascade, possibly by its interaction with c-Src or related family members. Crystals of human osteoclast-stimulating factor were obtained by the hanging-drop vapour-diffusion method using ammonium sulfate as a precipitant. The crystals are primitive orthorhombic and belong to P222 or a related space group, with unit-cell parameters a = 38.1, b = 54.9, c = 64.7 Å. text/html Crystallization and preliminary X-ray crystallographic analysis of osteoclast-stimulating factor text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 128 med@iucr.org 130 1744-3091 http://scripts.iucr.org/cgi-bin/paper?en5086 The structures of mistletoe lectin I (ML-I) from Viscum album complexed with lactose and galactose have been determined at 2.3 Å resolution and refined to R factors of 20.9% (Rfree = 23.6%) and 20.9 (Rfree = 24.6%), respectively. ML-I is a heterodimer and belongs to the class of ribosome-inactivating proteins of type II, which consist of two chains. The A-chain has rRNA N-glycosidase activity and irreversibly inhibits eukaryotic ribosomes. The B-chain is a lectin and preferentially binds to galactose-terminated glycolipids and glycoproteins on cell membranes. Saccharide binding is performed by two binding sites in subdomains α1 and γ2 of the ML-I B-chain separated by ∼62 Å from each other. The favoured binding of galactose in subdomain α1 is achieved via hydrogen bonds connecting the 4-hydroxyl and 3-hydroxyl groups of the sugar moiety with the side chains of Asp23B, Gln36B and Lys41B and the main chain of 26B. The aromatic ring of Trp38B on top of the preferred binding pocket supports van der Waals packing of the apolar face of galactose and stabilizes the sugar–lectin complex. In the galactose-binding site II of subdomain γ2, Tyr249B provides the hydrophobic stacking and the side chains of Asp235B, Gln238B and Asn256B are hydrogen-bonding partners for galactose. In the case of the galactose-binding site I, the 2-hydroxyl group also stabilizes the sugar–protein complex, an interaction thus far rarely detected in galactose-specific lectins. Finally, a potential third low-affinity galactose-binding site in subunit β1 was identified in the present ML-I structures, in which a glycerol molecule from the cryoprotectant buffer has bound, mimicking the sugar compound. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Mikeska, R. Wacker, R. Arni, R. Singh, T.P. Mikhailov, A. Gabdoulkhakov, A. Voelter, W. Betzel, C. 2005-01-01 doi:10.1107/S1744309104031501 International Union of Crystallography The structures of mistletoe lectin I in complex with lactose and galactose reveal differences in binding by the two known sites in subdomains α1 and γ2 and suggest the presence of a third low-affinity site in subdomain β1. en RIBOSOME-INACTIVATION PROTEINS; MISTLETOE LECTIN I; SUGAR-BINDING SITES The structures of mistletoe lectin I (ML-I) from Viscum album complexed with lactose and galactose have been determined at 2.3 Å resolution and refined to R factors of 20.9% (Rfree = 23.6%) and 20.9 (Rfree = 24.6%), respectively. ML-I is a heterodimer and belongs to the class of ribosome-inactivating proteins of type II, which consist of two chains. The A-chain has rRNA N-glycosidase activity and irreversibly inhibits eukaryotic ribosomes. The B-chain is a lectin and preferentially binds to galactose-terminated glycolipids and glycoproteins on cell membranes. Saccharide binding is performed by two binding sites in subdomains α1 and γ2 of the ML-I B-chain separated by ∼62 Å from each other. The favoured binding of galactose in subdomain α1 is achieved via hydrogen bonds connecting the 4-hydroxyl and 3-hydroxyl groups of the sugar moiety with the side chains of Asp23B, Gln36B and Lys41B and the main chain of 26B. The aromatic ring of Trp38B on top of the preferred binding pocket supports van der Waals packing of the apolar face of galactose and stabilizes the sugar–lectin complex. In the galactose-binding site II of subdomain γ2, Tyr249B provides the hydrophobic stacking and the side chains of Asp235B, Gln238B and Asn256B are hydrogen-bonding partners for galactose. In the case of the galactose-binding site I, the 2-hydroxyl group also stabilizes the sugar–protein complex, an interaction thus far rarely detected in galactose-specific lectins. Finally, a potential third low-affinity galactose-binding site in subunit β1 was identified in the present ML-I structures, in which a glycerol molecule from the cryoprotectant buffer has bound, mimicking the sugar compound. text/html Mistletoe lectin I in complex with galactose and lactose reveals distinct sugar-binding properties text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 protein structure communications 17 med@iucr.org 25 1744-3091 http://scripts.iucr.org/cgi-bin/paper?za5079 The crystallization of three catalytically inactive mutants of penicillin V acylase (PVA) from Bacillus sphaericus in precursor and processed forms is reported. The mutant proteins crystallize in different primitive monoclinic space groups that are distinct from the crystal forms for the native enzyme. Directed mutants and clone constructs were designed to study the post-translational autoproteolytic processing of PVA. The catalytically inactive mutants will provide three-dimensional structures of precursor PVA forms, plus open a route to the study of enzyme–substrate complexes for this industrially important enzyme. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Chandra, P.M. Brannigan, J.A. Prabhune, A. Pundle, A. Turkenburg, J.P. Dodson, G.G. Suresh, C.G. 2005-01-01 doi:10.1107/S1744309104031227 International Union of Crystallography The production, crystallization and characterization of three inactive mutants of penicillin V acylase from B. sphaericus in their respective precursor and processed forms are reported. The space groups are different for the native enzyme and the mutants. en AUTOPROTEOLYSIS; NTN HYDROLASES; OXYANION HOLES; POST-TRANSLATIONAL PROCESSING; PRECURSOR PROTEINS; PRO-PEPTIDES The crystallization of three catalytically inactive mutants of penicillin V acylase (PVA) from Bacillus sphaericus in precursor and processed forms is reported. The mutant proteins crystallize in different primitive monoclinic space groups that are distinct from the crystal forms for the native enzyme. Directed mutants and clone constructs were designed to study the post-translational autoproteolytic processing of PVA. The catalytically inactive mutants will provide three-dimensional structures of precursor PVA forms, plus open a route to the study of enzyme–substrate complexes for this industrially important enzyme. text/html Cloning, preparation and preliminary crystallographic studies of penicillin V acylase autoproteolytic processing mutants text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 124 med@iucr.org 127 1744-3091 http://scripts.iucr.org/cgi-bin/paper?gx5030 Mycobacterium tuberculosis, the causative agent of tuberculosis, depends on the secretion of salicylate-based siderophores called mycobactins for the acquisition of extracellular iron, which is essential for the growth and virulence of the bacterium. The protein MbtI is thought to be the isochorismate synthase enzyme responsible for the conversion of chorismate to isochorismate, the first step in the salicylate production required for mycobactin biosynthesis. MbtI has been overexpressed in Escherichia coli, purified and crystallized. The crystals diffract to a maximum resolution of 1.8 Å. They belong to space group P212121, with unit-cell parameters a = 51.8, b = 163.4, c = 194.9 Å, consistent with the presence of either two, three or four molecules in the asymmetric unit. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Harrison, A.J. Ramsay, R.J. Baker, E.N. Lott, J.S. 2005-01-01 doi:10.1107/S1744309104031215 International Union of Crystallography MbtI, the putative isochorismate synthase essential for siderophore biosynthesis in M. tuberculosis, has been crystallized. Diffraction data have been collected to 1.8 Å resolution. en MYCOBACTERIUM TUBERCULOSIS; IRON ACQUISITION; SIDEROPHORE BIOSYNTHESIS Mycobacterium tuberculosis, the causative agent of tuberculosis, depends on the secretion of salicylate-based siderophores called mycobactins for the acquisition of extracellular iron, which is essential for the growth and virulence of the bacterium. The protein MbtI is thought to be the isochorismate synthase enzyme responsible for the conversion of chorismate to isochorismate, the first step in the salicylate production required for mycobactin biosynthesis. MbtI has been overexpressed in Escherichia coli, purified and crystallized. The crystals diffract to a maximum resolution of 1.8 Å. They belong to space group P212121, with unit-cell parameters a = 51.8, b = 163.4, c = 194.9 Å, consistent with the presence of either two, three or four molecules in the asymmetric unit. text/html Crystallization and preliminary X-ray crystallographic analysis of MbtI, a protein essential for siderophore biosynthesis in Mycobacterium tuberculosis text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 121 med@iucr.org 123 1744-3091 http://scripts.iucr.org/cgi-bin/paper?en5070 Inhibitors of the enzyme catechol-O-methyltransferase (COMT) are used as co-adjuvants in the therapy of Parkinson's disease. A recombinant form of the soluble cytosolic COMT from rat has been co-crystallized with a new potent inhibitor, BIA 8-176 [(3,4-dihydroxy-2-nitrophenyl)phenylmethanone], by the vapour-diffusion method using PEG 6K as precipitant. Crystals diffract to 1.6 Å resolution on a synchrotron-radiation source and belong to the monoclinic space group P21, with unit-cell parameters a = 52.77, b = 79.63, c = 61.54 Å, β = 91.14°. