Iron–sulfur cluster coordination in the [FeFe]-hydrogenase H cluster biosynthetic factor HydF

Iron–sulfur cluster coordination was probed in the [FeFe]-hydrogenase H cluster maturation scaffold HydF. Putative Cys thiol and His imidazole ligation identified through multiple sequence alignments and structural studies were subjected to amino acid substitution and the variants were biochemically characterized. The results implicate a role for C304, C353, C356, and H306 of Clostridium acetobutylicum HydF in FeS cluster binding. Individual ligand substitutions affect both [4Fe–4S] and [2Fe–2S] cluster coordination suggesting shared coordination or cluster interconversion. Substitutions at C353 and H306 appear to preferentially impact the presence of the [2Fe–2S] cluster complement of the resulting variants of HydF. The results implicate a potential role for these residues in biosynthesis specifically and potential in bridging the [4Fe–4S] cluster to 2Fe subcluster biosynthetic intermediates.Structured summary of protein interactionsHydF and HydFbind by molecular sieving(View interaction: 1, 2)

► Iron–sulfur cluster coordination in HydF was investigated by site-specific amino acid substitution studies.
► Cys and His residues are implicated in [4Fe–4S] and [2Fe–2S] cluster coordination.
► Key substitutions preferentially impact [2Fe–2S] cluster indicating these residues may have a specific role in 2Fe subcluster biosynthesis.