NifX and NifEN exchange NifB cofactor and the VK-cluster, a newly isolated intermediate of the iron-molybdenum cofactor biosynthetic pathway
NifX and NifEN exchange NifB cofactor and the VK-cluster, a newly isolated intermediate of the iron-molybdenum cofactor biosynthetic pathway
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DOI：
10.1111/j.1365-2958.2006.05514.x
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发表时间：
2007-01-01
影响因子：
3.6
通讯作者：
Rubio, Luis M.
中科院分区：
文献类型：
--
作者：
Hernandez, Jose A.;Igarashi, Robert Y.;Rubio, Luis M.
固氮酶的铁钼辅因子（FeMo-co）是在几种Nif蛋白的催化下在多步过程中合成的，并且最终被插入到预先合成的脱辅基二固氮酶中以产生成熟的二固氮酶蛋白。NifEN复合物充当该合成的某些步骤的支架，而NifX属于小蛋白家族，在辅因子合成期间结合FeMo-co前体或FeMo-co。在这项工作中，FeMo-co前体的结合和纯化棕色固氮菌NifX和NifEN蛋白之间的转移进行了研究，揭示了NifX的FeMo-co合成的作用。纯化的NifX以高亲和力结合NifB辅因子（NifB-co）（FeMo-co的前体），并且能够将其转移到NifEN复合物。此外，NifEN和NifX交换另一个[Fe-S]簇，作为FeMo-co前体，我们将其命名为VK-簇。与NifB-co相反，VK簇在还原和氧化状态下具有电子顺磁共振（EPR）活性。NifX/VK-簇复合物在不存在NifEN的情况下不能支持体外FeMo-co合成，因为将VK-簇进一步加工成FeMo-co需要NifEN和NifH的同时活性。我们的体外研究表明，NifX在体内的作用是作为FeMo-co前体的瞬时储存库，从而帮助控制它们在FeMo-co合成过程中的通量。
The iron-molybdenum cofactor of nitrogenase (FeMo-co) is synthesized in a multistep process catalysed by several Nif proteins and is finally inserted into a pre-synthesized apo-dinitrogenase to generate mature dinitrogenase protein. The NifEN complex serves as scaffold for some steps of this synthesis, while NifX belongs to a family of small proteins that bind either FeMo-co precursors or FeMo-co during cofactor synthesis. In this work, the binding of FeMo-co precursors and their transfer between purified Azotobacter vinelandii NifX and NifEN proteins was studied to shed light on the role of NifX on FeMo-co synthesis. Purified NifX binds NifB cofactor (NifB-co), a precursor to FeMo-co, with high affinity and is able to transfer it to the NifEN complex. In addition, NifEN and NifX exchange another [Fe-S] cluster that serves as a FeMo-co precursor, and we have designated it as the VK-cluster. In contrast to NifB-co, the VK-cluster is electronic paramagnetic resonance (EPR)-active in the reduced and the oxidized states. The NifX/VK-cluster complex is unable to support in vitro FeMo-co synthesis in the absence of NifEN because further processing of the VK-cluster into FeMo-co requires the simultaneous activities of NifEN and NifH. Our in vitro studies suggest that the role of NifX in vivo is to serve as transient reservoir of FeMo-co precursors and thus help control their flux during FeMo-co synthesis.