Schlessman J L, Woo D, Joshua-Tor L, Howard J B, Rees D C
Division of Chemistry and Chemical Engineering, California Institute of Technology, 147-75CH, Pasadena, CA 91125, USA.
J Mol Biol. 1998 Jul 24;280(4):669-85. doi: 10.1006/jmbi.1998.1898.
The nitrogenase iron (Fe) protein performs multiple functions during biological nitrogen fixation, including mediating the mechanistically essential coupling between ATP hydrolysis and electron transfer to the nitrogenase molybdenum iron (MoFe) protein during substrate reduction, and participating in the biosynthesis and insertion of the FeMo-cofactor into the MoFe-protein. To establish a structural framework for addressing the diverse functions of Fe-protein, crystal structures of the Fe-proteins from Azotobacter vinelandii and Clostridium pasteurianum have been determined at resolutions of 2.2 A and 1.93 A, respectively. These two Fe-proteins are among the more diverse in terms of amino acid sequence and biochemical properties. As described initially for the A. vinelandii Fe-protein in a different crystal form at 2.9 A resolution, each subunit of the dimeric Fe-protein adopts a polypeptide fold related to other mononucleotide-binding proteins such as G-proteins, with the two subunits bridged by a 4Fe:4S cluster. The overall similarities in the subunit fold and dimer arrangement observed in the structures of the A. vinelandii and C. pasteurianum Fe-proteins indicate that they are representative of the conformation of free Fe-protein that is not in complex with nucleotide or the MoFe-protein. Residues in the cluster and nucleotide-binding sites are linked by a network of conserved hydrogen bonds, salt-bridges and water molecules that may conformationally couple these regions. Significant variability is observed in localized regions, especially near the 4Fe:4S cluster and the MoFe-protein binding surface, that change conformation upon formation of the ADP.AlF4- stabilized complex with the MoFe-protein. A core of 140 conserved residues is identified in an alignment of 59 Fe-protein sequences that may be useful for the identification of homologous proteins with functions comparable to that of Fe-protein in non-nitrogen fixing systems.
固氮酶铁（Fe）蛋白在生物固氮过程中发挥多种功能，包括在底物还原过程中介导ATP水解与向固氮酶钼铁（MoFe）蛋白的电子转移之间机制上必不可少的偶联，以及参与FeMo辅因子的生物合成并将其插入MoFe蛋白中。为了建立一个用于研究Fe蛋白多种功能的结构框架，分别以2.2 Å和1.93 Å的分辨率测定了来自棕色固氮菌和巴氏梭菌的Fe蛋白的晶体结构。这两种Fe蛋白在氨基酸序列和生化特性方面是差异较大的蛋白。正如最初在2.9 Å分辨率下以不同晶体形式描述的棕色固氮菌Fe蛋白那样，二聚体Fe蛋白的每个亚基都采用与其他单核苷酸结合蛋白（如G蛋白）相关的多肽折叠结构，两个亚基由一个4Fe:4S簇桥接。在棕色固氮菌和巴氏梭菌Fe蛋白结构中观察到的亚基折叠和二聚体排列的总体相似性表明，它们代表了未与核苷酸或MoFe蛋白结合的游离Fe蛋白的构象。簇和核苷酸结合位点中的残基通过保守的氢键、盐桥和水分子网络相连，这些网络可能在构象上耦合这些区域。在局部区域观察到显著的变异性，特别是在4Fe:4S簇和MoFe蛋白结合表面附近，在与MoFe蛋白形成ADP·AlF4-稳定复合物时这些区域的构象会发生变化。在59个Fe蛋白序列的比对中鉴定出140个保守残基的核心区域，这可能有助于鉴定在非固氮系统中具有与Fe蛋白功能相当的同源蛋白。