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Rodrigues, M.L. Bonifácio, M.J. Soares-da-Silva, P. Carrondo, M.A. Archer, M. 2005-01-01 doi:10.1107/S1744309104031197 International Union of Crystallography Catechol-O-methyltransferase has been co-crystallized with a novel inhibitor, which has potential therapeutic application in the Parkinson's disease therapy. en CATECHOL-O-METHYLTRANSFERASE; PARKINSON'S DISEASE; CATECHOL-O-METHYLTRANSFERASE INHIBITORS Inhibitors of the enzyme catechol-O-methyltransferase (COMT) are used as co-adjuvants in the therapy of Parkinson's disease. A recombinant form of the soluble cytosolic COMT from rat has been co-crystallized with a new potent inhibitor, BIA 8-176 [(3,4-dihydroxy-2-nitrophenyl)phenylmethanone], by the vapour-diffusion method using PEG 6K as precipitant. Crystals diffract to 1.6 Å resolution on a synchrotron-radiation source and belong to the monoclinic space group P21, with unit-cell parameters a = 52.77, b = 79.63, c = 61.54 Å, β = 91.14°. text/html Crystallization and preliminary X-ray diffraction studies of a catechol-O-methyltransferase/inhibitor complex text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 118 med@iucr.org 120 1744-3091 http://scripts.iucr.org/cgi-bin/paper?bw5073 The hyperthermophilic archaeon Sulfolobus solfataricus grows optimally above 353 K and can metabolize glucose and its C4 epimer galactose via a non-phosphorylative variant of the Entner–Doudoroff pathway involving catalytically promiscuous enzymes that can operate with both sugars. The initial oxidation step is catalysed by glucose dehydrogenase (SsGDH), which can utilize both NAD and NADP as cofactors. The enzyme operates with glucose and galactose at similar catalytic efficiency, while its substrate profile also includes a range of other five- and six-carbon sugars. Crystals of the 164 kDa SsGDH homotetramer have been grown under a variety of conditions. The best crystals to date diffract to 1.8 Å on a synchrotron source, have orthorhombic symmetry and belong to space group P21212. Attempts are being made to solve the structure by MAD and MR. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Theodossis, A. Milburn, C.C. Heyer, N.I. Lamble, H.J. Hough, D.W. Danson, M.J. Taylor, G.L. 2005-01-01 doi:10.1107/S174430910403101X International Union of Crystallography A glucose dehydrogenase from the hyperthermophilic archaeon S. solfataricus has been crystallized and subjected to preliminary crystallographic analysis. en SULFOLOBUS SOLFATARICUS; GLUCOSE DEHYDROGENASE; ENTNER-DOUDOROFF PATHWAY The hyperthermophilic archaeon Sulfolobus solfataricus grows optimally above 353 K and can metabolize glucose and its C4 epimer galactose via a non-phosphorylative variant of the Entner–Doudoroff pathway involving catalytically promiscuous enzymes that can operate with both sugars. The initial oxidation step is catalysed by glucose dehydrogenase (SsGDH), which can utilize both NAD and NADP as cofactors. The enzyme operates with glucose and galactose at similar catalytic efficiency, while its substrate profile also includes a range of other five- and six-carbon sugars. Crystals of the 164 kDa SsGDH homotetramer have been grown under a variety of conditions. The best crystals to date diffract to 1.8 Å on a synchrotron source, have orthorhombic symmetry and belong to space group P21212. Attempts are being made to solve the structure by MAD and MR. text/html Preliminary crystallographic studies of glucose dehydrogenase from the promiscuous Entner–Doudoroff pathway in the hyperthermophilic archaeon Sulfolobus solfataricus text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 112 med@iucr.org 115 1744-3091 http://scripts.iucr.org/cgi-bin/paper?ll5007 This is the first report of the crystallization of a sucrose phosphate synthase (SPS; EC 2.4.1.14). It also constitutes the first study of a sucrose phosphate synthase from a non-photosynthetic thermohalophilic anaerobic bacterium, Halothermothrix orenii. The purified recombinant spsA protein has been crystallized in the monoclinic space group C2, with unit-cell parameters a = 154.2, b = 47.9, c = 72.3 Å, β = 103.16°, using the hanging-drop vapour-diffusion method. The crystal diffracts X-rays to a resolution limit of 3.01 Å. Heavy-metal and halide-soaking trials are currently in progress to solve the structure. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Huynh, F. Tan, T.-C. Swaminathan, K. Patel, B.K.C. 2005-01-01 doi:10.1107/S174430910403091X International Union of Crystallography The first crystallographic study of a sucrose phosphate synthase from H. orenii, an organism that is both thermophilic and halophilic, is reported. The protein crystal diffracts X-rays to 3.01 Å. en SUCROSE PHOSPHATE SYNTHASE; HALOTHERMOTHRIX ORENII; THERMOPHILES; HALOPHILES; SUCROSE METABOLISM This is the first report of the crystallization of a sucrose phosphate synthase (SPS; EC 2.4.1.14). It also constitutes the first study of a sucrose phosphate synthase from a non-photosynthetic thermohalophilic anaerobic bacterium, Halothermothrix orenii. The purified recombinant spsA protein has been crystallized in the monoclinic space group C2, with unit-cell parameters a = 154.2, b = 47.9, c = 72.3 Å, β = 103.16°, using the hanging-drop vapour-diffusion method. The crystal diffracts X-rays to a resolution limit of 3.01 Å. Heavy-metal and halide-soaking trials are currently in progress to solve the structure. text/html Expression, purification and preliminary crystallographic analysis of sucrose phosphate synthase (SPS) from Halothermothrix orenii text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 116 med@iucr.org 117 1744-3091 http://scripts.iucr.org/cgi-bin/paper?gx5032 Disproportionating enzyme (D-enzyme; EC 2.4.1.25) is a 59 kDa protein that belongs to the α-amylase family. D-enzyme catalyses intramolecular and intermolecular transglycosylation reactions of α-1,4 glucan. A crystal of the D-­enzyme from potato was obtained by the hanging-drop vapour-diffusion method. Preliminary X-ray data showed that the crystal diffracts to 2.0 Å resolution and belongs to space group C2221, with unit-cell parameters a = 69.7, b = 120.3, c = 174.2 Å. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Imamura, K. Matsuura, T. Ye, Z. Takaha, T. Fujii, K. Kusunoki, M. Nitta, Y. 2005-01-01 doi:10.1107/S1744309104030829 International Union of Crystallography Disproportionating enzyme from potato was crystallized and preliminarily analyzed using X-ray diffraction. en DISPROPORTIONATING ENZYME; INTRAMOLECULAR AND INTERMOLECULAR TRANSGLYCOSYLATION REACTIONS Disproportionating enzyme (D-enzyme; EC 2.4.1.25) is a 59 kDa protein that belongs to the α-amylase family. D-enzyme catalyses intramolecular and intermolecular transglycosylation reactions of α-1,4 glucan. A crystal of the D-­enzyme from potato was obtained by the hanging-drop vapour-diffusion method. Preliminary X-ray data showed that the crystal diffracts to 2.0 Å resolution and belongs to space group C2221, with unit-cell parameters a = 69.7, b = 120.3, c = 174.2 Å. text/html Crystallization and preliminary X-ray crystallographic study of disproportionating enzyme from potato text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 109 med@iucr.org 111 1744-3091 http://scripts.iucr.org/cgi-bin/paper?hv5029 The arylamine N-acetyltransferase (NAT) enzymes have been found in a broad range of both eukaryotic and prokaryotic organisms. The NAT enzymes catalyse the transfer of an acetyl group from acetyl Co-enzyme A onto the terminal nitrogen of a range of arylamine, hydrazine and arylhydrazine compounds. Recently, several NAT structures have been reported from different prokaryotic sources including Salmonella typhimurium, Mycobacterium smegmatis and Pseudomonas aeruginosa. Bioinformatics analysis of the Mesorhizobium loti genome revealed two NAT paralogues, the first example of multiple NAT isoenzymes in a eubacterial organism. The M. loti NAT 1 enzyme was recombinantly expressed and purified for X-ray crystallographic studies. The purified enzyme was crystallized in 0.5 M Ca(OAc)2, 16% PEG 3350, 0.1 M Tris–HCl pH 8.5 using the sitting-drop vapour-diffusion method. A data set diffracting to 2.0 Å was collected from a single crystal at 100 K. The crystal belongs to the orthorhombic spacegroup P212121, with unit-cell parameters a = 53.2, b = 97.3, c = 114.3 Å. The structure was refined to a final free-R factor of 24.8%. The structure reveals that despite low sequence homology, M. loti NAT1 shares the common fold as reported in previous NAT structures and exhibits the same catalytic triad of residues (Cys-His-Asp) in the active site. