The Influence of Oxygen on [NiFe]–Hydrogenase Cofactor Biosynthesis and How Ligation of Carbon Monoxide Precedes Cyanation
نویسندگان
چکیده
The class of [NiFe]-hydrogenases is characterized by a bimetallic cofactor comprising low-spin nickel and iron ions, the latter of which is modified with a single carbon monoxide (CO) and two cyanide (CN-) molecules. Generation of these ligands in vivo requires a complex maturation apparatus in which the HypC-HypD complex acts as a 'construction site' for the Fe-(CN)2CO portion of the cofactor. The order of addition of the CO and CN- ligands determines the ultimate structure and catalytic efficiency of the cofactor; however much debate surrounds the succession of events. Here, we present an FT-IR spectroscopic analysis of HypC-HypD isolated from a hydrogenase-competent wild-type strain of Escherichia coli. In contrast to previously reported samples, HypC-HypD showed spectral contributions indicative of an electron-rich Fe-CO cofactor, at the same time lacking any Fe-CN- signatures. This immature iron site binds external CO and undergoes oxidative damage when in contact with O2. Binding of CO protects the site against loss of spectral features associated with O2 damage. Our findings strongly suggest that CO ligation precedes cyanation in vivo. Furthermore, the results provide a rationale for the deleterious effects of O2 on in vivo cofactor biosynthesis.
منابع مشابه
Identification of an Isothiocyanate on the HypEF Complex Suggests a Route for Efficient Cyanyl–Group Channeling during [NiFe]–Hydrogenase Cofactor Generation
[NiFe]-hydrogenases catalyze uptake and evolution of H2 in a wide range of microorganisms. The enzyme is characterized by an inorganic nickel/ iron cofactor, the latter of which carries carbon monoxide and cyanide ligands. In vivo generation of these ligands requires a number of auxiliary proteins, the so-called Hyp family. Initially, HypF binds and activates the precursor metabolite carbamoyl ...
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