Studies of the human testis. XX--Effect of NADH on 17 alpha-hydroxypregnene C17-20 lyase activity.
A study of the cofactor requirements of C17-20 lyase was carried out using human testis tissue obtained at the time of orchiectomy for treatment of prostatic carcinoma. Washed microsomal fractions were prepared from frozen human testes using a KCl containing buffer. The preparation revealed dose-dependent activity of C17-20 lyase in the presence of either NADPH or commercial or purified NADH. The Km of NADH for the enzyme was of the order of 10(-3) M and the Km of NADPH was determined as 1.6 X 10(-5) M. NADH also provided synergistic enhancement of NADPH-mediated lyase activity, and decreased the Km of NADPH for the lyase but did not change the Vmax of NADPH-mediated lyase activity. Carbon monoxide inhibited both NADH and NADPH-mediated lyase activities indicating that both activities are catalyzed by cytochrome P-450. Cations including Ca2+, Mg2+ and Mn2+ were found to inhibit the NADPH-mediated lyase activity but enhanced the lyase activity in the presence of NADH. The results indicate both the presence of NADH-mediated C17-20 lyase activity and the synergistic effect of NADH on NADPH-mediated lyase activity in the human testis.
D000368:Aged; D000446:Aldehyde-Lyases / Q000032:analysis*; D002118:Calcium / Q000494:pharmacology; D002248:Carbon Monoxide / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008297:Male; D008875:Middle Aged; D009243:NAD / Q000494:pharmacology*; D013254:Steroid 17-alpha-Hydroxylase; D013737:Testis / Q000201:enzymology*
D009243:NAD; D002248:Carbon Monoxide; D013254:Steroid 17-alpha-Hydroxylase; D000446:Aldehyde-Lyases; D008274:Magnesium; D002118:Calcium