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Holton, S.J. Dairou, J. Sandy, J. Rodrigues-Lima, F. Dupret, J.-M. Noble, M.E.M. Sim, E. 2005-01-01 doi:10.1107/S1744309104030659 International Union of Crystallography The crystal structure of a M. loti arylamine N-acetyltransferase 1 has been determined at 2.0 Å resolution. en ARYLAMINE N-ACETYLTRANSFERASE 1 The arylamine N-acetyltransferase (NAT) enzymes have been found in a broad range of both eukaryotic and prokaryotic organisms. The NAT enzymes catalyse the transfer of an acetyl group from acetyl Co-enzyme A onto the terminal nitrogen of a range of arylamine, hydrazine and arylhydrazine compounds. Recently, several NAT structures have been reported from different prokaryotic sources including Salmonella typhimurium, Mycobacterium smegmatis and Pseudomonas aeruginosa. Bioinformatics analysis of the Mesorhizobium loti genome revealed two NAT paralogues, the first example of multiple NAT isoenzymes in a eubacterial organism. The M. loti NAT 1 enzyme was recombinantly expressed and purified for X-ray crystallographic studies. The purified enzyme was crystallized in 0.5 M Ca(OAc)2, 16% PEG 3350, 0.1 M Tris–HCl pH 8.5 using the sitting-drop vapour-diffusion method. A data set diffracting to 2.0 Å was collected from a single crystal at 100 K. The crystal belongs to the orthorhombic spacegroup P212121, with unit-cell parameters a = 53.2, b = 97.3, c = 114.3 Å. The structure was refined to a final free-R factor of 24.8%. The structure reveals that despite low sequence homology, M. loti NAT1 shares the common fold as reported in previous NAT structures and exhibits the same catalytic triad of residues (Cys-His-Asp) in the active site. text/html Structure of Mesorhizobium loti arylamine N-acetyltransferase 1 text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 protein structure communications 14 med@iucr.org 16 1744-3091 http://scripts.iucr.org/cgi-bin/paper?bw5072 The trehalulose synthase (MutB) from Pseudomonas mesoacidophila MX-45, belonging to glycoside hydrolase family 13, catalyses the isomerization of sucrose to trehalulose (α-d-glucosylpyranosyl-1,1-d-fructofuranose) and isomaltulose (α-­d-glucosylpyranosyl-1,6-d-fructofuranose) as main products and glucose and fructose in residual amounts from the hydrolytic reaction. To date, a three-dimensional structure of a sucrose isomerase that produces mainly trehalulose, as is the case for MutB, has been lacking. Crystallographic studies of this 64 kDa enzyme have therefore been initiated in order to contribute to the understanding of the molecular basis of sucrose decomposition, isomerization and of the selectivity of this enzyme that leads to the formation of different products. The MutB protein has been overexpressed, purified and crystallized using the hanging-drop vapour-diffusion method. Two different crystal forms have been obtained: one diffracts X-rays to 1.6 Å resolution using synchrotron radiation and belongs to space group P1, with unit-cell parameters a = 63.8, b = 72.0, c = 82.2 Å, α = 67.5, β = 73.1, γ = 70.8°, while the other form diffracts to 1.8 Å resolution using synchrotron radiation and belongs to space group P21, with unit-cell parameters a = 63.7, b = 85.9, c = 119.7 Å, β = 97.7°. A molecular-replacement solution has been found using the structure of the isomaltulose synthase (PalI) from Klebsiella sp. LX3 as a search model. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Ravaud, S. Watzlawick, H. Haser, R. Mattes, R. Aghajari, N. 2005-01-01 doi:10.1107/S1744309104030623 International Union of Crystallography The trehalulose synthase MutB from P. mesoacidophila MX-45 has been crystallized in two different crystal forms and diffraction data have been collected to 1.6 and 1.8 Å, respectively. en ISOMERASES; HYDROLASES; TREHALULOSE SYNTHASE The trehalulose synthase (MutB) from Pseudomonas mesoacidophila MX-45, belonging to glycoside hydrolase family 13, catalyses the isomerization of sucrose to trehalulose (α-d-glucosylpyranosyl-1,1-d-fructofuranose) and isomaltulose (α-­d-glucosylpyranosyl-1,6-d-fructofuranose) as main products and glucose and fructose in residual amounts from the hydrolytic reaction. To date, a three-dimensional structure of a sucrose isomerase that produces mainly trehalulose, as is the case for MutB, has been lacking. Crystallographic studies of this 64 kDa enzyme have therefore been initiated in order to contribute to the understanding of the molecular basis of sucrose decomposition, isomerization and of the selectivity of this enzyme that leads to the formation of different products. The MutB protein has been overexpressed, purified and crystallized using the hanging-drop vapour-diffusion method. Two different crystal forms have been obtained: one diffracts X-rays to 1.6 Å resolution using synchrotron radiation and belongs to space group P1, with unit-cell parameters a = 63.8, b = 72.0, c = 82.2 Å, α = 67.5, β = 73.1, γ = 70.8°, while the other form diffracts to 1.8 Å resolution using synchrotron radiation and belongs to space group P21, with unit-cell parameters a = 63.7, b = 85.9, c = 119.7 Å, β = 97.7°. A molecular-replacement solution has been found using the structure of the isomaltulose synthase (PalI) from Klebsiella sp. LX3 as a search model. text/html Expression, purification, crystallization and preliminary X-ray crystallographic studies of the trehalulose synthase MutB from Pseudomonas mesoacidophila MX-45 text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 100 med@iucr.org 103 1744-3091 http://scripts.iucr.org/cgi-bin/paper?ll5005 No crystal structures are yet available for homologues of a predicted acetamidase/formamidase (Amds/Fmds) from the archaeon Thermoanaerobacter tengcongensis. The Amds/Fmds gene was cloned and expressed as a soluble protein in Escherichia coli. Native Amds/Fmds and its SeMet-substituted form were purified and crystallized by vapour diffusion in hanging drops at 296 K. The native crystals, which were grown in PEG 8000, belong to the monoclinic space group P21, with unit-cell parameters a = 41.23 (3), b = 152.88 (6), c = 100.26 (7) Å, β = 99.49 (3)°. The diffraction data were collected to 2.00 Å resolution using synchrotron radiation. Based on a predicted solvent content of 50%, a Matthews coefficient of 2.44 Å3 Da−1 and two main peaks in the self-rotation function, the asymmetric unit is predicted to contain two dimers of the 32 kDa native protein. MAD data were collected for the SeMet protein, but the corresponding crystals display different unit-cell parameters and appear to contain four dimers in the asymmetric unit. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Wu, G. Huang, Q. Tang, Y. Unno, H. Kusunoki, M. 2005-01-01 doi:10.1107/S1744309104030519 International Union of Crystallography A predicated acetamidase/formanidase from the archaeon T. tengcongensis and its SeMet substitute have been crystallized and undergone preliminarily crystallographic studies including MAD data collection. en ACETAMIDASE/FORMAMIDASE; THERMOANAEROBACTER TENGCONGENSIS No crystal structures are yet available for homologues of a predicted acetamidase/formamidase (Amds/Fmds) from the archaeon Thermoanaerobacter tengcongensis. The Amds/Fmds gene was cloned and expressed as a soluble protein in Escherichia coli. Native Amds/Fmds and its SeMet-substituted form were purified and crystallized by vapour diffusion in hanging drops at 296 K. The native crystals, which were grown in PEG 8000, belong to the monoclinic space group P21, with unit-cell parameters a = 41.23 (3), b = 152.88 (6), c = 100.26 (7) Å, β = 99.49 (3)°. The diffraction data were collected to 2.00 Å resolution using synchrotron radiation. Based on a predicted solvent content of 50%, a Matthews coefficient of 2.44 Å3 Da−1 and two main peaks in the self-rotation function, the asymmetric unit is predicted to contain two dimers of the 32 kDa native protein. MAD data were collected for the SeMet protein, but the corresponding crystals display different