Kinetic and Stereochemical Characterization of Hamster Liver 3α-Hydroxysteroid Dehydrogenase and 3α (17β)-Hydroxysteroid Dehydrogenase

抄録

The kinetic mechanism of NADP⁺ -dependent 3α-hydroxysteroid dehydrogenase and NAD⁺ -dependent 3α(17β)-hydroxysteroid dehydrogenase, purified from hamster liver cytosol, was studied in both directions. For 3α-hydroxysteroid dehydrogenase, the initial velocity and product inhibition studies indicated that the enzyme reaction sequence is ordered with NADP+ binding to the free enzyme and NADP⁺ being the last product to be released. Inhibition patterns by Cibacron blue and hexestrol, and binding studies of coenzyme and substrate are also consistent with an ordered bi bi mechanism. For 3α(17β)-hydroxysteroid dehydrogenase, the steady-state kinetic measurements and sub-strate binding studies suggest a random binding pattern of the substrates and an ordered release of product; NADH is released last. However, the two enzymes transferred the pro-R-hydrogen atom of NAD(P)H to the carbonyl substrate.