Molecular cloning of monkey P450 1A1 cDNA and expression in yeast.
Monkey P450 1A1 cDNA (MKah1) was isolated from the lambda gt11 cDNA library of a liver from a 3-methylcholanthrene (3MC)-treated crab-eating monkey using a dog P450 1A1 cDNA fragment as a probe. MKah1 was 2453 bp long and contained an entire coding region for a polypeptide of 512 residues. The nucleotide and deduced amino acid sequences of MKah1 displayed 95% and 94% identity with those of the human P450 1A1 gene, respectively. Even in the 3' noncoding region, MKah1 showed 94% homology with human P450 1A1, whereas it showed less than 69% homology with other mammalian P450 1A1. Monkey P450 1A1 mRNA was not detectable in untreated livers, but was induced by polychlorinated biphenyl and 3MC. The expression plasmid (designated as pMKC-1) was constructed by introduction of the coding region of MKah1 into a yeast expression vector (pAM82) containing the promoter of acid phosphatase (APase). Northern blot analysis revealed that monkey P450 1A1 mRNA was expressed in yeast under the control of the APase promoter. Microsomes from yeast transformed by pMKC-1 catalyzed 7-ethoxycoumarin O-deethylation, benzo(a)pyrene hydroxylation and the mutagenic activation of 2-amino-3-methyl-imidazo[4,5-f]quinoline (IQ), 3-amino-1-methyl-5H-pyrido(4,3-b)-indole acetate (Trp-P-2) and 2-amino-6-methyldipyrido(1,2-a:3',2'-d)imidazole acetate (Glu-P-1).
Komori
M
M
Division of Analytical Biochemistry, Faculty of Pharmaceutical Sciences, Hokkaido University, Japan.
Kikuchi
O
O
Kitada
M
M
Kamataki
T
T
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D003062:Codon / Q000032:analysis; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics*; D004247:DNA / Q000302:isolation & purification; D005260:Female; D000882:Haplorhini / Q000235:genetics*; D008099:Liver / Q000201:enzymology; D008748:Methylcholanthrene; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000302:isolation & purification; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*
D003062:Codon; D012333:RNA, Messenger; D008748:Methylcholanthrene; D004247:DNA; D003577:Cytochrome P-450 Enzyme System