Bordo D, Matak D, Djinovic-Carugo K, Rosano C, Pesce A, Bolognesi M, Stroppolo M E, Falconi M, Battistoni A, Desideri A
Department of Physics - INFM and Advanced Biotechnology Center - IST, University of Genova, Largo R. Benzi, 10, Genova, I-16132, Italy.
J Mol Biol. 1999 Jan 8;285(1):283-96. doi: 10.1006/jmbi.1998.2267.
Prokaryotic Cu,Zn superoxide dismutases are characterized by a distinct quaternary structure, as compared to that of the homologous eukaryotic enzymes. Here we report a newly determined crystal structure of the dimeric Cu,Zn superoxide dismutase from Photobacterium leiognathi (crystallized in space group R32, refined at 2.5 A resolution, R-factor 0.19) and analyse it in comparison with that of the monomeric enzyme from Escherichia coli. The dimeric assembly, observed also in a previously studied monoclinic crystal form of P. leiognathi Cu,Zn superoxide dismutase, is based on a ring-shaped subunit contact region, defining a solvated interface cavity. Three clusters of neighbouring residues play a direct role in the stabilization of the quaternary assembly. The present analysis, extended to the amino acid sequences of the other 11 known prokaryotic Cu,Zn superoxide dismutases, shows that at least in five other prokaryotic enzymes the interface residue clusters are under strong evolutionary constraint, suggesting the attainment of a quaternary structure coincident with that of P. leiognathi Cu,Zn superoxide dismutase. Calculation of electrostatic fields for both the enzymes from E. coli and P. leiognathi shows that the monomeric/dimeric association behaviour displayed by prokaryotic Cu, Zn superoxide dismutases is related to the distribution of surface charged residues. Moreover, Brownian dynamics simulations reproduce closely the observed enzyme:substrate association rates, highlighting the role of the active site neighbouring residues in determining the dismutase catalytic properties.
与同源的真核酶相比，原核铜锌超氧化物歧化酶具有独特的四级结构。在此，我们报道了新测定的来自鱼发光杆菌的二聚体铜锌超氧化物歧化酶的晶体结构（在空间群R32中结晶，在2.5埃分辨率下精修，R因子为0.19），并将其与来自大肠杆菌的单体酶的晶体结构进行比较分析。在先前研究的鱼发光杆菌铜锌超氧化物歧化酶的单斜晶型中也观察到的二聚体组装，基于一个环形亚基接触区域，界定了一个溶剂化的界面腔。三簇相邻残基在四级组装的稳定中起直接作用。本分析扩展到其他11种已知原核铜锌超氧化物歧化酶的氨基酸序列，结果表明至少在其他五种原核酶中，界面残基簇受到强烈的进化限制，这表明它们获得了与鱼发光杆菌铜锌超氧化物歧化酶一致的四级结构。对大肠杆菌和鱼发光杆菌的两种酶的静电场计算表明，原核铜锌超氧化物歧化酶表现出的单体/二聚体缔合行为与表面带电残基的分布有关。此外，布朗动力学模拟紧密再现了观察到的酶与底物的缔合速率，突出了活性位点相邻残基在决定歧化酶催化特性中的作用。
