The HydG Enzyme Generates an Fe(CO) 2 (CN) Synthon in Assembly of the FeFe Hydrogenase H-Cluster

Abstract

Three iron-sulfur proteins–HydE, HydF, and HydG–play a key role in the synthesis of the [2Fe] H component of the catalytic H-cluster of FeFe hydrogenase. The radical S -adenosyl- l -methionine enzyme HydG lyses free tyrosine to produce p -cresol and the CO and CN − ligands of the [2Fe] H cluster. Here, we applied stopped-flow Fourier transform infrared and electron-nuclear double resonance spectroscopies to probe the formation of HydG-bound Fe-containing species bearing CO and CN − ligands with spectroscopic signatures that evolve on the 1- to 1000-second time scale. Through study of the 13 C, 15 N, and 57 Fe isotopologs of these intermediates and products, we identify the final HydG-bound species as an organometallic Fe(CO) 2 (CN) synthon that is ultimately transferred to apohydrogenase to form the [2Fe] H component of the H-cluster.