Final Stages in the Biosynthesis of the [FeFe]‐Hydrogenase Active Site

Abstract

The paper aims to elucidate the final stages in the biosynthesis of the [2Fe]H active site of the [FeFe]‐hydrogenases. The recently hypothesized intermediate [Fe2(SCH2NH2)2(CN)2(CO)4]2‐ ([1]2‐) was prepared by a multistep route from [Fe2(S2)(CN)(CO)5]‐. The following synthetic intermediates were characterized in order: [Fe2(SCH2NHFmoc)2(CNBEt3)(CO)5]‐, [Fe2(SCH2NHFmoc)2(CN)‐(CO)5]‐, and [Fe2(SCH2NHFmoc)2(CN)2(CO)4]2‐, where Fmoc is fluorenylmethoxycarbonyl). Derivatives of these anions include [K(18‐crown‐6)]+, PPh4+ and PPN+ salts as well as the 13CD2‐isotopologues. These Fe2 species exist as a mixture of two isomers attributed to diequatorial (ee) and axial‐equatorial (ae) stereochemistry at sulfur. In vitro experiments demonstrate that [1]2‐ maturates HydA1 in the presence of HydF and a cocktail of reagents. HydA1 can also be maturated using a highly simplified cocktail, omitting HydF and other proteins. This result is consistent with HydA1 participating in the maturation process and refines the roles of HydF.