Acta Crystallographica Section D
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Acta Crystallographica Section D: Biological Crystallography welcomes the submission of articles covering any aspect of structural biology, with a particular emphasis on the structures of biological macromolecules and the methods used to determine them. Reports on new protein structures are particularly encouraged, as are structure-function articles that could include crystallographic binding studies, or structural analysis of mutants or other modified forms of a known protein structure. The key criterion is that such articles should present new insights into biology, chemistry or structure. Articles on crystallographic methods should be oriented towards biological crystallography, and may include new approaches to any aspect of structure determination or analysis. Articles on the crystallization of biological molecules will be accepted providing that these focus on new methods or other features that are of general importance or applicability.enCopyright (c) 2024 International Union of Crystallography2024-03-15International Union of CrystallographyInternational Union of Crystallographyhttp://journals.iucr.orgurn:issn:2059-7983Acta Crystallographica Section D: Biological Crystallography welcomes the submission of articles covering any aspect of structural biology, with a particular emphasis on the structures of biological macromolecules and the methods used to determine them. Reports on new protein structures are particularly encouraged, as are structure-function articles that could include crystallographic binding studies, or structural analysis of mutants or other modified forms of a known protein structure. The key criterion is that such articles should present new insights into biology, chemistry or structure. Articles on crystallographic methods should be oriented towards biological crystallography, and may include new approaches to any aspect of structure determination or analysis. Articles on the crystallization of biological molecules will be accepted providing that these focus on new methods or other features that are of general importance or applicability.text/htmlActa Crystallographica Section D: Structural Biology, Volume 80, Part 4, 2024textweekly12002-01-01T00:00+00:004802024-03-15Copyright (c) 2024 International Union of CrystallographyActa Crystallographica Section D: Structural Biology147urn:issn:2059-7983med@iucr.orgMarch 20242024-03-15Acta Crystallographica Section Dhttp://journals.iucr.org/logos/rss10d.gif
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Still imageStructural determination and modeling of ciliary microtubules
http://scripts.iucr.org/cgi-bin/paper?ih5006
The axoneme, a microtubule-based array at the center of every cilium, has been the subject of structural investigations for decades, but only recent advances in cryo-EM and cryo-ET have allowed a molecular-level interpretation of the entire complex to be achieved. The unique properties of the nine doublet microtubules and central pair of singlet microtubules that form the axoneme, including the highly decorated tubulin lattice and the docking of massive axonemal complexes, provide opportunities and challenges for sample preparation, 3D reconstruction and atomic modeling. Here, the approaches used for cryo-EM and cryo-ET of axonemes are reviewed, while highlighting the unique opportunities provided by the latest generation of AI-guided tools that are transforming structural biology.Copyright (c) 2024 International Union of Crystallographyurn:issn:2059-7983Walton, T.Doran, M.H.Brown, A.2024-03-07doi:10.1107/S2059798324001815International Union of CrystallographyThis review describes how electron-microscopy methods have helped to reveal the structure of the ciliary axoneme.ENcryo-EMcryo-ETciliaaxonemesmicrotubulesThe axoneme, a microtubule-based array at the center of every cilium, has been the subject of structural investigations for decades, but only recent advances in cryo-EM and cryo-ET have allowed a molecular-level interpretation of the entire complex to be achieved. The unique properties of the nine doublet microtubules and central pair of singlet microtubules that form the axoneme, including the highly decorated tubulin lattice and the docking of massive axonemal complexes, provide opportunities and challenges for sample preparation, 3D reconstruction and atomic modeling. Here, the approaches used for cryo-EM and cryo-ET of axonemes are reviewed, while highlighting the unique opportunities provided by the latest generation of AI-guided tools that are transforming structural biology.text/htmlStructural determination and modeling of ciliary microtubulestext4802024-03-07Copyright (c) 2024 International Union of CrystallographyActa Crystallographica Section Dresearch papers00VitroJet: new features and case studies
http://scripts.iucr.org/cgi-bin/paper?ih5005
