Raman and Infrared Spectroscopy of Cyanide-Inhibited CO Dehydrogenase/Acetyl-CoA Synthase from Clostridium thermoaceticum: Evidence for Bimetallic Enzymatic CO Oxidation

Abstract

Clostridium thermoaceticum and other autotrophic anaerobic bacteria contain a bifunctional enzyme, carbon monoxide dehydrogenase/acetyl-CoA synthase (CODH/ACS), that catalyzes two reactions of CO at two separate Ni−FeS clusters. Oxidation of CO to CO2 is catalyzed by Cluster C, while incorporation of CO into acetyl-CoA occurs at Cluster A. In this study, resonance Raman [RR] and infrared [IR] spectroscopy are applied to the adduct of Cluster C with cyanide, a selective inhibitor of CO oxidation. The RR spectra reveal that CN- binds simultaneously to Fe and Ni, because bands whose 13C and 15N shifts identify them as cyanide−metal stretching and bending modes are sensitive to incorporation of both 54Fe and 64Ni into the enzyme. The IR spectrum reveals a low frequency, 2037 cm-1, for the C−N stretch, indicative of FeII binding via the C end. Vibrational modeling of the frequencies and isotope shifts indicates a bent Fe−CN−Ni bridging geometry, with a ∼140° C−N−Ni angle. This geometry of the inhibitory adduct...