Estrophilic 3α,3β, 17β,20α-hydroxysteroid dehydrogenase from rabbit liver—I. Isolation and purification

A procedure for isolation of a highly-purified estrophilic hydroxysteroid dehydrogenase (EHSD) from rabbit liver, including ammonium sulphate fractionation, gel filtration, ion-exchange and affinity chromatography on estradiol-Sepharose, has been developed. The enzyme possesses NADP-dependent 3α,3β,17β,20α-HSD activities with a wide spectrum of androgenic, progestagenic, and estrogenic substrates. EHSD is a monomeric protein whose molecular mass determined by different methods is 35,000–39,000. The protein exhibits microheterogeneity due to the differences in molecular surface charge. The catalytic and hormone-binding properties and molecular sizes of the two protein fractions obtained by chromatography on DEAE-Toyopearl are close or identical. The enzymatic activity of EHSD is minor as compared to other HSDs from rabbit liver. However, the low values of K_m, the high affinity for steroid ligands, and high tissue levels of EHSD suggest the protein to play a role in the biodynamics of sex hormones.