Single-particle cryo-electron microscopy has become a widely adopted method in structural biology due to many recent technological advances in microscopes, detectors and image processing. Before being able to inspect a biological sample in an electron microscope, it needs to be deposited in a thin layer on a grid and rapidly frozen. The VitroJet was designed with this aim, as well as avoiding the delicate manual handling and transfer steps that occur during the conventional grid-preparation process. Since its creation, numerous technical developments have resulted in a device that is now widely utilized in multiple laboratories worldwide. It features plasma treatment, low-volume sample deposition through pin printing, optical ice-thickness measurement and cryofixation of pre-clipped Autogrids through jet vitrification. This paper presents recent technical improvements to the VitroJet and the benefits that it brings to the cryo-EM workflow. A wide variety of applications are shown: membrane proteins, nucleosomes, fatty-acid synthase, Tobacco mosaic virus, lipid nanoparticles, tick-borne encephalitis viruses and bacteriophages. These case studies illustrate the advancement of the VitroJet into an instrument that enables accurate control and reproducibility, demonstrating its suitability for time-efficient cryo-EM structure determination.Copyright (c) 2024 International Union of Crystallographyurn:issn:2059-7983Henderikx, R.J.M.Mann, D.Domanska, A.Dong, J.Shahzad, S.Lak, B.Filopoulou, A.Ludig, D.Grininger, M.Momoh, J.Laanto, E.Oksanen, H.M.Bisikalo, K.Williams, P.A.Butcher, S.J.Peters, P.J.Beulen, B.W.A.M.M.2024-03-15doi:10.1107/S2059798324001852International Union of CrystallographyThis paper presents recent technical improvements to the VitroJet and the benefits that it brings to the cryo-EM workflow, as well as a wide variety of case studies. This illustrates the advancement of the VitroJet into an instrument that enables accurate control and reproducibility, demonstrating its suitability for time-efficient cryo-EM structure determination.ENcryo-EMVitroJetice thicknesspin printingjet vitrificationSingle-particle cryo-electron microscopy has become a widely adopted method in structural biology due to many recent technological advances in microscopes, detectors and image processing. Before being able to inspect a biological sample in an electron microscope, it needs to be deposited in a thin layer on a grid and rapidly frozen. The VitroJet was designed with this aim, as well as avoiding the delicate manual handling and transfer steps that occur during the conventional grid-preparation process. Since its creation, numerous technical developments have resulted in a device that is now widely utilized in multiple laboratories worldwide. It features plasma treatment, low-volume sample deposition through pin printing, optical ice-thickness measurement and cryofixation of pre-clipped Autogrids through jet vitrification. This paper presents recent technical improvements to the VitroJet and the benefits that it brings to the cryo-EM workflow. A wide variety of applications are shown: membrane proteins, nucleosomes, fatty-acid synthase, Tobacco mosaic virus, lipid nanoparticles, tick-borne encephalitis viruses and bacteriophages. These case studies illustrate the advancement of the VitroJet into an instrument that enables accurate control and reproducibility, demonstrating its suitability for time-efficient cryo-EM structure determination.text/htmlVitroJet: new features and case studiestext4802024-03-15Copyright (c) 2024 International Union of CrystallographyActa Crystallographica Section Dresearch papers00AlphaFold-assisted structure determination of a bacterial protein of unknown function using X-ray and electron crystallography
http://scripts.iucr.org/cgi-bin/paper?jb5060
Macromolecular crystallography generally requires the recovery of missing phase information from diffraction data to reconstruct an electron-density map of the crystallized molecule. Most recent structures have been solved using molecular replacement as a phasing method, requiring an a priori structure that is closely related to the target protein to serve as a search model; when no such search model exists, molecular replacement is not possible. New advances in computational machine-learning methods, however, have resulted in major advances in protein structure predictions from sequence information. Methods that generate predicted structural models of sufficient accuracy provide a powerful approach to molecular replacement. Taking advantage of these advances, AlphaFold predictions were applied to enable structure determination of a bacterial protein of unknown function (UniProtKB Q63NT7, NCBI locus BPSS0212) based on diffraction data that had evaded phasing attempts using MIR and anomalous scattering methods. Using both X-ray and micro-electron (microED) diffraction data, it was possible to solve the structure of the main fragment of the protein using a predicted model of that domain as a starting point. The use of predicted structural models importantly expands the promise of electron diffraction, where structure determination relies critically on molecular replacement.Copyright (c) 2024 International Union of Crystallographyurn:issn:2059-7983Miller, J.E.Agdanowski, M.P.Dolinsky, J.L.Sawaya, M.R.Cascio, D.Rodriguez, J.A.Yeates, T.O,2024-03-07doi:10.1107/S205979832400072XInternational