CYP201A2, a cytochrome P450 from Rhodopseudomonas palustris, plays a key role in the biodegradation of tributyl phosphate.
Tributyl phosphate (TBP) is a toxic organophosphorous compound widely used in nuclear fuel processing and chemical industries. Rhodopseudomonas palustris, one of the most metabolically versatile photosynthetic bacteria, is shown here to degrade TBP efficiently under photosynthetic conditions. This study shows that this O(2)- and NADPH/FMNH(2)-dependent process was also catalyzed when TBP was incubated with membrane-associated proteins extracted from this strain. The effects of several regulators of cytochrome P450 activity on the TBP consumption suggest a key role for a cytochrome P450 in this process. Disruption of the rpa0241 gene encoding a putative cytochrome P450 led to a 60% decrease of the TBP catabolism, whereas reintroducing the gene in the mutant restored the wild-type phenotype. The rpa0241 gene was expressed and purified in Escherichia coli. Characterization by UV-visible spectroscopy of the purified recombinant membrane-bound protein (CYP201A2) encoded by the rpa0241 gene revealed typical spectral characteristics of cytochrome P450 with a large spin state change of the heme iron associated with binding of TBP (K (d) approximately 65 microM). It is proposed that CYP201A2 catalyzes the initial step of the biodegradation process of TBP.
Berne
Cécile
C
DSV/IBEB/SBVME/LBC, Unité Mixte de Recherche 6191, Centre National de la Recherche Scientifique/CEA/Univ. Aix-Marseille, CEA Cadarache, Saint Paul lez Durance, France.
Pignol
David
D
Lavergne
Jérôme
J
Garcia
Daniel
D
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D002384:Catalysis; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005816:Genetic Complementation Test; D007700:Kinetics; D009154:Mutation; D009249:NADP / Q000378:metabolism; D010755:Organophosphates / Q000378:metabolism*; D012241:Rhodopseudomonas / Q000201:enzymology* / Q000235:genetics