Export of cytochrome P450 105D1 to the periplasmic space of Escherichia coli

CYP105D1, a cytochrome P450 from Streptomyces griseus, was appended at its amino terminus to the secretory signal of Escherichia coli alkaline phospha tase and placed under the transcriptional control of the native phoA promot er. Heterologous expression in E. coli phosphate-limited medium resulted in abundant synthesis of recombinant CYP105D1 that was translocated across th e bacterial inner membrane and processed to yield authentic, heme-incorpora ted P450 within the periplasmic space. Cell extract and whole-cell activity studies shun;ed that the periplasmically located CYP105D1 competently cata lyzed NADH-dependent oxidation of the xenobiotic compounds benzo[a]pyrene a nd erythromycin, further revealing the presence in the E. coli periplasm of endogenous functional redox partners, This system offers substantial advan tages for the application of P450 enzymes to whole-cell biotransformation s trategies, H here the ability of cells to take up substrates or discard pro ducts may be limited.