Union of CrystallographyThe structure determination of a small protein of unknown function by molecular replacement using a search model predicted by new machine-learning methods is reported. Notably, the approach was successful using electron diffraction data collected from a protein microcrystal, highlighting a potentially important new route for structure determination.ENelectron diffractionprotein structure predictionAlphaFoldbacterial proteinsmolecular replacementMacromolecular crystallography generally requires the recovery of missing phase information from diffraction data to reconstruct an electron-density map of the crystallized molecule. Most recent structures have been solved using molecular replacement as a phasing method, requiring an a priori structure that is closely related to the target protein to serve as a search model; when no such search model exists, molecular replacement is not possible. New advances in computational machine-learning methods, however, have resulted in major advances in protein structure predictions from sequence information. Methods that generate predicted structural models of sufficient accuracy provide a powerful approach to molecular replacement. Taking advantage of these advances, AlphaFold predictions were applied to enable structure determination of a bacterial protein of unknown function (UniProtKB Q63NT7, NCBI locus BPSS0212) based on diffraction data that had evaded phasing attempts using MIR and anomalous scattering methods. Using both X-ray and micro-electron (microED) diffraction data, it was possible to solve the structure of the main fragment of the protein using a predicted model of that domain as a starting point. The use of predicted structural models importantly expands the promise of electron diffraction, where structure determination relies critically on molecular replacement.text/htmlAlphaFold-assisted structure determination of a bacterial protein of unknown function using X-ray and electron crystallographytext4802024-03-07Copyright (c) 2024 International Union of CrystallographyActa Crystallographica Section Dresearch papers00Efficient in situ screening of and data collection from microcrystals in crystallization plates
http://scripts.iucr.org/cgi-bin/paper?wa5148
A considerable bottleneck in serial crystallography at XFEL and synchrotron sources is the efficient production of large quantities of homogenous, well diffracting microcrystals. Efficient high-throughput screening of batch-grown microcrystals and the determination of ground-state structures from different conditions is thus of considerable value in the early stages of a project. Here, a highly sample-efficient methodology to measure serial crystallography data from microcrystals by raster scanning within standard in situ 96-well crystallization plates is described. Structures were determined from very small quantities of microcrystal suspension and the results were compared with those from other sample-delivery methods. The analysis of a two-dimensional batch crystallization screen using this method is also described as a useful guide for further optimization and the selection of appropriate conditions for scaling up microcrystallization.Copyright (c) 2024 International Union of Crystallographyurn:issn:2059-7983Thompson, A.J.Sanchez-Weatherby, J.Williams, L.J.Mikolajek, H.Sandy, J.Worrall, J.A.R.Hough, M.A.2024-03-15doi:10.1107/S2059798324001955International Union of CrystallographyA sample- and time-efficient method to obtain serial crystallography data from batch-grown microcrystals dispensed as drops on a 96-well crystallization plate is described. This offers a versatile method to obtain low-dose room-temperature structures and guide the optimization of microcrystallization for synchrotron and XFEL serial crystallography experiments.ENserial crystallographyin situ data collectionperoxidaseradiation damagemicrocrystalsA considerable bottleneck in serial crystallography at XFEL and synchrotron sources is the efficient production of large quantities of homogenous, well diffracting microcrystals. Efficient high-throughput screening of batch-grown microcrystals and the determination of ground-state structures from different conditions is thus of considerable value in the early stages of a project. Here, a highly sample-efficient methodology to measure serial crystallography data from microcrystals by raster scanning within standard in situ 96-well crystallization plates is described. Structures were determined from very small quantities of microcrystal suspension and the results were compared with those from other sample-delivery methods. The analysis of a two-dimensional batch crystallization screen using this method is also described as a useful guide for further optimization and the selection of appropriate conditions for scaling up microcrystallization.text/htmlEfficient in situ screening of and data collection from microcrystals in crystallization platestext4802024-03-15Copyright (c) 2024 International Union of CrystallographyActa Crystallographica Section Dresearch papers00