Stereoselective Formation and Metabolism of 4-Hydroxy-Retinoic Acid Enantiomers by Cytochrome P450 Enzymes

• Published in: The Journal of biological chemistry 2012-12, Vol.287 (50), p.42223-42232
• Main Authors: Shimshoni, Jakob A., Roberts, Arthur G., Scian, Michele, Topletz, Ariel R., Blankert, Sean A., Halpert, James R., Nelson, Wendel L., Isoherranen, Nina
• Format: Article
• Language: English
• Subjects: Cytochrome P-450 Enzyme System - chemistry; Cytochrome P-450 Enzyme System - metabolism; Cytochrome P450; Enzyme Kinetics; Humans; Hydroxylation - physiology; Metabolism; Molecular Docking Simulation; Molecular Structure; Retinoid; Retinoid-binding Protein; Stereoisomerism; Tretinoin - analogs & derivatives; Tretinoin - pharmacokinetics
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M112.404475

All-trans-retinoic acid (atRA), the major active metabolite of vitamin A, plays a role in many biological processes, including maintenance of epithelia, immunity, and fertility and regulation of apoptosis and cell differentiation. atRA is metabolized mainly by CYP26A1, but other P450 enzymes such as CYP2C8 and CYP3As also contribute to atRA 4-hydroxylation. Although the primary metabolite of atRA, 4-OH-RA, possesses a chiral center, the stereochemical course of atRA 4-hydroxylation has not been studied previously. (4S)- and (4R)-OH-RA enantiomers were synthesized and separated by chiral column HPLC. CYP26A1 was found to form predominantly (4S)-OH-RA. This stereoselectivity was rationalized via docking of atRA in the active site of a CYP26A1 homology model. The docked structure showed a well defined niche for atRA within the active site and a specific orientation of the β-ionone ring above the plane of the heme consistent with stereoselective abstraction of the hydrogen atom from the pro-(S)-position. In contrast to CYP26A1, CYP3A4 formed the 4-OH-RA enantiomers in a 1:1 ratio and CYP3A5 preferentially formed (4R)-OH-RA. Interestingly, CYP3A7 and CYP2C8 preferentially formed (4S)-OH-RA from atRA. Both (4S)- and (4R)-OH-RA were substrates of CYP26A1 but (4S)-OH-RA was cleared 3-fold faster than (4R)-OH-RA. In addition, 4-oxo-RA was formed from (4R)-OH-RA but not from (4S)-OH-RA by CYP26A1. Overall, these findings show that (4S)-OH-RA is preferred over (4R)-OH-RA by the enzymes regulating atRA homeostasis. The stereoselectivity observed in CYP26A1 function will aid in better understanding of the active site features of the enzyme and the disposition of biologically active retinoids. Background: CYP26A1 is a critical enzyme in the metabolism of all-trans-retinoic acid (atRA). Results: CYP26A1 metabolizes atRA stereoselectively to (4S)-OH-RA, which corresponds to atRA orientation within the CYP26A1 active site. Conclusion: Preference for (4S)-OH-RA is conserved in atRA metabolism and in elimination of 4-OH-RA. Significance: Determination of stereoselectivity in atRA metabolism is important for better understanding of retinoid biology and CYP26A1 biochemistry.