CYP11A1 stimulates the hydroxylase activity of CYP11B1 in mitochondria of recombinant yeast in vivo and in vitro

In mammals, hydrocortisone synthesis from cholesterol is catalyzed by a set of five specialized enzymes, four of them belonging to the superfamily of cytochrome P-450 monooxygenases. A recombinant yeast expression system was recently developed for the CYPI11B1 (P450(11 beta)) enzyme, which performs the 11 beta hydroxylation of steroids such as 11-deoxycortisol into hydroco rtisone, one of the three mitochondrial cytochrome P-450 proteins involved in steroidogenesis in mammals. This heterologous system was used to test th e potential interaction between CYPI11B1 and CYP11A1 (P450(scc)), the mitoc hondrial cytochrome P-450 enzyme responsible for the side chain cleaving of cholesterol. Recombinant CYPI11B1 and CYP11A1 were targeted to Saccharomyc es cerevisiae mitochondria using the yeast cytochrome oxidase subunit 6 mit ochondrial presequence fused to the mature form of the two proteins. In yea st, the presence or CYP11A1 appears to improve 11 beta hydroxylase activity of CYP11B1 in vivo and in vitro. Fractionation experiments indicate the pr esence of the two proteins in the same membrane fractions, i.e. inner membr ane and contact sites of mitochondria. Thus, yeast mitochondria provide int eresting insights to study some molecular and cellular aspects of mammalian steroid synthesis. In particular, recombinant yeast should permit a better understanding of the mechanism permitting the synthesis of steroids (sex s teroids, minerallocorticoids and glucocorticoids) with a minimal set of enz ymes at physiological level, thus avoiding disease states.