unit-cell parameters and appear to contain four dimers in the asymmetric unit. text/html Crystallization and preliminary crystallographic study of a recombinant predicted acetamidase/formamidase from the thermophile Thermoanaerobacter tengcongensis text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 106 med@iucr.org 108 1744-3091 http://scripts.iucr.org/cgi-bin/paper?en5073 Pim kinases, including Pim-1, Pim-2 and Pim-3, belong to a distinctive serine/threonine protein-kinase family. They are involved in cytokine-induced signal transduction and the development of lymphoid malignancies. Their kinase domains are highly homologous to one another, but share low sequence identity to other kinases. Specifically, there are two proline residues in the conserved hinge-region sequence ERPXPX separated by a residue that is non-conserved among Pim kinases. Full-length human Pim-1 kinase (1–313) was cloned and expressed in Escherichia coli as a GST-fusion protein and truncated to Pim-1 (14–313) by thrombin digestion during purification. The Pim-1 (14–313) protein was purified to high homogeneity and monodispersity. This protein preparation yielded small crystals in the initial screening and large crystals after optimization. The large crystals of apo Pim-1 enzyme diffracted to 2.1 Å resolution and belong to space group P65, with unit-cell parameters a = b = 95.9, c = 80.0 Å, β = 120° and one molecule per asymmetric unit. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Qian, K.C. Studts, J. Wang, L. Barringer, K. Kronkaitis, A. Peng, C. Baptiste, A. LaFrance, R. Mische, S. Farmer, B. 2005-01-01 doi:10.1107/S1744309104029963 International Union of Crystallography Pim kinases, belong to a distinctive serine/threonine protein-kinase family and are involved in cytokine-induced signal transduction and the development of lymphoid malignancies. Human Pim-1 kinase has been cloned, expressed and crystallized en PIM-1 KINASE Pim kinases, including Pim-1, Pim-2 and Pim-3, belong to a distinctive serine/threonine protein-kinase family. They are involved in cytokine-induced signal transduction and the development of lymphoid malignancies. Their kinase domains are highly homologous to one another, but share low sequence identity to other kinases. Specifically, there are two proline residues in the conserved hinge-region sequence ERPXPX separated by a residue that is non-conserved among Pim kinases. Full-length human Pim-1 kinase (1–313) was cloned and expressed in Escherichia coli as a GST-fusion protein and truncated to Pim-1 (14–313) by thrombin digestion during purification. The Pim-1 (14–313) protein was purified to high homogeneity and monodispersity. This protein preparation yielded small crystals in the initial screening and large crystals after optimization. The large crystals of apo Pim-1 enzyme diffracted to 2.1 Å resolution and belong to space group P65, with unit-cell parameters a = b = 95.9, c = 80.0 Å, β = 120° and one molecule per asymmetric unit. text/html Expression, purification, crystallization and preliminary crystallographic analysis of human Pim-­1 kinase text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 96 med@iucr.org 99 1744-3091 http://scripts.iucr.org/cgi-bin/paper?vr5024 Alzheimer's disease is thought to be triggered by production of the amyloid β (Aβ) peptide through proteolytic cleavage of the amyloid precursor protein (APP). The binding of Cu2+ to the copper-binding domain (CuBD) of APP reduces the production of Aβ in cell-culture and animal studies. It is expected that structural studies of the CuBD will lead to a better understanding of how copper binding causes Aβ depletion and will define a potential drug target. The crystallization of CuBD in two different forms suitable for structure determination is reported here. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Kong, G.K.-W. Galatis, D. Barnham, K.J. Polekhina, G. Adams, J.J. Masters, C.L. Cappai, R. Parker, M.W. McKinstry, W.J. 2005-01-01 doi:10.1107/S1744309104029744 International Union of Crystallography The binding of Cu2+ ions to the copper-binding domain of the amyloid precursor protein of Alzheimer's disease reduces the production of the amyloid β peptide, which is centrally involved in Alzheimer's disease. Structural studies of the copper-binding domain will provide a basis for structure-based drug design that might prove useful in treating this devastating disease. en ALZHEIMER'S DISEASE; AMYLOID PRECURSOR PROTEIN; COPPER BINDING Alzheimer's disease is thought to be triggered by production of the amyloid β (Aβ) peptide through proteolytic cleavage of the amyloid precursor protein (APP). The binding of Cu2+ to the copper-binding domain (CuBD) of APP reduces the production of Aβ in cell-culture and animal studies. It is expected that structural studies of the CuBD will lead to a better understanding of how copper binding causes Aβ depletion and will define a potential drug target. The crystallization of CuBD in two different forms suitable for structure determination is reported here. text/html Crystallization and preliminary crystallographic studies of the copper-binding domain of the amyloid precursor protein of Alzheimer's disease text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 93 med@iucr.org 95 1744-3091 http://scripts.iucr.org/cgi-bin/paper?en5076 Bacteriophage Mu baseplate protein gene product 44 (gp44) is an essential protein required for the assembly of viable phages. To investigate the roles of gp44 in baseplate assembly and infection, gp44 was crystallized at pH 6.0 in the presence of 20% 2-methyl-2,4-pentanediol. The crystals belong to space group R3, with unit-cell parameters a = b = 127.47, c = 63.97 Å. The crystals diffract X-­rays to at least 2.1 Å resolution and are stable in the X-ray beam and are therefore appropriate for structure determination. Native data have been collected to 2.1 Å resolution using a DIP6040 image-plate system at beamline BL44XU at the SPring-8 facility in Japan. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Kondou, Y. Kitazawa, D. Takeda, S. Yamashita, E. Mizuguchi, M. Kawano, K. Tsukihara, T. 2005-01-01 doi:10.1107/S1744309104029574 International Union of Crystallography Bacteriophage Mu baseplate protein gene product 44 was crystallized. The crystal belongs to space group R3, with unit-cell parameters a = b = 126.6, c = 64.2 Å. en GENE PRODUCT 44; BACTERIOPHAGE MU Bacteriophage Mu baseplate protein gene product 44 (gp44) is an essential protein required for the assembly of viable phages. To investigate the roles of gp44 in baseplate assembly and infection, gp44 was crystallized at pH 6.0 in the presence of 20% 2-methyl-2,4-pentanediol. The crystals belong to space group R3, with unit-cell parameters a = b = 127.47, c = 63.97 Å. The crystals diffract X-­rays to at least 2.1 Å resolution and are stable in the X-ray beam and are therefore appropriate for structure determination. Native data have been collected to 2.1 Å resolution using a DIP6040 image-plate system at beamline BL44XU at the SPring-8 facility in Japan. text/html Crystallization and preliminary X-ray analysis of gene product 44 from bacteriophage Mu text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 104 med@iucr.org 105 1744-3091 http://scripts.iucr.org/cgi-bin/paper?za5078 The putative metal-dependent hydrolase gene TTE1006 from Thermoanaerobacter tengcongensis strain MB4T (T = type strain; Genbank accession No. AE008691) was heterologously expressed in Escherichia coli. The 205-amino-acid gene product was purified and crystallized. The crystal used for data collection belongs to space group P21, with unit-cell parameters a = 85.2, b = 62.1, c = 172.4 Å, β = 104.2°. Using a synchrotron-radiation source, the resolution limit of the data reached 1.87 Å. Eight molecules were estimated to be present in the asymmetric unit, with a solvent content of 48%. Structure determination is ongoing using the multiple-wavelength anomalous diffraction (MAD) method and also the molecular-replacement (MR) method. