Oxidation of dithiothreitol during turnover of nitric oxide reductase: evidence for generation of nitroxyl with the enzyme from Paracoccus denitrificans

Summary The stoichiometric relationship between thiol oxidized and NO reduced was studied for the reaction catalyzed by nitric oxide reductase from Paracoccus denitrificans . The reaction systems consisted of dithiothreitol, ascorbate, phenazine methosulfate, enzyme and NO, or that system minus ascorbate. The mole ratio of thiol groups oxidized to NO reduced was observed to be 2.3 to 1.5 over a range of NO from 0.09 to 0.35 μ mol. A ratio of 1.0 was expected for the simple reduction of NO by 1-electron to N 2 O. The oxidation of additional thiol is attributed to the trapping of nitrosyl hydride (nitroxyl, NO − /NOH) by thiol.