Amphibacillus xylanus NADH oxidase and Salmonella typhimurium alkyl-hydroperoxide reductase flavoprotein components show extremely high scavenging activity for both alkyl hydroperoxide and hydrogen peroxide in the presence of S. typhimurium alkyl-hydroperoxide reductase 22-kDa protein component.
The flavoprotein NADH oxidase from Amphibacillus xylanus consumes oxygen to produce hydrogen peroxide. The amino acid sequence of this flavoprotein shows 51.2% identity to the F-52a component, denoted AhpF, of the alkyl-hydroperoxide reductase from Salmonella typhimurium. AhpF also catalyzes NADH-dependent hydrogen peroxide formation under aerobic conditions, albeit at a somewhat slower rate than the Amphibacillus protein. In the presence of the 22-kDa colorless component (AhpC) of the Salmonella alkyl-hydroperoxide reductase, both proteins catalyze the 4-electron reduction of oxygen to water. Both flavoproteins are active as AhpC reductases and mediate electron transfer, resulting in the NADH-dependent reduction of hydrogen peroxide and cumene hydroperoxide. Both enzymes' Km values for hydrogen peroxide, cumene hydroperoxide, and NADH are so low that they could not be determined accurately. Vmax values for hydrogen peroxide or cumene hydroperoxide reduction are > 10,000 min(-1) at 25 degrees C. These values are almost the same as the reduction rate of the flavoprotein component by NADH. The involvement in catalysis of a redox-active disulfide of the A. xylanus flavoprotein was shown by construction of three mutant enzymes, C337S, C340S, and C337S/C40SC337S/C340S. Very little activity for hydrogen peroxide or cumene hydroperoxide was found with the single mutants (C337S and C340S), and none with the double mutant (C337S/C340S). Analysis of the DNA sequence upstream of the Amphibacillus flavoprotein structural gene indicated the presence of a partial open reading frame homologous to the Salmonella ahpC structural gene (64.3% identical at the amino acid sequence level), suggesting that the NADH oxidase protein of A. xylanus is also part of a functional alkyl-hydroperoxide reductase system within these catalase-lacking bacteria.
Niimura
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Y
Department of Biological Chemistry, University of Michigan Medical School, Ann Arbor 48109-0606, USA.
Poole
L B
LB
Massey
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V
D000595:Amino Acid Sequence; D001419:Bacteria / Q000201:enzymology* / Q000235:genetics; D001421:Bacteria, Anaerobic / Q000201:enzymology / Q000235:genetics; D001555:Benzene Derivatives / Q000378:metabolism; D005420:Flavoproteins / Q000378:metabolism*; D016166:Free Radical Scavengers / Q000378:metabolism*; D005798:Genes, Bacterial; D006861:Hydrogen Peroxide / Q000378:metabolism; D008954:Models, Biological; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000235:genetics / Q000378:metabolism*; D009247:NADH, NADPH Oxidoreductases / Q000235:genetics / Q000378:metabolism*; D009994:Osmolar Concentration; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000378:metabolism*; D010101:Oxygen Consumption; D010544:Peroxidases; D010545:Peroxides / Q000378:metabolism*; D054464:Peroxiredoxins; D012486:Salmonella typhimurium / Q000201:enzymology
D001555:Benzene Derivatives; D005420:Flavoproteins; D016166:Free Radical Scavengers; D009097:Multienzyme Complexes; D010545:Peroxides; D006861:Hydrogen Peroxide; D010088:Oxidoreductases; D010544:Peroxidases; D054464:Peroxiredoxins; C023107:NADH oxidase; D009247:NADH, NADPH Oxidoreductases; C007164:cumene hydroperoxide