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J.Health Sci., 50(5), 503-510, 2004
Changes in the Enzymatic Properties of CYP2D6 by the Substitution of Phenylalanine at Position
120 by Alanine
Kazufumi Masuda,a Hiroki
Hashimoto,a Keietsu
Tamagake,a Yukie
Okuda,b Daisuke
Tsuzuki,b Takashi
Isobe,b Hiroyuki
Hichiya,b Nobumitsu
Hanioka,b Shigeo
Yamamoto,c and Shizuo Narimatsu*, b
Laboratories of aPharmaceutical Physical Chemistry,
bHealth Chemistry, and cBiomolecular Sciences, Faculty of
Pharmaceutical Sciences, Okayama University, 1-1-1 Tsushima-naka, Okayama 700-8530, Japan
The functional roles of phenylalanine at position 120 (Phe-120) in the oxidation of bunitrolol (BTL),
debrisoquine (DB) and bufuralol (BF) by cytochrome P450 2D6 (CYP2D6) were examined using a yeast cell
expression system (Saccharomyces
cerevisiae AH-22 strain). The substitution of Phe-120 by alanine markedly
increased the activities of enantiomeric BTL 4-hydroxylase and DB 4-hydroxylase, whereas it did not
remarkably affect BF 1''-hydroxylase activities. Kinetic studies revealed that the substitution of Phe-120 by alanine
increased the Km and Vmax values for enantiomeric BTL 4-hydroxylation, but increased only the
Vmax value for DB 4-hydroxylation without changing the
Km value. Km and Vmax values for BF
1''-hydroxylation were similar between the mutant and the wild-type. The dissociation constants of the mutant calculated from the
binding spectra for BTL enantiomers were higher than those of the wild-type, suggesting that the substitution of
Phe-120 by alanine decreased the affinity of CYP2D6 for BTL enantiomers. These results indicate that Phe-120
has an important role in the oxidation of substrates by CYP2D6.
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