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Liu, S. Wu, G. Huang, Q. Lai, L. Tang, Y. Unno, H. Kusunoki, M. 2005-01-01 doi:10.1107/S1744309104029392 International Union of Crystallography Crystallization and preliminary crystallographic study of a potential metal-dependent hydrolase with cyclase activity from T. tengcongensis. en METAL-DEPENDENT HYDROLASES The putative metal-dependent hydrolase gene TTE1006 from Thermoanaerobacter tengcongensis strain MB4T (T = type strain; Genbank accession No. AE008691) was heterologously expressed in Escherichia coli. The 205-amino-acid gene product was purified and crystallized. The crystal used for data collection belongs to space group P21, with unit-cell parameters a = 85.2, b = 62.1, c = 172.4 Å, β = 104.2°. Using a synchrotron-radiation source, the resolution limit of the data reached 1.87 Å. Eight molecules were estimated to be present in the asymmetric unit, with a solvent content of 48%. Structure determination is ongoing using the multiple-wavelength anomalous diffraction (MAD) method and also the molecular-replacement (MR) method. text/html Expression, purification, crystallization and preliminary crystallographic study of a potential metal-dependent hydrolase with cyclase activity from Thermoanaerobacter tengcongensis text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 90 med@iucr.org 92 1744-3091 http://scripts.iucr.org/cgi-bin/paper?za5077 A lectin from Canavalia maritima seeds (ConM) was purified and submitted to crystallization experiments. The best crystals were obtained using the vapour-diffusion method at a constant temperature of 293 K and grew in 7 d. A complete structural data set was collected to 2.1 Å resolution using a synchrotron-radiation source. The ConM crystal belongs to the orthorhombic space group P21212, with unit-cell parameters a = 67.15, b = 70.90, c = 97.37 Å. A molecular-replacement search found a solution with a correlation coefficient of 69.2% and an R factor of 42.5%. Crystallographic refinement is under way. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Gadelha, C.A.A. Moreno, F.B.M.B. Santi-Gadelha, T. Cajazeiras, J.B. da Rocha, B.A.M. Rustiguel, J.K.R. Freitas, B.T. Canduri, F. Delatorre, P. de Azevedo, W.F. Cavada, B.S. 2005-01-01 doi:10.1107/S1744309104029197 International Union of Crystallography A lectin from C. maritima was crystallized using the vapour-diffusion method and crystals diffracted to 2.1 Å resolution. A molecular-replacement search found a solution with a correlation coefficient of 69.2% and an R factor of 42.5%, refinement is in progress. en LECTINS; CANAVALIA MARITIMA A lectin from Canavalia maritima seeds (ConM) was purified and submitted to crystallization experiments. The best crystals were obtained using the vapour-diffusion method at a constant temperature of 293 K and grew in 7 d. A complete structural data set was collected to 2.1 Å resolution using a synchrotron-radiation source. The ConM crystal belongs to the orthorhombic space group P21212, with unit-cell parameters a = 67.15, b = 70.90, c = 97.37 Å. A molecular-replacement search found a solution with a correlation coefficient of 69.2% and an R factor of 42.5%. Crystallographic refinement is under way. text/html Crystallization and preliminary X-ray diffraction analysis of a lectin from Canavalia maritima seeds text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 87 med@iucr.org 89 1744-3091 http://scripts.iucr.org/cgi-bin/paper?ll5004 The reaction center–light-harvesting 1 (RC–LH1) core complex is the photosynthetic apparatus in the membrane of the purple photosynthetic bacterium Rhodopseudomonas viridis. The RC is surrounded by an LH1 complex that is constituted of oligomers of three types of apoproteins (α, β and γ chains) with associated bacteriochlorophyll bs and carotenoid. It has been crystallized by the sitting-drop vapour-diffusion method. A promising crystal diffracted to beyond 8.0 Å resolution. It belonged to space group P1, with unit-cell parameters a = 141.4, b = 136.9, c = 185.3 Å, α = 104.6, β = 94.0, γ = 110.7°. A Patterson function calculated using data between 15.0 and 8.0 Å resolution suggested that the LH1 complex is distributed with quasi-16-fold rotational symmetry around the RC. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Saijo, S. Sato, T. Kumasaka, T. Tanaka, N. Harata, K. Odahara, T. 2005-01-01 doi:10.1107/S1744309104028945 International Union of Crystallography The reaction center–light-harvesting 1 core complex from R. viridis was crystallized and X-ray diffraction data were collected to 8.0 Å resolution. en PHOTOSYNTHESIS; REACTION CENTER-LIGHT-HARVESTING 1 CORE COMPLEX; RHODOPSEUDOMONAS VIRIDIS; INTEGRAL MEMBRANE PROTEINS The reaction center–light-harvesting 1 (RC–LH1) core complex is the photosynthetic apparatus in the membrane of the purple photosynthetic bacterium Rhodopseudomonas viridis. The RC is surrounded by an LH1 complex that is constituted of oligomers of three types of apoproteins (α, β and γ chains) with associated bacteriochlorophyll bs and carotenoid. It has been crystallized by the sitting-drop vapour-diffusion method. A promising crystal diffracted to beyond 8.0 Å resolution. It belonged to space group P1, with unit-cell parameters a = 141.4, b = 136.9, c = 185.3 Å, α = 104.6, β = 94.0, γ = 110.7°. A Patterson function calculated using data between 15.0 and 8.0 Å resolution suggested that the LH1 complex is distributed with quasi-16-fold rotational symmetry around the RC. text/html Crystallization and preliminary X-ray studies on the reaction center–light-harvesting 1 core complex from Rhodopseudomonas viridis text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 83 med@iucr.org 86 1744-3091 http://scripts.iucr.org/cgi-bin/paper?ll5003 Four crystal forms have been grown and characterized by X-ray diffraction of a Bence-Jones protein collected from the urine of a multiple myeloma patient more than 40 years ago. Closely related tetragonal and orthorhombic forms belonging to space groups P43212 and P212121, with unit-cell parameters a = b = 68.7, c = 182.1 and a = 67.7, b = 69.4, c = 87.3 Å, diffract to 1.5 and 1.9 Å, respectively. Two closely related trigonal forms, both belonging to space group P3121 with unit-cell parameters a = b = 154.3 Å but differing by a doubling of the c axis, one 46.9 Å and the other 94.0 Å, diffract to 2.9 and 2.6 Å resolution, respectively. The trigonal crystal of short c-axis length shows a positive indication of twinning. The trigonal crystal of longer c axis, which appeared only after eight months of incubation at room temperature, is likely to be composed of proteolytically degraded molecules and unlike the other crystal forms contains two entire Bence-Jones dimers in the asymmetric unit. This latter crystal form may shed some light on the formation of fibrils common to certain storage diseases. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Makino, D.L. Henschen-Edman, A.H. McPherson, A. 2005-01-01 doi:10.1107/S1744309104028532 International Union of Crystallography Four crystal forms have been grown and characterized by X-ray diffraction of a Bence-Jones protein collected from the urine of a multiple myeloma patient more than 40 y ago. The trigonal crystal form may shed some light on the formation of fibrils common to certain storage diseases. en STORAGE DISEASES; MULTIPLE MYELOMA; IMMUNOGLOBULINS; PROTEOLYSIS; TWINNING Four crystal forms have been grown and characterized by X-ray diffraction of a Bence-Jones protein collected from the urine of a multiple myeloma patient more than 40 years ago. Closely related tetragonal and orthorhombic forms belonging to space groups P43212 and P212121, with unit-cell parameters a = b = 68.7, c = 182.1 and a = 67.7, b = 69.4, c = 87.3 Å, diffract to 1.5 and 1.9 Å, respectively. Two closely related trigonal forms, both belonging to space group P3121 with unit-cell parameters a = b = 154.3 Å but differing by a doubling of the c axis, one 46.9 Å and the other 94.0 Å, diffract to 2.9 and 2.6 Å resolution, respectively. The trigonal crystal of short c-axis length shows a positive indication of twinning. The trigonal crystal of longer c axis, which appeared only after eight months of incubation at room temperature, is likely to be composed of proteolytically degraded molecules and unlike the other crystal forms contains two entire Bence-Jones dimers in the asymmetric unit. This latter crystal form may shed some light on the formation of fibrils common to certain storage diseases. text/html Four crystal forms of a Bence-Jones protein text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 79 med@iucr.org 82 1744-3091 http://scripts.iucr.org/cgi-bin/paper?ll5002 Group I introns are catalytic RNAs that are capable of performing a variety of phosphotransesterification reactions including self-splicing and RNA cleavage. The reactions are efficient, accurate and dependent only on the presence of guanosine-nucleotide substrate and sufficient magnesium ion to stabilize the structure of the RNA. To understand how the group I intron active-site facilitates catalysis, crystals of a 242-nucleotide ribozyme bound to a four-nucleotide product RNA have been produced that diffract to 3.6 Å resolution. The space group of these crystals is I212121 and the unit-cell parameters are a = 94.6, b = 141.0, c = 210.9 Å. A single heavy-atom derivative has been synthesized by covalent modification of the product RNA with iodine. