Studies on NADH oxidase and alkyl hydroperoxide reductase produced by Porphyromonas gingivalis.
Enzymes that detoxify oxygen or oxygen radicals are important to anaerobic microorganisms that inhabit oxygenated environments. In previous studies we have determined that Porphyromonas gingivalis W50 cell extracts possess NADH oxidase-like activity, which increases slightly under oxygenated conditions. The aim of this study was to characterize the protein responsible for this activity in order to establish whether it protects the microorganism from oxidative stress. Protein purification based on NADH oxidase activity did not isolate a conventional NADH oxidase. Instead, the NADH oxidase activity was found to be associated with a FAD-dependent enzyme identified as 4-hydroxybutyryl-CoA dehydratase (AbfD). The biological significance of this activity with respect to protection against oxidative stress is not clear; hydrogen peroxide (H2O2) was present after completion of the NADH oxidase assay with the purified protein. Northern blot analysis, examining the expression of other proteins likely to function as NADH oxidases/peroxidases in P. gingivalis, revealed the transcription of a protein similar to alkyl-hydroperoxide reductase (AhpF-C), which could serve as an NADH oxidase and H2O2-detoxification system. AhpF is transcribed in a polycystronic way with its neighboring gene, which encodes for the coupling protein AhpC. No transcript could be detected for the closest match to an NADH oxidase identified in the P. gingivalis genome sequence. In conclusion, P. gingivalis seems to lack a protective NADH oxidase but AhpF-C could contribute to its moderate tolerance to reactive oxygen species by metabolizing H2O2.
Diaz
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PI
Microbiology Laboratory, Dental School, The University of Adelaide, Australia.
Zilm
P S
PS
Wasinger
V
V
Corthals
G L
GL
Rogers
A H
AH
D004591:Electrophoresis, Polyacrylamide Gel; D005182:Flavin-Adenine Dinucleotide / Q000378:metabolism; D005798:Genes, Bacterial / Q000235:genetics; D006801:Humans; D006836:Hydro-Lyases / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000378:metabolism; D013058:Mass Spectrometry; D009097:Multienzyme Complexes / Q000302:isolation & purification / Q000378:metabolism*; D009247:NADH, NADPH Oxidoreductases / Q000302:isolation & purification / Q000378:metabolism*; D018384:Oxidative Stress / Q000502:physiology; D010544:Peroxidases / Q000302:isolation & purification / Q000378:metabolism*; D054464:Peroxiredoxins; D010741:Phospholipases A / Q000302:isolation & purification / Q000378:metabolism*; D016966:Porphyromonas gingivalis / Q000201:enzymology*; D017382:Reactive Oxygen Species / Q000378:metabolism; D014158:Transcription, Genetic / Q000235:genetics
D009097:Multienzyme Complexes; D017382:Reactive Oxygen Species; D005182:Flavin-Adenine Dinucleotide; D006861:Hydrogen Peroxide; D010544:Peroxidases; D054464:Peroxiredoxins; C023107:NADH oxidase; D009247:NADH, NADPH Oxidoreductases; D010741:Phospholipases A; C066370:4-hydroxybutyryl-CoA dehydratase; D006836:Hydro-Lyases