期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2015
卷号：112
期号：37
页码：11455-11460
DOI：10.1073/pnas.1508440112
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：SignificanceHydrogen production is central to a solar fuel paradigm, and a variety of metabolic processes use H2 as an electron donor or protons as an electron acceptor. Hydrogenases mediate the biological redox interconversion of protons and H2, with FeFe hydrogenases among the most active. This reactivity occurs at the "H cluster," which features an organometallic subcluster that is synthesized and inserted in a complex series of steps. The accessory protein HydG generates an [Fe(CO)2(CN)] intermediate en route to the H cluster, and the mechanism of this process is under intensive investigation. We now report that free L-cysteine serves as the ligand platform on which the [Fe(CO)2(CN)] synthon is built and plays a role in both Fe2+ binding and synthon release. Hydrogenases catalyze the redox interconversion of protons and H2, an important reaction for a number of metabolic processes and for solar fuel production. In FeFe hydrogenases, catalysis occurs at the H cluster, a metallocofactor comprising a [4Fe-4S]H subcluster coupled to a [2Fe]H subcluster bound by CO, CN-, and azadithiolate ligands. The [2Fe]H subcluster is assembled by the maturases HydE, HydF, and HydG. HydG is a member of the radical S-adenosyl-L-methionine family of enzymes that transforms Fe and L-tyrosine into an [Fe(CO)2(CN)] synthon that is incorporated into the H cluster. Although it is thought that the site of synthon formation in HydG is the "dangler" Fe of a [5Fe] cluster, many mechanistic aspects of this chemistry remain unresolved including the full ligand set of the synthon, how the dangler Fe initially binds to HydG, and how the synthon is released at the end of the reaction. To address these questions, we herein show that L-cysteine (Cys) binds the auxiliary [4Fe-4S] cluster of HydG and further chelates the dangler Fe. We also demonstrate that a [4Fe-4S]aux[CN] species is generated during HydG catalysis, a process that entails the loss of Cys and the [Fe(CO)2(CN)] fragment; on this basis, we suggest that Cys likely completes the coordination sphere of the synthon. Thus, through spectroscopic analysis of HydG before and after the synthon is formed, we conclude that Cys serves as the ligand platform on which the synthon is built and plays a role in both Fe2+ binding and synthon release.