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Chase, E. Golden, B.L. 2005-01-01 doi:10.1107/S1744309104028337 International Union of Crystallography A group I self-splicing RNA has been synthesized and cocrystallized with a four-nucleotide product RNA. Iodination of the product RNA produces a heavy-atom derivative suitable for structure determination. en CATALYTIC RNA; GROUP I INTRON; RNA SPLICING; RNA STRUCTURE Group I introns are catalytic RNAs that are capable of performing a variety of phosphotransesterification reactions including self-splicing and RNA cleavage. The reactions are efficient, accurate and dependent only on the presence of guanosine-nucleotide substrate and sufficient magnesium ion to stabilize the structure of the RNA. To understand how the group I intron active-site facilitates catalysis, crystals of a 242-nucleotide ribozyme bound to a four-nucleotide product RNA have been produced that diffract to 3.6 Å resolution. The space group of these crystals is I212121 and the unit-cell parameters are a = 94.6, b = 141.0, c = 210.9 Å. A single heavy-atom derivative has been synthesized by covalent modification of the product RNA with iodine. text/html Crystallization and preliminary diffraction analysis of a group I ribozyme from bacteriophage Twort text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 71 med@iucr.org 74 1744-3091 http://scripts.iucr.org/cgi-bin/paper?bw5066 Pantothenate kinase is an essential enzyme in the bacterial life cycle. It catalyzes the phosphorylation of pantothenate (vitamin B5) to 4′-phosphopantothenate, the first step in the coenzyme A biosynthetic pathway. The enzyme from Mycobacterium tuberculosis, MW 35.7 kDa, has been cloned, expressed, purified and crystallized in two different trigonal crystal forms, both belonging to space group P3121. Two complete data sets of resolution 2.5 Å (form I) and 2.9 Å (form II) from crystals with unit-cell parameters a = b = 78.3, c = 115.45 Å and a = b = 107.63, c = 89.85 Å, respectively, were collected at room temperature on a home X-ray source. Structures of both crystal forms were solved for one subunit in the asymmetric unit by molecular replacement. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Das, S. Kumar, P. Bhor, V. Surolia, A. Vijayan, M. 2005-01-01 doi:10.1107/S1744309104028040 International Union of Crystallography Pantothenate kinase, the first enzyme of the universal coenzyme A biosynthetic pathway, from M. tuberculosis H37Rv has been cloned, expressed, purified and X-ray analysed in two different crystal forms. en PANTOTHENATE KINASE Pantothenate kinase is an essential enzyme in the bacterial life cycle. It catalyzes the phosphorylation of pantothenate (vitamin B5) to 4′-phosphopantothenate, the first step in the coenzyme A biosynthetic pathway. The enzyme from Mycobacterium tuberculosis, MW 35.7 kDa, has been cloned, expressed, purified and crystallized in two different trigonal crystal forms, both belonging to space group P3121. Two complete data sets of resolution 2.5 Å (form I) and 2.9 Å (form II) from crystals with unit-cell parameters a = b = 78.3, c = 115.45 Å and a = b = 107.63, c = 89.85 Å, respectively, were collected at room temperature on a home X-ray source. Structures of both crystal forms were solved for one subunit in the asymmetric unit by molecular replacement. text/html Expression, purification, crystallization and preliminary X-ray crystallographic analysis of pantothenate kinase from Mycobacterium tuberculosis text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 65 med@iucr.org 67 1744-3091 http://scripts.iucr.org/cgi-bin/paper?vr5029 Endo-1,3-β-glucanases hydrolyze internal 1,3-β-glucosyl linkages. The endo-1,3-β-glucanase from Arthrobacter sp. was crystallized by the hanging-drop vapour-diffusion method. The crystals belonged to space group P41, with unit-cell parameters a = 71.31, c = 60.07 Å, and contained one molecule per asymmetric unit. The Matthews coefficient (VM) and the solvent content were 2.35 Å3 Da−1 and 47.63%, respectively. Diffraction data were collected to a resolution of 1.66 Å at SPring-8 using a MAR CCD area detector and gave a data set with an overall Rmerge of 5.4% and a completeness of 99.4%. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Pang, Z. Kang, Y.-N. Ban, M. Oda, M. Kobayashi, R. Ohnishi, M. Mikami, B. 2005-01-01 doi:10.1107/S1744309104027915 International Union of Crystallography Endo-1,3-β-glucanase from Arthrobacter sp. has been crystallized and X-ray diffraction data have been collected to 1.66 Å resolution. en ENDO-1,3-[BETA]-GLUCANASE Endo-1,3-β-glucanases hydrolyze internal 1,3-β-glucosyl linkages. The endo-1,3-β-glucanase from Arthrobacter sp. was crystallized by the hanging-drop vapour-diffusion method. The crystals belonged to space group P41, with unit-cell parameters a = 71.31, c = 60.07 Å, and contained one molecule per asymmetric unit. The Matthews coefficient (VM) and the solvent content were 2.35 Å3 Da−1 and 47.63%, respectively. Diffraction data were collected to a resolution of 1.66 Å at SPring-8 using a MAR CCD area detector and gave a data set with an overall Rmerge of 5.4% and a completeness of 99.4%. text/html Crystallization and preliminary crystallographic analysis of endo-1,3-β-glucanase from Arthrobacter sp. text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 68 med@iucr.org 70 1744-3091 http://scripts.iucr.org/cgi-bin/paper?en5068 The crystallization and preliminary crystallographic analysis of agkicetin-C, a well known platelet glycoprotein Ib (GPIb) antagonist from the venom of Deinagkistrodon acutus found in Anhui Province, China is reported. Crystals of agkicetin-C suitable for structure determination were obtained from 1.8 M ammonium sulfate, 40 mM MES pH 6.5 with 2%(v/v) PEG 400. Interestingly, low buffer concentrations of MES seem to be necessary for crystal growth. The crystals of agkicetin-C belong to space group C2, with unit-cell parameters a = 177.5, b = 97.7, c = 106.8 Å, β = 118.5°, and diffract to 2.4 Å resolution. Solution of the phase problem by the molecular-replacement method shows that there are four agkicetin-C molecules in the asymmetric unit, with a VM value of 3.4 Å3 Da−1, which corresponds to a high solvent content of approximately 64%. Self-rotation function calculations show a single well defined non-crystallographic twofold axis with features that may represent additional elements of non-crystallographic symmetry. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Xu, G. Huang, Q. Teng, M. Liu, P. Dong, Y. Niu, L. 2005-01-01 doi:10.1107/S1744309104027241 International Union of Crystallography The crystallization and preliminary crystallographic analysis of agkicetin-C, a well known platelet glycoprotein Ib (GPIb) antagonist from the venom of Deinagkistrodon acutus found in Anhui Province, China is reported. en SNAKE VENOMS; C-TYPE LECTINS; AGKICETIN-C The crystallization and preliminary crystallographic analysis of agkicetin-C, a well known platelet glycoprotein Ib (GPIb) antagonist from the venom of Deinagkistrodon acutus found in Anhui Province, China is reported. Crystals of agkicetin-C suitable for structure determination were obtained from 1.8 M ammonium sulfate, 40 mM MES pH 6.5 with 2%(v/v) PEG 400. Interestingly, low buffer concentrations of MES seem to be necessary for crystal growth. The crystals of agkicetin-C belong to space group C2, with unit-cell parameters a = 177.5, b = 97.7, c = 106.8 Å, β = 118.5°, and diffract to 2.4 Å resolution. Solution of the phase problem by the molecular-replacement method shows that there are four agkicetin-C molecules in the asymmetric unit, with a VM value of 3.4 Å3 Da−1, which corresponds to a high solvent content of approximately 64%. Self-rotation function calculations show a single well defined non-crystallographic twofold axis with features that may represent additional elements of non-crystallographic symmetry. text/html Crystallization and preliminary X-ray crystallographic analysis of agkicetin-C from Deinagkistrodon acutus venom text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 75 med@iucr.org 78 1744-3091 http://scripts.iucr.org/cgi-bin/paper?bw5068 Bri3 is a recently identified proline-rich transmembrane polypeptide up-regulated during TNF-mediated inflammation and immunity. The polyproline-rich N-terminal (residues 1–60) domain of Bri3 was affinity-purified to homogeneity as a glutathione-S-transferase (GST) fusion protein. Crystals were obtained in ∼3 d by the equilibrium vapour-diffusion method from a solution containing 1.5–2.2 M ammonium sulfate and 0.1 M bis-tris pH 6.0. The crystals belong to space group P43212, with unit-cell parameters a = b = 91.66, c = 57.53 Å. An X-ray data set was collected to 1.6 Å resolution using synchrotron radiation, with an Rsym of 0.058 and a completeness of 95.3%. There is one molecule of the fusion protein in the asymmetric unit, which corresponds to ∼35% solvent content. