title: CYP11A1 stimulates the hydroxylase activity of CYP11B1 in mitochondria of recombinant yeast in vivo and in vitro.
abstract: 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 CYP11B1 (P45011beta) enzyme, which performs the 11beta hydroxylation of steroids such as 11-deoxycortisol into hydrocortisone, one of the three mitochondrial cytochrome P-450 proteins involved in steroidogenesis in mammals. This heterologous system was used to test the potential interaction between CYP11B1 and CYP11A1 (P450scc), the mitochondrial cytochrome P-450 enzyme responsible for the side chain cleaving of cholesterol. Recombinant CYP11B1 and CYP11A1 were targeted to Saccharomyces cerevisiae mitochondria using the yeast cytochrome oxidase subunit 6 mitochondrial presequence fused to the mature form of the two proteins. In yeast, the presence of CYP11A1 appears to improve 11beta hydroxylase activity of CYP11B1 in vivo and in vitro. Fractionation experiments indicate the presence of the two proteins in the same membrane fractions, i.e. inner membrane and contact sites of mitochondria. Thus, yeast mitochondria provide interesting 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 steroids, mineralocorticoids and glucocorticoids) with a minimal set of enzymes at physiological level, thus avoiding disease states.
authors.affiliation: Transgene S.A., Strasbourg, France.
authors.forename: G
authors.initials: G
authors.lastname: Cauet
authors.forename: D
authors.initials: D
authors.lastname: Balbuena
authors.forename: T
authors.initials: T
authors.lastname: Achstetter
authors.forename: B
authors.initials: B
authors.lastname: Dumas
mesh_terms: D002784:Cholesterol / Q000378:metabolism; D002786:Cholesterol Side-Chain Cleavage Enzyme / Q000235:genetics / Q000378:metabolism*; D003350:Cortodoxone / Q000378:metabolism; D006854:Hydrocortisone / Q000096:biosynthesis; D006900:Hydroxylation; D007425:Intracellular Membranes / Q000201:enzymology; D008928:Mitochondria / Q000201:enzymology*; D011994:Recombinant Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae; D013252:Steroid 11-beta-Hydroxylase / Q000235:genetics / Q000378:metabolism*
chemical_list: D011994:Recombinant Proteins; D002784:Cholesterol; D013252:Steroid 11-beta-Hydroxylase; D002786:Cholesterol Side-Chain Cleavage Enzyme; D003350:Cortodoxone; D006854:Hydrocortisone