By using the polymerase chain reaction technique combined with restriction enzyme fragment length polymorphism (PCR-RFLP), a novel polymorphism of CYP2A6, CYP2A6*6, was detected in 0.4% of the Japanese population. To study the enzymatic properties of the CYP2A6.6 protein with a single amino acid substitution of arginine 128 to glutamine, both this isozyme and the CYP2A6.1 protein (wild-type) were produced in insect cells using a baculovirus system. Coumarin 7-hydroxylation, which reflects CYP2A6 activity, was significantly reduced (one-eighth of normal) in cell lysate from CYP2A6*6-transfected Sf9 cells compared with that lysate from CYP2A6*1-transfected cells. To clarify the mechanism of inactivation of the CYP2A6.6 enzyme, the heme content and reduced CO difference spectrum were examined. Although CYP2A6.6 retained about one-half the heme content of CYP2A6.1, the reduced CO-bound Soret peak was completely lost. These results suggest that the inactivation of CYP2A6.6 is mainly due to disordering of the holoprotein structure rather than a failure of heme incorporation.
CITATION STYLE
Kitagawa, K., Kunugita, N., Kitagawa, M., & Kawamoto, T. (2001). CYP2A6*6, a Novel Polymorphism in Cytochrome P450 2A6, Has a Single Amino Acid Substitution (R128Q) that Inactivates Enzymatic Activity. Journal of Biological Chemistry, 276(21), 17830–17835. https://doi.org/10.1074/jbc.M009432200
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