title: The nitric oxide reductase of Paracoccus denitrificans.
abstract: The nitric oxide (NO) reductase activity of the cytoplasmic membrane of Paracoccus denitrificans can be solubilized in dodecyl maltoside with good retention of activity. The solubilized enzyme lacks NADH-dependent activity, but can be assayed with isoascorbate plus 2,3,5,6-tetramethylphenylene-1,4-diamine as electron donor and with horse heart cytochrome c as mediator. Reduction of NO was measured with an amperomeric electrode. The solubilized enzyme could be separated from other electron-transport components, including the cytochrome bc1 complex and nitrite reductase, by several steps of chromatography. The purified enzyme had a specific activity of 11 mumols.min-1.mg of protein-1 and the Km(NO) was estimated as less than 10 microM. The enzyme formed N2O from NO with the expected stoichiometry. These observations support the view that NO reductase is a discrete enzyme that participates in the denitrification process. The enzyme contained both b- and c-type haems. The former was associated with a polypeptide of apparent molecular mass 37 kDa and the latter with a polypeptide of 18 kDa. Polypeptides of 29 and 45 kDa were also identified in the purified protein which showed variable behaviour on electrophoresis in polyacrylamide gels.
authors.affiliation: Department of Biochemistry, University of Oxford, U.K.
authors.forename: G J
authors.initials: GJ
authors.lastname: Carr
authors.forename: S J
authors.initials: SJ
authors.lastname: Ferguson
mesh_terms: D001205:Ascorbic Acid / Q000378:metabolism; D002384:Catalysis; D002462:Cell Membrane / Q000201:enzymology; D002845:Chromatography; D003574:Cytochrome c Group / Q000378:metabolism; D003902:Detergents; D004579:Electron Transport; D004591:Electrophoresis, Polyacrylamide Gel; D005960:Glucosides; D007700:Kinetics; D008970:Molecular Weight; D009243:NAD / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000302:isolation & purification* / Q000378:metabolism; D010231:Paracoccus denitrificans / Q000201:enzymology*; D012995:Solubility; D013053:Spectrophotometry; D013772:Tetramethylphenylenediamine / Q000378:metabolism
chemical_list: D003574:Cytochrome c Group; D003902:Detergents; D005960:Glucosides; D009243:NAD; C005056:isoascorbic acid; D009569:Nitric Oxide; C040358:dodecyl maltoside; D010088:Oxidoreductases; C034747:nitric-oxide reductase; D013772:Tetramethylphenylenediamine; D001205:Ascorbic Acid