3-beta-HSD is a bifunctional enzyme, that catalyzes the oxidation and isomerization of cholesterol, pregnenolone, and dehydroepiandrosterone (DHEA) into cholest-4-en-3-one, progesterone, and androsterone, respectively. a 3beta-hydroxy-Delta(5)-steroid + NAD(+) = a 3-oxo-Delta(5)-steroid + H(+) + NADH cholesterol + NAD(+) = cholest-5-en-3-one + H(+) + NADH NAD(+) + pregnenolone = H(+) + NADH + pregn-5-ene-3,20-dione 3beta-hydroxyandrost-5-en-17-one + NAD(+) = androst-5-ene-3,17-dione + H(+) + NADH a 3-oxo-Delta(5)-steroid = a 3-oxo-Delta(4)-steroid cholest-5-en-3-one = cholest-4-en-3-one pregn-5-ene-3,20-dione = progesterone androst-5-ene-3,17-dione = androst-4-ene-3,17-dione Inhibited by EDTA, zinc, silver, copper, and activated by magnesium and calcium. A competitive inhibition is observed with NAD at concentrations higher than 5.6 mM, and with trilostane (3,17-dihydroxy-4,5-epoxyandrost-2-ene-2-carbonitrile). Optimum pH is between 8.5-9.5. No catalytic activity is detected below pH 7.5. Lipid metabolism; steroid biosynthesis. Monomer. Disruption reduces the cholesterol oxidation activity at least 90-fold. Belongs to the 3-beta-HSD family.