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Ye, Q. Singh, V.K. Blonde, J.D. Jia, Z. 2005-01-01 doi:10.1107/S1744309104026739 International Union of Crystallography The crystallization of the polyproline-rich polypeptide from human Bri3 overexpressed as a GST-fusion protein in Escherichia coli is presented. en BRI3; TRANSMEMBRANE PROTEINS Bri3 is a recently identified proline-rich transmembrane polypeptide up-regulated during TNF-mediated inflammation and immunity. The polyproline-rich N-terminal (residues 1–60) domain of Bri3 was affinity-purified to homogeneity as a glutathione-S-transferase (GST) fusion protein. Crystals were obtained in ∼3 d by the equilibrium vapour-diffusion method from a solution containing 1.5–2.2 M ammonium sulfate and 0.1 M bis-tris pH 6.0. The crystals belong to space group P43212, with unit-cell parameters a = b = 91.66, c = 57.53 Å. An X-ray data set was collected to 1.6 Å resolution using synchrotron radiation, with an Rsym of 0.058 and a completeness of 95.3%. There is one molecule of the fusion protein in the asymmetric unit, which corresponds to ∼35% solvent content. text/html Crystallization and preliminary X-ray analysis of the GST-fused human Bri3 N-terminal domain text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 62 med@iucr.org 64 1744-3091 http://scripts.iucr.org/cgi-bin/paper?bw5070 The vacuole-type ATPases in eukaryotic cells translocate protons across various biological membranes including the vacuolar membrane by consuming ATP molecules. The E subunit of the multisubunit complex V-ATPase from Pyrococcus horikoshii OT3, which has a molecular weight of 22.88 kDa, has been cloned, overexpressed in Escherichia coli, purified and crystallized by the microbatch method using PEG 4000 as a precipitant at 296 K. A data set to 1.85 Å resolution with 98.8% completeness and an Rmerge of 6.5% was collected from a single flash-cooled crystal using synchrotron radiation. The crystal belonged to the orthorhombic space group P212121, with unit-cell parameters a = 52.196, b = 55.317, c = 77.481 Å, and is most likely to contain one molecule per asymmetric unit. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Lokanath, N.K. Ukita, Y. Sugahara, M. Kunishima, N. 2005-01-01 doi:10.1107/S1744309104026430 International Union of Crystallography The E subunit of vacuole-type ATPase from P. horikoshii OT3 was overexpressed, purified and crystallized. The native crystals diffracted X-rays to 1.85 Å resolution. en VACUOLE-TYPE ATPASE SUBUNIT E; PYROCOCCUS HORIKOSHII OT3 The vacuole-type ATPases in eukaryotic cells translocate protons across various biological membranes including the vacuolar membrane by consuming ATP molecules. The E subunit of the multisubunit complex V-ATPase from Pyrococcus horikoshii OT3, which has a molecular weight of 22.88 kDa, has been cloned, overexpressed in Escherichia coli, purified and crystallized by the microbatch method using PEG 4000 as a precipitant at 296 K. A data set to 1.85 Å resolution with 98.8% completeness and an Rmerge of 6.5% was collected from a single flash-cooled crystal using synchrotron radiation. The crystal belonged to the orthorhombic space group P212121, with unit-cell parameters a = 52.196, b = 55.317, c = 77.481 Å, and is most likely to contain one molecule per asymmetric unit. text/html Purification, crystallization and preliminary crystallographic analysis of the vacuole-type ATPase subunit E from Pyrococcus horikoshii OT3 text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 56 med@iucr.org 58 1744-3091 http://scripts.iucr.org/cgi-bin/paper?gx5029 In the cell, propionate is mainly formed during β-oxidation of odd-numbered carbon-chain fatty acids, fermentation of carbohydrates and degradation of the amino acids threonine, valine, isoleucine and methionine. Recently, it has been shown that l-threonine is non-oxidatively cleaved to propionate via 2-­ketobutyrate. The last step in this process, conversion of propionyl phosphate and ADP to propionate and ATP, is catalysed by propionate kinase (EC 2.7.1.–). Here, the cloning of propionate kinase (molecular weight 44 kDa) from Salmonella typhimurium with an N-terminal hexahistidine affinity tag and its overexpression in Escherichia coli are reported. Purified propionate kinase was found to cocrystallize with ADP in the hanging-drop vapour-diffusion and microbatch methods. Crystals belong to space group P3121 or P3221, with unit-cell parameters a = b = 111.47, c = 66.52 Å. A complete data set to 2.2 Å resolution has been collected using an image-plate detector system mounted on a rotating-anode X-ray generator. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Simanshu, D.K. Murthy, M.R.N. 2005-01-01 doi:10.1107/S1744309104026429 International Union of Crystallography Propionate kinase (TdcD) from S. typhimurium has been expressed, purified and crystallized. A diffraction data set has been collected to 2.2 Å resolution. en TDCD; PROPIONATE KINASES; ACETATE KINASES; L-THREONINE METABOLISM In the cell, propionate is mainly formed during β-oxidation of odd-numbered carbon-chain fatty acids, fermentation of carbohydrates and degradation of the amino acids threonine, valine, isoleucine and methionine. Recently, it has been shown that l-threonine is non-oxidatively cleaved to propionate via 2-­ketobutyrate. The last step in this process, conversion of propionyl phosphate and ADP to propionate and ATP, is catalysed by propionate kinase (EC 2.7.1.–). Here, the cloning of propionate kinase (molecular weight 44 kDa) from Salmonella typhimurium with an N-terminal hexahistidine affinity tag and its overexpression in Escherichia coli are reported. Purified propionate kinase was found to cocrystallize with ADP in the hanging-drop vapour-diffusion and microbatch methods. Crystals belong to space group P3121 or P3221, with unit-cell parameters a = b = 111.47, c = 66.52 Å. A complete data set to 2.2 Å resolution has been collected using an image-plate detector system mounted on a rotating-anode X-ray generator. text/html Cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of propionate kinase (TdcD) from Salmonella typhimurium text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 52 med@iucr.org 55 1744-3091 http://scripts.iucr.org/cgi-bin/paper?bw5065 In living systems, the intramolecular cyclization of N-terminal glutamine residues is accomplished by glutaminyl cyclase enzymes (EC 2.3.2.5). While in mammals these enzymes are involved in the synthesis of hormonal and neurotransmitter peptides, the physiological role played by the corresponding plant enzymes still remains to be unravelled. Papaya glutaminyl cyclase (PQC), a 33 kDa enzyme found in the latex of the tropical tree Carica papaya, displays an exceptional resistance to chemical and thermal denaturation as well as to proteolysis. In order to elucidate its enzymatic mechanism and to gain insights into the structural determinants underlying its remarkable stability, PQC was isolated from papaya latex, purified and crystallized by the hanging-drop vapour-diffusion method. The crystals belong to the orthorhombic space group P212121, with unit-cell parameters a = 62.82, b = 81.23, c = 108.17 Å and two molecules per asymmetric unit. Diffraction data have been collected at ESRF beamline BM14 and processed to a resolution of 1.7 Å. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Azarkan, M. Clantin, B. Bompard, C. Belrhali, H. Baeyens-Volant, D. Looze, Y. Villeret, V. Wintjens, R. 2005-01-01 doi:10.1107/S1744309104025904 International Union of Crystallography The glutaminyl cyclase isolated from C. papaya latex has been crystallized using the hanging-drop method. Diffraction data have been collected at ESRF beamline BM14 and processed to 1.7 Å resolution. en GLUTAMINYL CYCLASE In living systems, the intramolecular cyclization of N-terminal glutamine residues is accomplished by glutaminyl cyclase enzymes (EC 2.3.2.5). While in mammals these enzymes are involved in the synthesis of hormonal and neurotransmitter peptides, the physiological role played by the corresponding plant enzymes still remains to be unravelled. Papaya glutaminyl cyclase (PQC), a 33 kDa enzyme found in the latex of the tropical tree Carica papaya, displays an exceptional resistance to chemical and thermal denaturation as well as to proteolysis. In order to elucidate its enzymatic mechanism and to gain insights into the structural determinants underlying its remarkable stability, PQC was isolated from papaya latex, purified and crystallized by the hanging-drop vapour-diffusion method. The crystals belong to the orthorhombic space group P212121, with unit-cell parameters a = 62.82, b = 81.23, c = 108.17 Å and two molecules per asymmetric unit. Diffraction data have been collected at ESRF beamline BM14 and processed to a resolution of 1.7 Å. text/html Crystallization and preliminary X-ray diffraction studies of the glutaminyl cyclase from Carica papaya latex text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 59 med@iucr.org 61 1744-3091 http://scripts.iucr.org/cgi-bin/paper?bw5067 The high-affinity calcium-mediated type II cohesin–dockerin interaction is responsible for the attachment of the multi-enzyme cellulose-degrading complex, termed the cellulosome, to the cell surface of the thermophilic anaerobe Clostridium thermocellum. A trimodular 40 kDa complex comprising the SdbA type II cohesin and the the CipA type II dockerin–X module modular pair from the cellulosome of C. thermocellum has been crystallized. The crystals belong to space group P212121, with unit-cell parameters a = 45.21, b = 52.34, c = 154.69 Å. The asymmetric unit contains one molecule of the protein complex and native and selenomethionine-derivative crystals diffracted to 2.1 and 2.0 Å, respectively. