1887

Abstract

Escherichia coli has two machineries for the synthesis of FeS clusters, namely Isc (iron–sulfur cluster) and Suf (sulfur formation). The Isc machinery, encoded by the iscRSUA-hscBA-fdx-iscXoperon, plays a crucial role in the biogenesis of FeS clusters for the oxidoreductases of aerobic metabolism. Less is known, however, about the role of ISC in the maturation of key multi-subunit metalloenzymes of anaerobic metabolism. Here, we determined the contribution of each iscoperon gene product towards the functionality of the major anaerobic oxidoreductases in E. coli, including three [NiFe]-hydrogenases (Hyd), two respiratory formate dehydrogenases (FDH) and nitrate reductase (NAR). Mutants lacking the cysteine desulfurase, IscS, lacked activity of all six enzymes, as well as the activity of fumaratereductase, and this was due to deficiencies in enzyme biosynthesis, maturation or FeS cluster insertion into electron-transfer components. Notably, based on anaerobic growth characteristics and metabolite patterns, the activity of the radical-S-adenosylmethionine enzyme pyruvate formate-lyase activase was independent of IscS, suggesting that FeS biogenesis for this ancient enzyme has different requirements. Mutants lacking either the scaffold protein IscU, the ferredoxin Fdx or the chaperones HscA or HscB had similar enzyme phenotypes: five of the oxidoreductases were essentially inactive, with the exception being the Hyd-3 enzyme, which formed part of the H2-producing formate hydrogenlyase (FHL) complex. Neither the frataxin-homologue CyaY nor the IscX protein was essential for synthesis of the three Hyd enzymes. Thus, while IscS is essential for H2 production in E. coli, the other ISC components are non-essential.

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2017-06-26
2019-10-15
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