Point mutation of cytochrome P450 2A6 (a polymorphic variant CYP2A6.25) confers new substrate specificity towards flavonoids |
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| Authors: | Tomohide Uno Chika Ogura Chiho Izumi Masahiko Nakamura Takeshi Yanase Hiroshi Yamazaki Hitoshi Ashida Kengo Kanamaru Hiroshi Yamagata Hiromasa Imaishi |
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| Affiliation: | 1. Laboratory of Biological Chemistry, Department of Biofunctional Chemistry, Faculty of Agriculture, Kobe University, Kobe, Hyogo, Japan;2. Department of Bioscience and Biotechnology, Faculty of Bioenvironmental Science, Kyoto Gakuen University, Kameoka, Kyoto, Japan;3. Laboratory of Drug Metabolism and Pharmacokinetics, Showa Pharmaceutical University, Machida, Tokyo, Japan;4. Laboratory of Biochemistry Frontiers, Graduate School of Agricultural Science, Kobe University, Kobe, Hyogo, Japan;5. Functional Analysis of Environmental Genes, Research Center for Environmental Genomics, Kobe University, Kobe, Hyogo, Japan |
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| Abstract: | CYP2A6 is a major hepatic member of the cytochrome P450 family in humans. Much variation in CYP2A6 levels and activity can be attributed to genetic polymorphisms of this gene. CYP2A6*25 comprises an amino acid substitution, F118L. To clarify the effect of the leucine substitution at position 118 in CYP2A6.25, this variant, wild type CYP2A6 and three additional variants consisting of artificial mutations at the substrate binding site (position 481) suggested by earlier reports using random mutagenesis studies [CYP2A6.1, CYP2A6.25, CYP2A6.1(F118A), CYP2A6.1(A481G) and CYP2A6.25(A481G)], were co‐expressed with NADPH‐cytochrome P450 reductase in E. coli. The hydroxylase activity of these variants toward 7‐ethoxycoumarin, coumarin, flavone, α‐naphthoflavone, flavanone and hydroxyflavanone were examined. All the mutants had lower activities for coumarin 7‐hydroxylation than the wild type. All the mutants showed higher activities for flavone and α‐naphthoflavone compared with CYP2A6.1. CYP2A6.1 had the highest flavanone 2′‐hydroxylase activity, whereas CYP2A6.25 had the highest 6‐ and 4′‐hydroxylase activities. CYP2A6.1(F118A), CYP2A6.1(A481G) and CYP2A6.25(A481G) had higher flavanone 3′‐hydroxylase activities than CYP2A6.1 and CYP2A6.25. Furthermore, 4′‐hydroxyflavanone was metabolized by CYP2A6.25. These results indicate that the CYP2A6.25 mutation confers new substrate specificity towards flavonoids. Copyright © 2015 John Wiley & Sons, Ltd. |
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| Keywords: | P450 monooxygenase polymorphism flavanone CYP2A6 flavone |
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