Studies of carbon monoxide dehydrogenase from Oligotropha carboxidovorans

Publication date: Available online 11 October 2016 Source:Journal of Molecular Catalysis B: Enzymatic Author(s): Stephanie Dingwall, Jarett Wilcoxen, Dimitri Niks, Russ Hille We have undertaken physicochemical studies of the CO dehydrogenase from the aerobe Oligotropha carboxidovorans, probing both the binuclear Mo- and Cu-containing active site where CO is oxidized to CO2 and the enzyme’s FAD, where the reducing equivalents obtained from CO are transferred to the quinone pool. Regarding the FAD site, we have characterized the semiquinone oxidation state by EPR and identified it to be of the blue neutral form with a linewidth of 20G. The signature long-wavelength absorbance of FADH• is also observed in the absorption spectrum of partially reduced enzyme at low pH. The enzyme exhibits a pH-dependent absorption spectrum in the oxidized state that is lost upon covalent modification of the enzyme by the flavin-specific agent diphenyliodonium cation. The pH dependence is attributed to Tyr 193 of the FAD-containing CoxM subunit, which sits atop the re face of the isoalloxazine ring in van der Waals contact with it. Electron equilibration among the enzyme’s four redox-active centers (including two [2Fe-2S] clusters in addition to the binuclear center and FAD) is found to be pH-dependent, but too fast to be followed using a stopped-flow pH jump protocol. Electron transfer from the iron-sulfur clusters to the FAD is thus much faster than in other members of the xanthine ox...

Source: Journal of Molecular Catalysis B: Enzymatic - October 11, 2016 Category: Biochemistry Source Type: research