Abstract
Human cytochrome P450 4F2 (CYP4F2) catalyzes the initial omega-hydroxylation reaction in the metabolism of tocopherols and tocotrienols to carboxychromanols and is, to date, the only enzyme shown to metabolize vitamin E. The objective of this study was to characterize this activity, particularly the influence of key features of tocochromanol substrate structure. The influence of the number and positions of methyl groups on the chromanol ring, and of stereochemistry and saturation of the side chain, were explored using HepG2 cultures and microsomal reaction systems. Human liver microsomes and microsomes selectively expressing recombinant human CYP4F2 exhibited substrate activity patterns similar to those of HepG2 cells. Although activity was strongly associated with substrate accumulation by cells or microsomes, substantial differences in specific activities between substrates remained under conditions of similar microsomal membrane substrate concentration. Methylation at C5 of the chromanol ring was associated with markedly low activity. Tocotrienols exhibited much higher Vmax values than their tocopherol counterparts. Side chain stereochemistry had no effect on omega-hydroxylation of alpha-tocopherol (alpha-TOH) by any system. Kinetic analysis of microsomal CYP4F2 activity revealed Michaelis-Menten kinetics for alpha-TOH but allosteric cooperativity for other vitamers, especially tocotrienols. Additionally, alpha-TOH was a positive effector of omega-hydroxylation of other vitamers. These results indicate that CYP4F2-mediated tocopherol-omega-hydroxylation is a central feature underlying the different biological half-lives, and therefore biopotencies, of the tocopherols and tocotrienols.
Highlights
Human cytochrome P450 4F2 (CYP4F2) catalyzes the initial v-hydroxylation reaction in the metabolism of tocopherols and tocotrienols to carboxychromanols and is, to date, the only enzyme shown to metabolize vitamin E
The comparative metabolism of the various forms of vitamin E was examined in hepatocyte culture and in various microsomal systems, including human liver microsomes and microsomes from insect cells in which recombinant human cytochrome P450 tocopherol-vhydroxylase (CYP4F2) was selectively expressed
We previously identified the first enzyme catalyzing the biotransformation of vitamin E and demonstrated its preferential metabolism of a- and g-tocopherol (g-TOH) over a-TOH [30]
Summary
Human cytochrome P450 4F2 (CYP4F2) catalyzes the initial v-hydroxylation reaction in the metabolism of tocopherols and tocotrienols to carboxychromanols and is, to date, the only enzyme shown to metabolize vitamin E. The influence of the number and positions of methyl groups on the chromanol ring, and of stereochemistry and saturation of the side chain, were explored using HepG2 cultures and microsomal reaction systems. Human liver microsomes and microsomes selectively expressing recombinant human CYP4F2 exhibited substrate activity patterns similar to those of HepG2 cells. Side chain stereochemistry had no effect on v-hydroxylation of a-tocopherol (a-TOH) by any system. Kinetic analysis of microsomal CYP4F2 activity revealed Michaelis-Menten kinetics for a-TOH but allosteric cooperativity for other vitamers, especially tocotrienols. A-TOH was a positive effector of v-hydroxylation of other vitamers. These results indicate that CYP4F2-mediated tocopherol-vhydroxylation is a central feature underlying the different biological half-lives, and biopotencies, of the tocopherols and tocotrienols.—Sontag, T.
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