Structural Basis for the Reaction Mechanism of S-Carbamoylation of HypE by HypF in the Maturation of [NiFe]-Hydrogenases

Crystal structures of the carbamoylated and cyanated forms of HypE for [NiFe] hydrogenase maturation

“…The aspartate residue in the DX 4 GAXP motif is involved in holding Mg 2+ ions and nucleotides. The crystal structures of HypE have been reported from Thermococcus kodakarensis (9), Desulfovibrio vulgaris (21), Escherichia coli (22), and Caldanaerobacter subterraneus (23). The structures of all of the other Hyp proteins have also been reported (9,(24)(25)(26)(27)(28)(29)(30)(31)(32).…”

“…The structures of all of the other Hyp proteins have also been reported (9,(24)(25)(26)(27)(28)(29)(30)(31)(32). The carbamoylation mechanism of the C-terminal cysteine residue of HypE was proposed based on the structure of the HypE-HypF complex (23). The N-terminal acylphosphataselike domain of HypF catalyzes the formation of a carbamate intermediate, which can be transferred to the active site through a tunnel inside the protein.…”

[NiFe]‐hydrogenase maturation: Isolation of a HypC–HypD complex carrying diatomic CO and CN− ligands

“…In an ATPdependent reaction the carbamoyltransferase HypF transfers a carbamoyl moiety to the C-terminal cysteinyl residue of HypE [7,8,13,14], which then catalyses the ATP-dependent dehydration of the thiocarboxamide to a thiocyanate group. In vitro studies have demonstrated that the CN À moiety can be transferred to the HypCD complex, provided that it is isolated anaerobically; however, neither component individually can accept the CN À group, suggesting the proteins share coordination of the cofactor [15].…”

Structural Foundations for O2 Sensitivity and O2 Tolerance in [NiFe]-Hydrogenases

“…Significant insight into the molecular aspects of the EcHyd-3 large subunit maturation has been provided by the X-ray structure determinations of HypA, HypB, HypC, HypD, HypE, HypF, HycI and SlyD (Watanabe et al 2009;Xia et al 2009;Gasper et al 2006;Chan et al 2012;Watanabe et al 2007;Shomura et al 2007;Rangarajan et al 2008;Shomura and Higuchi 2012;Petkun et al 2011;Kumarevel et al 2009;Loew et al 2010), but some details remain unclear. In addition, although apo pre HycE has been generally assumed to be devoid of metal, a recent report suggests that a mutant of HybC, the homologous unprocessed large subunit of EcHyd-2, may actually contain a labile [Fe 4 S 4 ] cluster at the active site position in the mature subunit (Soboh et al 2012).…”

“…CP is converted by HypF to H 2 NC(O)-AMP, also in an ATP-dependent reaction (reactions 4-5). After formation of a HypEF complex, of known structure (Shomura and Higuchi 2012), the H 2 NCO group is transferred to the C-terminal cysteine of HypE (reaction 6). The resulting thiocarbamate is subsequently dehydrated in yet another ATP-dependent reaction to thiocyanate (reaction 7), followed by CN transfer to Fe bound to HypCD (reaction 8) in a putative HypCDE complex (Watanabe et al 2007).…”