title: Export of cytochrome P450 105D1 to the periplasmic space of Escherichia coli.
abstract: CYP105D1, a cytochrome P450 from Streptomyces griseus, was appended at its amino terminus to the secretory signal of Escherichia coli alkaline phosphatase and placed under the transcriptional control of the native phoA promoter. Heterologous expression in E. coli phosphate-limited medium resulted in abundant synthesis of recombinant CYP105D1 that was translocated across the bacterial inner membrane and processed to yield authentic, heme-incorporated P450 within the periplasmic space. Cell extract and whole-cell activity studies showed that the periplasmically located CYP105D1 competently catalyzed NADH-dependent oxidation of the xenobiotic compounds benzo[a]pyrene and erythromycin, further revealing the presence in the E. coli periplasm of endogenous functional redox partners. This system offers substantial advantages for the application of P450 enzymes to whole-cell biotransformation strategies, where the ability of cells to take up substrates or discard products may be limited.
authors.affiliation: Institute of Biological Sciences, University of Wales Aberystwyth, Aberystwyth SY23 3DA, Wales, United Kingdom.
authors.forename: M A
authors.initials: MA
authors.lastname: Kaderbhai
authors.forename: C C
authors.initials: CC
authors.lastname: Ugochukwu
authors.forename: S L
authors.initials: SL
authors.lastname: Kelly
authors.forename: D C
authors.initials: DC
authors.lastname: Lamb
mesh_terms: D001426:Bacterial Proteins; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D010105:Oxygenases / Q000235:genetics* / Q000378:metabolism*; D019897:Periplasm / Q000201:enzymology*; D021381:Protein Transport; D013302:Streptomyces / Q000201:enzymology* / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D010105:Oxygenases; C122191:CYP105D1 protein, Streptomyces griseus