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Adams, J.J. Pal, G. Yam, K. Spencer, H.L. Jia, Z. Smith, S.P. 2005-01-01 doi:10.1107/S1744309104025837 International Union of Crystallography A trimodular complex comprising the type II cohesin–dockerin interaction from the cellulosome of C. thermocellum has been purified and crystallized by the hanging-drop vapour-diffusion method. A native crystal and a selenomethionine derivative have been analyzed using X-ray diffraction. en TYPE II COHESIN; DOCKERIN; CELLULOSOME The high-affinity calcium-mediated type II cohesin–dockerin interaction is responsible for the attachment of the multi-enzyme cellulose-degrading complex, termed the cellulosome, to the cell surface of the thermophilic anaerobe Clostridium thermocellum. A trimodular 40 kDa complex comprising the SdbA type II cohesin and the the CipA type II dockerin–X module modular pair from the cellulosome of C. thermocellum has been crystallized. The crystals belong to space group P212121, with unit-cell parameters a = 45.21, b = 52.34, c = 154.69 Å. The asymmetric unit contains one molecule of the protein complex and native and selenomethionine-derivative crystals diffracted to 2.1 and 2.0 Å, respectively. text/html Purification and crystallization of a trimodular complex comprising the type II cohesin–dockerin interaction from the cellulosome of Clostridium thermocellum text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 46 med@iucr.org 48 1744-3091 http://scripts.iucr.org/cgi-bin/paper?pu5065 Phosphoribosylpyrophosphate synthases (PRS; EC 2.7.6.1) are enzymes that are of central importance in several metabolic pathways in all cells. The sugar cane PRS enzyme contains 328 amino acids with a molecular weight of 36.6 kDa and represents the first plant PRS to be crystallized, as well as the first phosphate-independent PRS to be studied in molecular detail. Sugar cane PRS was overexpressed in Escherichia coli, purified and crystallized using the hanging-drop vapour-diffusion method. Using X-ray diffraction experiments it was determined that the crystals belong to the orthorhombic system, with space group P21212 and unit-cell parameters a = 213.2, b = 152.6, c = 149.3 Å. The crystals diffract to a maximum resolution of 3.3 Å and a complete data set to 3.5 Å resolution was collected and analysed. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Napolitano, H.B. Sculaccio, S.A. Thiemann, O.H. Oliva, G. 2005-01-01 doi:10.1107/S1744309104025825 International Union of Crystallography X-ray diffraction data have been collected from crystals of recombinant sugar cane phosphoribosylpyrophosphate synthase (PRS) and analysis has revealed its quaternary structure, localizing this PRS into the class of enzymes forming an hexameric oligomer of 223 kDa. en PHOSPHORIBOSYLPYROPHOSPHATE; PRPP SYNTHASE; SUGAR CANE Phosphoribosylpyrophosphate synthases (PRS; EC 2.7.6.1) are enzymes that are of central importance in several metabolic pathways in all cells. The sugar cane PRS enzyme contains 328 amino acids with a molecular weight of 36.6 kDa and represents the first plant PRS to be crystallized, as well as the first phosphate-independent PRS to be studied in molecular detail. Sugar cane PRS was overexpressed in Escherichia coli, purified and crystallized using the hanging-drop vapour-diffusion method. Using X-ray diffraction experiments it was determined that the crystals belong to the orthorhombic system, with space group P21212 and unit-cell parameters a = 213.2, b = 152.6, c = 149.3 Å. The crystals diffract to a maximum resolution of 3.3 Å and a complete data set to 3.5 Å resolution was collected and analysed. text/html Preliminary crystallographic analysis of sugar cane phosphoribosylpyrophosphate synthase text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography 1744-3091 crystallization communications 49 med@iucr.org 51 1744-3091 http://scripts.iucr.org/cgi-bin/paper?en5071 The venom of the common Indian krait (Bungarus caeruleus) contains about a dozen isoforms of phospholipase A2 (PLA2), which exist in different oligomeric forms as well as in complexes with low-molecular-weight ligands. The basic objective of multimerization and complexation is either to inactivate PLA2 in the venom for long-term storage, to generate a new PLA2 function or to make a more lethal assembly. The current isoform was isolated from the venom of B. caeruleus. Dynamic light-scattering studies indicated the presence of a stable trimeric association of this PLA2. Its primary sequence was determined by cDNA cloning. The purified protein was crystallized with 2.8 M NaCl as a precipitating agent using the sitting-drop vapour-diffusion method. The crystals belonged to the monoclinic space group C2, with unit-cell parameters a = 80.9, b = 80.5, c = 57.1 Å, β = 90.3°. The structure was refined to a final R factor of 0.198. This is a novel trimeric PLA2 structure in which the central pore formed by the association of three molecules is filled with water molecules. The interactions across the pore take place via multiple water bridges primarily to the side chains of Arg, Lys and Thr residues. Approximately 12% of the total solvent-accessible surface area is buried in the core of the trimer. The active sites of all three molecules are located on the surface and are fully exposed to the solvent, resulting in a highly potent enzymatic unit. Copyright (c) 2005 International Union of Crystallography urn:issn:1744-3091 Singh, G. Gourinath, S. Saravanan, K. Sharma, S. Bhanumathi, S. Betzel, C. Srinivasan, A. Singh, T.P. 2005-01-01 doi:10.1107/S1744309104025503 International Union of Crystallography The structure of a novel trimeric isoform of phospholipase A2 has been determined at 2.5 Å resolution. The trimer formation occurs in such a way that the active sites of all the three molecules are fully exposed to the solvent, making the trimer a highly potent enzymatic unit. en PHOSPHOLIPASE A2; OLIGOMERIZATION; AGGREGATION The venom of the common Indian krait (Bungarus caeruleus) contains about a dozen isoforms of phospholipase A2 (PLA2), which exist in different oligomeric forms as well as in complexes with low-molecular-weight ligands. The basic objective of multimerization and complexation is either to inactivate PLA2 in the venom for long-term storage, to generate a new PLA2 function or to make a more lethal assembly. The current isoform was isolated from the venom of B. caeruleus. Dynamic light-scattering studies indicated the presence of a stable trimeric association of this PLA2. Its primary sequence was determined by cDNA cloning. The purified protein was crystallized with 2.8 M NaCl as a precipitating agent using the sitting-drop vapour-diffusion method. The crystals belonged to the monoclinic space group C2, with unit-cell parameters a = 80.9, b = 80.5, c = 57.1 Å, β = 90.3°. The structure was refined to a final R factor of 0.198. This is a novel trimeric PLA2 structure in which the central pore formed by the association of three molecules is filled with water molecules. The interactions across the pore take place via multiple water bridges primarily to the side chains of Arg, Lys and Thr residues. Approximately 12% of the total solvent-accessible surface area is buried in the core of the trimer. The active sites of all three molecules are located on the surface and are fully exposed to the solvent, resulting in a highly potent enzymatic unit. text/html Sequence-induced trimerization of phospholipase A2: structure of a trimeric isoform of PLA2 from common krait (Bungarus caeruleus) at 2.5 Å resolution text 1 61 2005-01-01 Acta Crystallographica Section F: Structural Biology and Crystallization Communications Copyright (c) 2005 International Union of Crystallography