cyp7b1 catalyses the 7alpha-hydroxylation of dehydroepiandrosterone and 25-hydroxycholesterol in rat prostate.
Dehydroepiandrosterone (DHEA) is the most prominent circulating steroid in humans, and it is a precursor for sex-steroid synthesis in peripheral tissues, including the prostate. Recently, enzyme-mediated pre-receptor metabolism has been recognized as a key step in determining steroid action in vivo. Hydroxylation of 3beta-steroids at the 7alpha-position has been reported in rat and human prostate to be a major inhibitory pathway to sex-steroid synthesis/action. However, the molecular identity of the enzyme responsible is so far unknown. We recently described a novel cytochrome P450 enzyme, cyp7b1, strongly expressed in the hippocampus of rodent brain, which catalyses the metabolism of DHEA, pregnenolone and 25-hydroxycholesterol to 7alpha-hydroxy products. In the light of this new enzyme, we have examined its possible role in 7alpha-hydroxylation conversion in rat prostate. NADPH-dependent 7alpha-hydroxylation was confirmed for 3beta-hydroxysteroids including DHEA and androstenediol, as well as 25-hydroxycholesterol. Kinetic analysis yielded an apparent K(m) of 14+/-1 microM for 7alpha-hydroxylation of DHEA in the prostate gland, a value similar to that recorded for recombinant cyp7b1 enzyme [13.6 microM; Rose, Stapleton, Dott, Kieny, Best, Schwarz, Russell, Bjoorkheim, Seckl and Lathe (1997) Proc. Natl. Acad. Sci. U.S.A. 94, 4925-4930]. The V(max) value of the prostate was 46+/-2 pmol/min per mg, and this activity was inhibited by clotrimazole, a P450-enzyme blocker. Moreover, RNA analysis (reverse-transcription PCR, Northern blotting and in situ hybridization) revealed a high expression of cyp7b1 mRNA in the rat prostate, restricted to the epithelium, suggesting that cyp7b1 catalyses oxysterol 7alpha-hydroxylation in the prostate gland.
Martin
C
C
Molecular Endocrinology, Molecular Medicine Centre, Western General Hospital and Centre for Neuroscience, University of Edinburgh, Edinburgh EH4 2XU, Scotland, UK. cecile.martin@ed.ac.uk
Bean
R
R
Rose
K
K
Habib
F
F
Seckl
J
J
D000818:Animals; D002384:Catalysis; D002790:Cholesterol 7-alpha-Hydroxylase / Q000378:metabolism; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D003687:Dehydroepiandrosterone / Q000378:metabolism*; D004848:Epithelium / Q000201:enzymology; D006888:Hydroxycholesterols / Q000378:metabolism*; D006900:Hydroxylation; D017403:In Situ Hybridization; D007700:Kinetics; D008297:Male; D011467:Prostate / Q000201:enzymology* / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D013250:Steroid Hydroxylases / Q000235:genetics / Q000378:metabolism*; D013379:Substrate Specificity
D006888:Hydroxycholesterols; D012333:RNA, Messenger; D003687:Dehydroepiandrosterone; C007997:25-hydroxycholesterol; D003577:Cytochrome P-450 Enzyme System; D013250:Steroid Hydroxylases; C115867:oxysterol 7-alpha-hydroxylase; D002790:Cholesterol 7-alpha-Hydroxylase