In an E. coli expression system for human cytochrome P450 3A7 (CYP3A7), holo-CYP3A7 was not expressed as judged by CO-difference spectra, although apo-CYP3A7 was clearly detected by Western blot analysis. Unlike CYP3A7, CYP3A4 was expressed efficiently as a hemoprotein in E. coli transformed with a CYP3A4 expression plasmid. To achieve the high yield of the holo-CYP3A7 in E. coli, we examined a causal residue(s) preventing the expression of the holo-CYP3A7 using the chimeric gene of CYP3A4 with CYP3A7. It was found that the region between residues 405 and 503 of CYP3A7 was responsible for the prevention of the holo-CYP3A7 expression in E. coli. Among amino acids examined, substitution of Thr at position 485 in CYP3A7 with Pro, which is at the corresponding position of CYP3A4, resulted in an increase in the amount of holo-CYP3A7. The Thr residue was adjacent to the heme-binding region of CYP3A7. Thus, it appeared that the incorporation of heme into CYP3A7 was possibly affected by this particular amino acid residue. Moreover, holo-CYP3A7 was expressed efficiently when CYP3A7 was co-expressed with molecular chaperone GroEL, known to assist the correct folding of unfolded proteins. Dehydroepiandrosterone 16alpha-hydroxylation was catalyzed by CYP3A7 expressed in the presence of GroEL.
Copyright 2000 Academic Press.