Cloning and expression analysis of a new member of the cytochrome P450, CYP2A15 from the Chinese hamster, encoding testosterone 7alpha-hydroxylase.
We cloned a new cytochrome P450 cDNA encoding testosterone 7alpha-hydroxylase in the Chinese hamster, designated CYP2A15 which shares significant amino acid sequence homology with members of the CYP2A subfamily. The CYP2A15 cDNA was isolated by screening a liver cDNA library and the sequence contains an open reading frame of 1482 nucleotides encoding a polypeptide of 493 amino acids with a calculated molecular mass of 56,295 Da. This is flanked by a 5'-untranslated region of 2 bp and a 3' untranslated region of 191 bp including the poly(A) tail. We determined the catalytic activity of CYP2A15 using microsomes obtained by transient expression of its cDNA in transfected COS-7 cells. The heterologously expressed CYP2A15 was found to hydroxylate testosterone at position 7alpha in a reconstituted system. RT-PCR experiments revealed that the mRNA of CYP2A15 was expressed in liver, but not detected in kidney, lung, or small intestine. The expression of CYP2A15 mRNA was slightly induced by treatment with either rifampicin or 3-methylcholanthrene.
Kurose
K
K
Department of Pharmaceutical Sciences, National Institute of Public Health, 4-6-1 Shirokanedai, Minato-ku, Tokyo, 108-8638, Japan. kurose@iph.go.jp
Isozaki
E
E
Tohkin
M
M
Fukuhara
M
M
D000595:Amino Acid Sequence; D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases; D001483:Base Sequence; D003001:Cloning, Molecular; D006224:Cricetinae; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics* / Q000378:metabolism; D018076:DNA, Complementary / Q000235:genetics; D005260:Female; D015870:Gene Expression; D006900:Hydroxylation; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D008099:Liver / Q000187:drug effects / Q000201:enzymology; D008297:Male; D008748:Methylcholanthrene / Q000494:pharmacology; D008861:Microsomes / Q000201:enzymology; D008969:Molecular Sequence Data; D010634:Phenobarbital / Q000494:pharmacology; D010802:Phylogeny; D011994:Recombinant Proteins / Q000378:metabolism; D012293:Rifampin / Q000494:pharmacology; D013250:Steroid Hydroxylases / Q000235:genetics* / Q000378:metabolism; D013739:Testosterone / Q000378:metabolism*; D014018:Tissue Distribution
D018076:DNA, Complementary; D007527:Isoenzymes; D011994:Recombinant Proteins; D013739:Testosterone; D008748:Methylcholanthrene; D003577:Cytochrome P-450 Enzyme System; D013250:Steroid Hydroxylases; D001189:Aryl Hydrocarbon Hydroxylases; C022657:steroid hormone 6-beta-hydroxylase; C022724:testosterone 7-alpha-hydroxylase, hamster; D012293:Rifampin; D010634:Phenobarbital