Studies on the human testis. VI. NADH-linked reactions of microsomal steroid 20alpha-and 20beta-hydroxysteroid dehydrogenase and 17alpha-hydroxylase.
NADH-linked 20alpha- and 20beta-hydroxysteroid dehydrogenase and 17alpha-hydroxylase activities were demonstrated in the microsomal fraction of the human testis. The microsomal 20alpha-hydroxysteroid dehydrogenase showed substrate affinity to pregnenolone and progesterone and not to 17alpha-hydroxyprogesterone and preferred NADH to NADPH as a hydrogen donor. In the presence of NADH, the optimal pH for the enzyme was 7.7 and the apparent Michaelis constants of the enzyme for progesterone and pregnenolone at 37 C and pH 7.4 were 6.9-7.1 X 10-6M and in the order of 10-5M, respectively, 17alpha, 20beta-Dihydroxypregn-4-en-3-one was the only significant metabolite produced from 17alpha-hydroxyprogesterone by microsomal fraction of the human testis in the presence of NADH. The apparent Michaelis constant of microsomal 20beta-hydroxysteroid dehydrogenase for 17alpha-hydroxyprogesterone in the presence of NADH was in the order of 10-5M at 37 C and pH 7.4. The microsomal 17alpha-hydroxylase catalyzed the metabolism of pregnenolone and progesterone at a similar rate in the presence of NADH. The optimal pH and the apparent Michaelis constant at 37 C and pH 7.4 of the NADH-linked reaction of 17alpha-hydroxylase for progesterone were 7.7 and 5.3-5.4 X 10-7M, resepctively. The NADH-linked enzyme activity for progesterone was competitively inhibited by both pregn-5-ene-3beta, 20alpha-diol (inhibition constant: 1.7 X 10-7M) and 20alpha-hydroxypregn-4-en-3 one (inhibition constant: 6.6 X 10-7M), and was resistant to poor oxygen supply during incubation. The results indicate that the microsomal 20alpha-hydroxysteroid dehydrogenase is a different enzyme from the one in the soluble fraction of the human testis and that microsomal 17alpha-hydroxylase in the human testis is activated by NADH as well as NADPH.
D001667:Binding, Competitive; D006863:Hydrogen-Ion Concentration; D006908:Hydroxyprogesterones; D006913:Hydroxysteroid Dehydrogenases / Q000378:metabolism*; D006914:Hydroxysteroids; D008297:Male; D008861:Microsomes / Q000201:enzymology*; D006899:Mixed Function Oxygenases / Q000378:metabolism*; D009243:NAD / Q000378:metabolism*; D009249:NADP / Q000378:metabolism; D010100:Oxygen; D011283:Pregnenes / Q000378:metabolism; D011284:Pregnenolone; D011374:Progesterone / Q000031:analogs & derivatives / Q000378:metabolism; D013696:Temperature; D013737:Testis / Q000201:enzymology*
D006908:Hydroxyprogesterones; D006914:Hydroxysteroids; D011283:Pregnenes; D009243:NAD; D011374:Progesterone; D009249:NADP; D011284:Pregnenolone; D006899:Mixed Function Oxygenases; D006913:Hydroxysteroid Dehydrogenases; D010100:Oxygen