3b-Hydroxysteroid Dehydrogenase/D-Isomerase Activity Associated with the Human 17b-Hydroxysteroid Dehydrogenase Type 2 Isoform*

The type 2 isoform of human 17b-hydroxysteroid dehydrogenase (17bHSD2) efficiently catalyzes the oxidative metabolism of androgens and estrogens, and it is expressed in a large series of human peripheral tissues. To obtain a better understanding of the regulation of local steroid biosynthesis and metabolism in human tissues, we have established a dual steroidogenic activity of the 17bHSD2 enzyme after transfection of human 17bHSD2-transfected human embryonic kidney (293) cells. After transient transfection, the metabolism of testosterone, pregnenolone, and dehydroepiandrosterone (DHEA) in intact transfected 293 cells was evaluated by TLC-based radiometric assays. 17bHSD2-transfected cells converted 91% of testosterone (1 mmol/L) into androstenedione in a 2-h incubation period. In addition, pregnenolone (1 mmol/L) was converted to progesterone (18.5%), whereas DHEA (1 mmol/L) was metabolized to androstenedione (8.3% conversion) in a 15-h incubation period. The kinetics of the 3bhydroxysteroid dehydrogenase (3bHSD) and 17bHSD2 activities using cell homogenate protein of stably transfected 293 cells indicated that the catalytic efficiency (apparent catalytic efficiency 5 maximum velocity/Km) of this 3bHSD activity is approximately 2000-fold (pregnenolone as substrate) or 3000-fold (DHEA as substrate) weaker than that of 17bHSD2 activity. It is noteworthy, however, that the apparent catalytic efficiency of the HSD3B2 gene product is only approximately 50-fold higher than that of the 3bHSD aspect of the 17bHSD2 gene product. Pregnenolone or DHEA effectively inhibited 17bHSD2 activity in a noncompetitive fashion. Furthermore, the potent 5areductase/3bHSD inhibitor, 17b-N,N-diethylcarbamoyl-4-methyl-4aza-5a-androstane-3-one, inhibited neither 3bHSD nor 17bHSD2 activities. We conclude that human 17bHSD2 enzyme exhibits 3bHSD activity. Notwithstanding that this 3bHSD activity is reduced compared to 17bHSD oxidative activity, it may account for at least some of the reports of 3bHSD activity found in human peripheral tissues that express notable amounts of the 17bHSD2 isozyme as well as in individuals with severe classic 3bHSD deficiency. (J Clin Endocrinol Metab 85: 3669–3672, 2000) 1 dehydrogenase (17bHSD) enzymes, which catalyze the reversible reactions between estrone and estradiol and between androstenedione and testosterone, are believed to play essential roles in regulation of the tissue levels of bioactive steroids. In recent years various isoforms of 17bHSD have been identified (1–4), and it is revealed that 17b-reduction and oxidation of androgens and estrogens are catalyzed by different isozymes. 17bHSD type 1 catalyzes the 17b-reduction of estrogens (1), type 3 is the most efficient in the reduction of testosterone to androstenedione (3), whereas 17bHSD type 2 (17bHSD2) is NAD dependent and efficiently catalyzes the oxidative metabolism of estrogens and androgens (2). 17bHSD2 enzyme is expressed in a large series of human peripheral tissues, including the placenta, liver, pancreas, kidney, small intestine, and secretory endometrium (5), and it is believed to be a key enzyme in the transformation of bioactive steroids to inactive