UDP-Glucuronosyltransferases in Conjugation of 5α- and 5β-Androstane Steroids

We have examined the glucuronidation of androsterone (5 alpha-androstane-3 alpha-ol-17-one), etiocholanolone (5 beta-androstane-3 alpha-ol-17-one), 5 alpha-androstane-3 alpha-, 17 beta-diol (5 alpha-diol), and 5 beta-androstane-3 alpha-, 17 beta-diol (5 beta-diol) by 19 recombinant human UDP-glucuronosyltransferases (UGTs). The results reveal large differences in stereo-and regioselectivity between UGT2B7, UGT2B15, and UGT2B17. UGT2B7 conjugated all four androgens at the 3-OH but not at the 17-OH that is available in both diols. UGT2B7 exhibited a higher glucuronidation rate toward the steroids with a flat backbone, androsterone and 5 alpha-diol, compared with etiocholanolone and 5 beta-diol, which have a bent backbone. UGT2B17 readily glucuronidated androsterone and, particularly, etiocholanolone at the 3-OH, but in the two diols it exhibited high preference for the 17-OH and low glucuronidation rate at the 3-OH. UGT2B15 did not glucuronidate any of the studied four androgens at the 3-OH, but it did conjugate both diols at the 17-OH, with a clear preference for 5 alpha-diol. Of the UGT1A subfamily, only UGT1A4 catalyzed the glucuronidation of androsterone and 5 alpha-diol at measurable rates, even if low. UGT2A1 and UGT2A2 glucuronidated most compounds in this study, but mostly at rather low rates. An exception was the glucuronidation of etiocholanolone by UGT2A1 that revealed a very low substrate affinity in combination with very high V-max value. The results shed new light on the substrate selectivity of individual UGTs in steroid glucuronidation. In addition they bear implications for doping analyses and its dependence of genetic polymorphism because testosterone is a precursor in the biosynthesis of these four androgens, whereas the contribution of UGT2B17 to their glucuronidation varies greatly.