The NAD(P)H:flavin oxidoreductase from Escherichia coli as a source of superoxide radicals.
The NAD(P)H:flavin oxidoreductase (encoded by the fre gene) of Escherichia coli is a soluble enzyme which, under aerobic conditions and together with NAD(P)H and flavins, generates superoxide radicals selectively. This was demonstrated from spin trapping experiments and from the ability of the flavin reductase to achieve a superoxide dismutase (SOD)-sensitive reduction of cytochrome c. The participation of the flavin reductase to O2-. generation in E. coli cells has been studied. Superoxide production in dialyzed cytosolic fraction of SOD-deficient E. coli was stimulated by the addition of flavins. There was no stimulation in soluble extracts of flavin reductase-deficient strains. Moreover, using fusions of sodA promoter to lacZ, we showed that sodA transcription was diminished in flavin reductase-deficient E. coli and that the induction of MnSOD by flavin reductase was SoxRS-independent. These results suggest that the flavin reductase might: (i) in vivo, be an important cytosolic site of O2-. generation; (ii) in vitro, serve as a simple, efficient, and selective O2-. generator.
Gaudu
P
P
Laboratoire d'Etudes Dynamiques et Structurales de la Sélectivité, Unité de Recherche Associée au Centre National de la REcherche Scientifique 332, Université Joseph Fourier, Grenoble, France.
Touati
D
D
Nivière
V
V
Fontecave
M
M
D003497:Cyclic N-Oxides; D004578:Electron Spin Resonance Spectroscopy; D004790:Enzyme Induction; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D038181:FMN Reductase; D005486:Flavin Mononucleotide / Q000378:metabolism; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D009243:NAD / Q000378:metabolism; D009247:NADH, NADPH Oxidoreductases / Q000235:genetics / Q000378:metabolism*; D009249:NADP / Q000378:metabolism; D010084:Oxidation-Reduction; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012256:Riboflavin / Q000494:pharmacology; D013113:Spin Labels; D013482:Superoxide Dismutase / Q000096:biosynthesis / Q000235:genetics; D013481:Superoxides / Q000378:metabolism*; D014158:Transcription, Genetic; D001616:beta-Galactosidase / Q000096:biosynthesis / Q000378:metabolism
D003497:Cyclic N-Oxides; D011993:Recombinant Fusion Proteins; D013113:Spin Labels; D009243:NAD; D013481:Superoxides; D009249:NADP; C017245:5,5-dimethyl-1-pyrroline-1-oxide; D005486:Flavin Mononucleotide; D013482:Superoxide Dismutase; D038181:FMN Reductase; D009247:NADH, NADPH Oxidoreductases; D001616:beta-Galactosidase; D012256:Riboflavin