CYP2A13-catalysed coumarin metabolism: comparison with CYP2A5 and CYP2A6

Abstract

1. We investigated the total metabolism of coumarin by baculovirus (BV)-expressed CYP2A13 and compared it with metabolism by BV-expressed CYP2A6. The major coumarin metabolite formed by CYP2A13 was 7-hydroxycoumarin, which accounted for 43% of the total metabolism. The product of 3,4-epoxidation, o -hydroxyphenylacetaldehyde (o -HPA), accounted for 30% of the total metabolites. 2. The K m and V max for CYP2A13-mediated coumarin 7-hydroxylation were 0.48 ± 0.07 µm and 0.15 ± 0.006 nmol min − 1 nmol − 1 CYP, respectively. The V max of coumarin 7-hydroxylation by CYP2A13 was about 16-fold lower than that of CYP2A6, whereas the K m was 10-fold lower. 3. In the mouse, there were two orthologues for CYP2A6: CYP2A4 and CYP2A5, which differed by only 11 amino acids. However, CYP2A5 is an efficient coumarin 7-hydroxylase, where as CYP2A4 is not. We report here that BV-expressed CYP2A4 metabolizes coumarin by 3,4-epoxidation. Two products of the 3,4-epoxidation pathway, o -HPA and o -hydroxyphenylacetic acid (o -HPAA), were detected by radioflow HPLC. 4. The K m and V max for the coumarin 3,4-epoxidation by CYP2A4 were 8.7 ± 3.6 µm and 0.20 ± 0.04 nmol min − 1 nmol − 1 CYP, respectively. Coumarin 7-hydroxylation by CYP2A5 was more than 200 times more efficient than 3,4 epoxidation by CYP2A4.