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Cloning and functional expression of a novel marmoset cytochrome P450 2D enzyme, CYP2D30: comparison with the known marmoset CYP2D19

Biochemical Pharmacology, ISSN: 0006-2952, Vol: 68, Issue: 1, Page: 165-175
2004
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Article Description

Using a primer set designed on the cDNA encoding the known marmoset cytochrome P450 2D19 (CYP2D19), a cDNA encoding a novel CYP2D enzyme (CYP2D30) was cloned from the liver of a female marmoset bred at Kyoto University (KYU). In addition, a cDNA encoding CYP2D19 was cloned from the liver of a female marmoset bred at Kagoshima University (KAU). CYP2D30 and CYP2D19 showed homologies of 93.6 and 93.4% in their nucleotide and amino acid sequences, respectively. Reverse transcription polymerase chain reaction (RT-PCR) and digestion with Nde I demonstrated that the KYU-marmoset liver contained mainly mRNA for CYP2D30, while the KAU-marmoset liver contained mainly mRNA for CYP2D19. Marmoset CYP2D30, like human CYP2D6, exhibited high debrisoquine (DB) 4-hydroxylase activity and relatively low DB 5-, 6-, 7- and 8-hydroxylase activities, whereas CYP2D19 lacked DB 4-hydroxylase but exhibited marked 5-, 6-, 7- and 8-hydroxylase activities. The two marmoset recombinant enzymes showed enantioselective bufuralol (BF) 1″-hydroxylase activities, similar to CYP2D6. BF 1″-hydroxylation by CYP2D30 exhibited product-enantioselectivity of (1″ R -OH-BF ⪡ 1″ S -OH-BF), similar to that observed with human CYP2D6, whereas CYP2D19 showed a reversed selectivity of (1″ R -OH-BF ≥ 1″ S -OH-BF). BF 1″-hydroxylation in marmoset liver microsomes from both sources was inhibited by antibodies raised against rat CYP2D1 in a concentration-dependent manner. A known inhibitor of CYP2D6, quinidine, effectively inhibited the BF 1″-hydroxylation activities in liver microsomal fractions prepared from KYU- and KAU-marmosets. These results suggest that CYP2D19 and CYP2D30 proteins can be expressed as functional enzymes in marmoset livers, although it is unresolved whether both enzymes coexist in the same marmoset liver.

Bibliographic Details

Hichiya, Hiroyuki; Kuramoto, Shino; Yamamoto, Shigeo; Shinoda, Sumio; Hanioka, Nobumitsu; Narimatsu, Shizuo; Asaoka, Kazuo; Miyata, Atsuro; Iwata, Shinichi; Nomoto, Masahiro; Satoh, Tetsuo; Kiryu, Kimio; Ueda, Nobuhiko; Naito, Shinsaku; Tucker, Geoffrey T; Ellis, S Wynne

Elsevier BV

Biochemistry, Genetics and Molecular Biology; Pharmacology, Toxicology and Pharmaceutics

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