Human NADPH-P450 oxidoreductase: complementary DNA cloning, sequence and vaccinia virus-mediated expression and localization of the CYPOR gene to chromosome 7.
The cDNA containing the full coding sequence of human NADPH-P450 oxidoreductase was isolated and completely sequenced. The cDNA contained 2398 base pairs, including 9 and 358 base pairs of 5' and 3' noncoding sequences, respectively. The human NADPH-P450 oxidoreductase protein deduced from the cDNA has 677 amino acids, with a calculated molecular weight of 76,656. The cDNA nucleotide and deduced amino acid sequences displayed 83 and 92% similarities, respectively, with those of the rat NADPH-P450 oxidoreductase. By use of somatic cell hybrids, the NADPH-P450 oxidoreductase gene was regionally localized to human chromosome 7 (7p15-q35). The levels of NADPH-P450 oxidoreductase protein and mRNA were analyzed in 13 human liver specimens and less than 3-fold variation was found among the different livers. The NADPH-P450 oxidoreductase cDNA was inserted into vaccinia virus and expressed in cell culture. The cDNA-expressed enzyme was active in reducing the electron acceptor cytochrome c. In addition, the NADPH-P450 oxidoreductase stimulated the enzymatic activity of vaccinia virus-expressed human P3(450) when both recombinant viruses were used to coinfect human cells in culture. An approximate equal mole level of NADPH-P450 oxidoreductase and P3(450) was required to achieve maximal activity for both ethoxycoumarin O-deethylase and aryl hydrocarbon hydroxylase.
Yamano
S
S
Laboratory of Molecular Carcinogenesis, National Cancer Institute, Bethesda, Maryland 20892.
Aoyama
T
T
McBride
O W
OW
Hardwick
J P
JP
Gelboin
H V
HV
Gonzalez
F J
FJ
D000818:Animals; D001483:Base Sequence; D002874:Chromosome Mapping; D002897:Chromosomes, Human, Pair 7; D003001:Cloning, Molecular; D003577:Cytochrome P-450 Enzyme System / Q000032:analysis / Q000235:genetics*; D004247:DNA / Q000032:analysis*; D006801:Humans; D008969:Molecular Sequence Data; D009251:NADPH-Ferrihemoprotein Reductase / Q000032:analysis / Q000235:genetics*; D051381:Rats; D014616:Vaccinia virus / Q000201:enzymology*
D004247:DNA; D003577:Cytochrome P-450 Enzyme System; D009251:NADPH-Ferrihemoprotein Reductase