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![[Figure 4]](lp5050fig4mag.jpg)
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Figure 4
Putative strategy of promoter recognition by dimeric osAca2. (a) Cartoon representation of dimeric osAca2 colored according to the degree of amino-acid sequence conservation as analyzed by the ConSurf server. (b) Sequence alignment of Aca2 from different species. Mostly conserved and partially conserved residues are colored red and blue, respectively. The location of the three helices α1–α3 at the N-terminus containing the HTH motif are shown above the sequence. # indicates conserved residues involved in formation of the dimeric interface. * indicates conserved residues that might be involved in DNA recognition. (c) Structural model of the osAca2–P1 promoter complex generated by the HDOCK server. The sequence of the P1 putative osAca2 binding site used for modeling and docking is provided below the structural model. Arg30, Gln33, Tyr34 and Arg39 involved in the readout of the palindromic GTT sequence are labeled in the structure. (d) Close-up view of the putative binding region between osAca2 and promoter DNA. Residues involved in the readout of the palindromic GTT sequence are labeled. (e) Cartoon representation showing the distance between Arg30 in each molecule of dimeric osAca2. The black dashed box indicates the position magnified in (d). (f) Mobility shift of synthesized DNA probes observed on an agarose gel with increasing concentrations of DNA binding-disturbed mutants, including R30D, Q33R, Y34W and R39W. The DNA probes used in the experiment are indicated below the gel. Concentration-dependence is indicated by the black triangle. A concentration gradient, 0, 0.07, 0.18 and 0.36 mM, was applied for each mutant protein. Lane M contains molecular size markers. (g) Verification of the effect of mutagenesis on the stoichiometric alteration of osAca2. The SEC profiles compare the positions of the eluted peaks for the wild type and each mutant protein. (h) MALS profile of the R30D mutant of osAca2. The red line indicates the experimental molecular mass analyzed by MALS. |
![[Figure 4]](lp5050fig4.jpg)
 | STRUCTURAL BIOLOGY |
ISSN: 2059-7983
