Peters J W, Fisher K, Newton W E, Dean D R
Department of Biochemistry and Anaerobic Microbiology, Virginia Polytechnic Institute and State University, Blacksburg 24061, USA.
J Biol Chem. 1995 Nov 10;270(45):27007-13. doi: 10.1074/jbc.270.45.27007.
Nitrogenase is the catalytic component of biological nitrogen fixation, and it is comprised of two component proteins called the Fe protein and MoFe protein. The Fe protein contains a single Fe4S4 cluster, and the MoFe protein contains two metallocluster types called the P cluster (Fe8S8) and FeMo-cofactor (Fe7S9Mo-homocitrate). During turnover, electrons are delivered one at a time from the Fe protein to the MoFe protein in a reaction coupled to component-protein association-dissociation and MgATP hydrolysis. Under conditions of optimum activity, the rate of component-protein dissociation is rate-limiting. The Fe protein's Fe4S4 cluster is the redox entity responsible for intermolecular electron delivery to the MoFe protein, and FeMo-cofactor provides the substrate reduction site. In contrast, the role of the P cluster in catalysis is not well understood although it is believed to be involved in accumulating electrons delivered from the Fe protein and brokering their intramolecular delivery to the substrate reduction site. A nitrogenase component-protein docking model, which is based on the crystallographic structures of the component proteins and which pairs the 2-fold symmetric surface of the Fe protein with the exposed surface of the MoFe protein's pseudosymmetric alpha beta interface, is now available. During component-protein interaction, this model places the P cluster between the Fe protein's Fe4S4 cluster and FeMo-cofactor, which implies that the P cluster is involved in mediating intramolecular electron transfer between the clusters. In the present study, evidence supporting this idea was obtained by demonstrating that it is possible to alter the rate of substrate reduction by perturbing the polypeptide environment between the P cluster and FeMo-cofactor without necessarily disrupting the metallocluster polypeptide environments or altering component-protein interaction.
固氮酶是生物固氮作用的催化成分，由两种称为铁蛋白和钼铁蛋白的蛋白质组成。铁蛋白含有一个单一的Fe4S4簇，钼铁蛋白含有两种金属簇类型，称为P簇（Fe8S8）和铁钼辅因子（Fe7S9Mo-高柠檬酸）。在催化循环过程中，电子一次一个地从铁蛋白传递到钼铁蛋白，该反应与蛋白质组分的缔合-解离以及MgATP水解相偶联。在最佳活性条件下，蛋白质组分解离的速率是限速的。铁蛋白的Fe4S4簇是负责分子间电子传递到钼铁蛋白的氧化还原实体，铁钼辅因子提供底物还原位点。相比之下，尽管人们认为P簇参与积累从铁蛋白传递来的电子并促成其向底物还原位点的分子内传递，但它在催化中的作用仍未得到很好的理解。现在有一个基于蛋白质组分晶体结构的固氮酶蛋白质组分对接模型，该模型将铁蛋白的2倍对称表面与钼铁蛋白假对称αβ界面的暴露表面配对。在蛋白质组分相互作用过程中，该模型将P簇置于铁蛋白的Fe4S4簇和铁钼辅因子之间，这意味着P簇参与介导簇之间的分子内电子转移。在本研究中，通过证明有可能通过扰动P簇和铁钼辅因子之间的多肽环境来改变底物还原速率，而不必破坏金属簇多肽环境或改变蛋白质组分相互作用，从而获得了支持这一观点的证据。
