Pregnenolone esterification in Saccharomyces cerevisiae - A potential detoxification mechanism

While studying the effect of steroids on the growth of the yeast Saccharomy ces cerevisiae, we found that pregnenolone was converted into the acetate e ater. This reaction was identified as a transfer of the acetyl group from a cetyl-CoA to the 3 beta-hydroxyl group of pregnenolone. The corresponding e nzyme, acetyl-CoA: pregnenolone acetyltransferase (APAT) is specific for De lta(5)- or Delta(4)-3 beta-hydroxysteroids and short-chain acyl-CoAs. The a pparent K-m for pregnenolone is approximate to 0.5 mu M. The protein associ ated with APAT activity was partially purified and finally isolated from an SDS/polyacrylamide gel. Tryptic peptides were generated and N-terminally s equenced. Two peptide sequences allowed the identification of an open readi ng frame (YGR177c, in the S. cerevisiae genome database) translating into a 62-kDa protein of hitherto unknown function. This protein encoded by a gen e known as ATF2 displays 37% identity with an alcohol acetyltransferase enc oded by the yeast gene ATF1. Disruption of ATF2 led to the complete elimina tion of APAT activity and consequently abolished the esterification of preg nenolone. In addition, a toxic effect of pregnenolone linked to the disrupt ion of ATF2 was observed. Pregnenolone toxicity is more pronounced when the atf 2-Delta mutation is introduced in a yeast strain devoid of the ATP-bin ding cassette transporters, PDR5 and SNQ2. Our results suggest that Atf2p ( APAT) plays an active role in the detoxification of 3 beta-hydroxysteroids in association with the efflux pumps Pdr5p and Snq2p.