Open AccessIncorporation of Ni2+, Co2+, and Selenocysteine into the Auxiliary Fe-S Cluster of the Radical SAM Enzyme HydG
Author(s) -
Guodong Rao,
Katherine B. Alwan,
Ninian J. Blackburn,
R. David Britt
Publication year - 2019
Publication title -
inorganic chemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.348
H-Index - 233
eISSN - 1520-510X
pISSN - 0020-1669
DOI - 10.1021/acs.inorgchem.9b01293
Subject(s) - selenocysteine , chemistry , crystallography , cluster (spacecraft) , stereochemistry , x ray absorption spectroscopy , extended x ray absorption fine structure , cluster chemistry , active site , group 2 organometallic chemistry , ligand (biochemistry) , molecule , absorption spectroscopy , cysteine , enzyme , organic chemistry , biochemistry , physics , quantum mechanics , computer science , programming language , receptor
The radical SAM enzyme HydG generates CO- and CN - -containing Fe complexes that are involved in the bioassembly of the [FeFe] hydrogenase active cofactor, the H-cluster. HydG contains a unique 5Fe-4S cluster in which the fifth "dangler" Fe and the coordinating cysteine molecule have both been shown to be essential for its function. Here, we demonstrate that this dangler Fe can be replaced with Ni 2+ or Co 2+ and that the cysteine can be replaced with selenocysteine. The resulting HydG variants were characterized by electron paramagnetic resonance and X-ray absorption spectroscopy, as well as subjected to a Tyr cleavage assay. Both Ni 2+ and Co 2+ are shown to be exchange-coupled to the 4Fe-4S cluster, and selenocysteine substitution does not alter the electronic structure significantly. XAS data provide details of the coordination environments near the Ni, Co, and Se atoms and support a close interaction of the dangler metal with the FeS cluster via an asymmetric SeCys bridge. Finally, while we were unable to observe the formation of novel organometallic species for the Ni 2+ and Co 2+ variants, the selenocysteine variant retains the activity of wild type HydG in forming [Fe(CO) x (CN) y ] species. Our results provide more insights into the unique auxiliary cluster in HydG and expand the scope of artificially generated Fe-S clusters with heteroatoms.