Rypniewski W R, Mangani S, Bruni B, Orioli P L, Casati M, Wilson K S
European Molecular Biology Laboratory (EMBL), Hamburg, Germany.
J Mol Biol. 1995 Aug 11;251(2):282-96. doi: 10.1006/jmbi.1995.0434.
Cu,Zn superoxide dismutase was investigated crystallographically in the reduced form. Co-ordinate errors were estimated by comparing two independently refined models, based on two different data sets. This gave a detailed error estimation as opposed to the standard sigma A and Luzzati plots, which estimate only the overall error. The high quality of the final model, obtained after scaling together the two data sets, combined with the error estimates allowed a detailed analysis of the protein and solvent structures. An automatic procedure for building and refining solvent structure was tested and found to give reproducible results. Contrary to results obtained from spectroscopic studies, the co-ordination of the metal ions in the catalytic site is preserved in the crystal structure of the reduced enzyme, as compared with the crystal structure of the oxidised form. Analysis of the solvent reveals a well-defined chain of closely packed, hydrogen-bonded water molecules filling the active site groove. This structural feature could serve as a hydrogen bond relay for efficient delivery of protons to the active centre. Analysis of electron density suggests that Glu119 is covalently modified. The modification, if originated in vivo, could have a role in the catalytic mechanism and could affect the overall electrostatic field in the active site. There are significant differences between the active sites of the two crystallographically independent monomers. They are explained in terms of local differences in the crystal environment.
对还原态的铜锌超氧化物歧化酶进行了晶体学研究。通过比较基于两个不同数据集独立精修的两个模型来估计坐标误差。与仅估计总体误差的标准西格玛A和卢扎蒂图不同，这给出了详细的误差估计。将两个数据集一起缩放后获得的最终模型质量很高，结合误差估计，能够对蛋白质和溶剂结构进行详细分析。测试了一种构建和精修溶剂结构的自动程序，发现其结果具有可重复性。与光谱学研究结果相反，与氧化态的晶体结构相比，还原酶晶体结构中催化位点的金属离子配位得以保留。对溶剂的分析揭示了一条排列紧密、通过氢键相连的水分子链，填充了活性位点凹槽。这一结构特征可作为氢键中继，将质子高效传递至活性中心。电子密度分析表明，Glu119发生了共价修饰。如果这种修饰发生在体内，可能在催化机制中起作用，并可能影响活性位点的整体静电场。两个晶体学独立单体的活性位点之间存在显著差异。这些差异可以从晶体环境的局部差异方面进行解释。
