title: Characterization of the metal centers of the Ni/Fe-S component of the carbon-monoxide dehydrogenase enzyme complex from Methanosarcina thermophila.
abstract: Methanosarcina thermophila contains a multienzyme complex called the carbon-monoxide dehydrogenase complex, which has been resolved into a nickel/iron-sulfur and a corrinoid/iron-sulfur component. This complex plays a central role in acetoclastic methanogenesis. The Ni/Fe-S component catalyzes CO oxidation and has been proposed to be involved in cleavage of acetyl-CoA into its methyl, carbonyl, and CoA moieties. In the work reported here, three metal centers in the Ni/Fe-S component were characterized by electron paramagnetic resonance (EPR) spectroscopy and spectroelectrochemistry and pre-steady state kinetics. Center A contains nickel and iron and forms an EPR active adduct with CO, which is called the NiFeC species. The EPR spectrum of the NiFeC species has g values of 2.059, 2.051, and 2.029 and is observable at temperatures as high as 150 K. This signal had previously been observed only in the carbon-monoxide dehydrogenase complex of M. thermophila and the acetyl-CoA synthase from acetate-producing bacteria. Incubation of the CO-reduced Ni/Fe-S component with acetyl-CoA resulted in an increase in intensity of the NiFeC signal, which supports a role for the component in the cleavage of acetyl-CoA. Generation of the NiFeC EPR signal occurs with a rate constant of 0.4 s-1, a result that demonstrates the kinetic competence of this species in the acetyl-CoA cleavage reaction but rules it out as the site of oxidation of CO to CO2. Center B is likely to be a [4Fe-4S]2+/1+ center with g values of 2.04, 1.93, and 1.89 (gav = 1.95) and a standard reduction potential (E'0) of -444 mV. At potentials less than -500 mV, another EPR signal develops that appears to originate from another state of Center B. Center C is a fast relaxing center with g values of 2.02, 1.88, and 1.71 (gav = 1.87) and an E'0 of -154 mV.
authors.affiliation: Department of Biochemistry, University of Nebraska, Lincoln 68583-0718.
authors.forename: W P
authors.initials: WP
authors.lastname: Lu
authors.forename: P E
authors.initials: PE
authors.lastname: Jablonski
authors.forename: M
authors.initials: M
authors.lastname: Rasche
authors.forename: J G
authors.initials: JG
authors.lastname: Ferry
authors.forename: S W
authors.initials: SW
authors.lastname: Ragsdale
mesh_terms: D000445:Aldehyde Oxidoreductases / Q000737:chemistry* / Q000302:isolation & purification; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods; D019855:Ethylene Glycol; D005026:Ethylene Glycols / Q000494:pharmacology; D005615:Freezing; D007501:Iron / Q000032:analysis; D007506:Iron-Sulfur Proteins / Q000737:chemistry* / Q000302:isolation & purification; D017020:Methanosarcina / Q000201:enzymology*; D009097:Multienzyme Complexes; D009532:Nickel / Q000032:analysis*; D010084:Oxidation-Reduction; D011487:Protein Conformation; D013045:Species Specificity; D013455:Sulfur / Q000032:analysis; D013816:Thermodynamics
chemical_list: D005026:Ethylene Glycols; D007506:Iron-Sulfur Proteins; D009097:Multienzyme Complexes; D013455:Sulfur; D009532:Nickel; D007501:Iron; D000445:Aldehyde Oxidoreductases; C019823:carbon monoxide dehydrogenase; D019855:Ethylene Glycol