High catalytic activity of human cytochrome P450 co-expressed with human NADPH-cytochrome P450 reductase in Escherichia coli.
Forms of human cytochrome P450 (P450 or CYP), such as CYP1A1, CYP1A2, CYP2A6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, and CYP3A4, were expressed or co-expressed together with human NADPH-P450 reductase in Escherichia coli. When P450 was expressed alone in E. coli, the expression level of holo-P450 ranged from 310 to 1620 nmol/L of culture. The expression level of holo-P450 decreased by co-expression with the reductase, and the level ranged from 66 to 381 nmol/L of culture. The expression level of the reductase varied depending on the forms of P450 co-expressed, and ranged from 204 to 937 U/L of culture. We assayed the catalytic activity of P450 using E. coli cells disrupted by freeze-thaw. When co-expressed with the reductase, human P450 catalyzed the oxidation of representative substrates at efficient rates. The rates appeared comparable to the reported activities of P450 in a reconstituted system containing purified preparations of P450 and the reductase.
Iwata
H
H
Division of Drug Metabolism, Faculty of Pharmaceutical Sciences, Hokkaido University, Sapporo, Japan.
Fujita
K
K
Kushida
H
H
Suzuki
A
A
Konno
Y
Y
Nakamura
K
K
Fujino
A
A
Kamataki
T
T
D000595:Amino Acid Sequence; D002384:Catalysis; D003001:Cloning, Molecular; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli; D006801:Humans; D008969:Molecular Sequence Data; D009247:NADH, NADPH Oxidoreductases / Q000235:genetics / Q000378:metabolism*; D009251:NADPH-Ferrihemoprotein Reductase; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
D011994:Recombinant Proteins; D003577:Cytochrome P-450 Enzyme System; D009247:NADH, NADPH Oxidoreductases; D009251:NADPH-Ferrihemoprotein Reductase