17β-Hydroxysteroid Dehydrogenase from Human Red Blood Cells

Abstract 17β-Hydroxysteroid dehydrogenase has been isolated from human red cells and purified 740 times. The enzyme is sensitive to ionic strength and is most effective at sodium chloride concentrations of 0.1 m and above. 17β-Estradiol 3-sulfate reacts at a faster rate than 17β-estradiol. The Km for estradiol is 9.1 x 10-5, and for estradiol sulfate it is 3.8 x 10-4. The free compound has a pH optimum of 9.7; the sulfate reacts optimally at pH 9.0. The enzyme is specific for TPN; DPN shows one-sixth to one-tenth as much activity. The Km values for TPN are 2.7 x 10-5 with estradiol and 2.4 x 10-5 with estradiol sulfate. Acetyl-TPN is less active than TPN. Albumin inhibits the reaction, presumably by binding the steroids. Organic solvents also act inhibitorily. The enzyme appears to be of the sulfhydryl type. Enzyme activity was assayed by measurement of TPNH produced and by yield of estrone. Reduction of estrone or its sulfate occurs at slower rates than the corresponding reverse reactions. The molecular weight of the enzyme, calculated from mobility data on Sephadex gel, is approximately 75,500.