Spectroscopic studies of nickel-deficient carbon monoxide dehydrogenase from Rhodospirillum rubrum: nature of the iron-sulfur clusters.

Carbon monoxide dehydrogenase (CODH) from Rhodospirillum rubrum utilizes three types of Fe-S clusters to catalyze the reversible oxidation of CO to CO(2): a novel [Ni4Fe5S] active site (C cluster) and two distinct [4Fe4S] electron-transfer sites (B and D clusters). While recent X-ray data show the geometric arrangement of the five metal centers at the C cluster, electronic structures of the various [Ni4Fe5S] oxidation states remain ambiguous. These studies report magnetic circular dichroism (MCD), variable temperature, variable field MCD (VTVH MCD), and resonance Raman (rR) spectroscopic properties of the Fe-S clusters contained in Ni-deficient CODH. Essentially homogeneous sample preparations aided in the resolution of the reduced [4Fe4S](1+) (S = (1)/(2)) B cluster and the reduced Ni-deficient C cluster (denoted C, S > (1)/(2)) by MCD. The three Fe atoms derived from the [Ni3Fe4S] cubane component appear to dominate the reduced C cluster MCD spectrum, while the presence of a fourth Fe center can be inferred from the ground state spin. The same underlying MCD features present in Ni-deficient CODH spectra are also observed for Ni-containing CODH, suggesting that both proteins contain the same C cluster Fe-S component. Overlooked in all spectroscopic studies to date, the D cluster was confirmed by rR to be a typical [4Fe4S] site with cysteinyl coordination. Together, MCD and rR data show that the D cluster remains in the oxidized [4Fe4S](2+) (S = 0) state at potentials > or = -530 mV (versus SHE), thus exhibiting an unusually low redox potential for a standard [4Fe4S](2+/1+) electron-transfer site.