Purification and kinetic properties of 3 beta-hydroxysteroid dehydrogenase from bovine adrenocortical microsomes.
3 beta-Hydroxysteroid dehydrogenase was purified from bovine adrenocortical microsomes and its properties were studied. The purified dehydrogenase gave a single homogeneous protein band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and showed no steroid delta 5-delta-4 isomerase activity. The molecular weight of the dehydrogenase was estimated to be 41,000 for the monomer and the isoelectric point was determined to be at pH 6.3. The Km values of the dehydrogenase were 6.2 microM for NAD+, 4.9 mM for NADP+, 2.0 microM for pregnenolone, and 5.3 microM for 17 alpha-hydroxypregnenolone. The mechanism of inhibition by trilostane of the dehydrogenase was also examined kinetically. The inhibition was found to be competitive, with Ki values of 0.14 microM for 17 alpha-hydroxypregnenolone and 0.38 microM for pregnenolone.
D006907:17-alpha-Hydroxypregnenolone / Q000378:metabolism; D015096:3-Hydroxysteroid Dehydrogenases / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D000302:Adrenal Cortex / Q000201:enzymology*; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D002417:Cattle; D013196:Dihydrotestosterone / Q000031:analogs & derivatives / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D007526:Isoelectric Point; D007700:Kinetics; D008861:Microsomes / Q000201:enzymology*; D008970:Molecular Weight; D009243:NAD / Q000378:metabolism; D009249:NADP / Q000378:metabolism; D011284:Pregnenolone / Q000378:metabolism
D013196:Dihydrotestosterone; D009243:NAD; D006907:17-alpha-Hydroxypregnenolone; D009249:NADP; D011284:Pregnenolone; D015096:3-Hydroxysteroid Dehydrogenases; C009954:trilostane