Characterization of molecular and catalytic properties of intact and truncated human 17 beta-hydroxysteroid dehydrogenase type 2 enzymes: Intracellular localization of the wild-type enzyme in the endoplasmic reticulum

Human 17 beta-hydroxysteroid dehydrogenase (17HSD) type 2 is a widely distr ibuted enzyme that primarily converts the highly active 17 beta-hydroxyster oids to their inactive keto forms. In the present study, full-length human 17HSD type 2 was localized in the endoplasmic reticulum using a double immu nofluorescence labeling technique. As a consequence of its strong membrane interaction, full-length human 17HSD type 2 could not be solubilized as a b iologically active form in vitro. However, by deleting the first 29 amino a cids from the N-terminus, we were able to purify a catalytically active enz yme from the cytosolic fraction of Sf9 insect cells. Biochemical and cataly tic properties of the purified truncated human 17HSD type 2 protein confirm its suitability for structure-function analyses of the enzyme. Both intact and truncated 17HSD type 2 enzymes efficiently catalyzed the oxidation of estradiol, testosterone, dihydrotestosterone, androstenediol, and 20 alpha- dihydroprogesterone. The oxidation of estradiol brought about by human 17HS D type 2 was effectively inhibited by several other steroidal compounds, su ch as a-hydroxyestradiol, 5 beta-androstan-3 alpha,17 beta-diol, 5 alpha-an drostan-3 alpha,17 beta-diol, and 5 alpha-androstan-3 beta,17 beta-diol. Th e broad substrate specificity of human 17HSD type 2 together with its predo minant oxidative activity and intracellular location, as observed in this s tudy, indicate the physiological role of the enzyme to be primarily an inac tivator of highly active 17 beta-hydroxysteroids.