Affinity labeling of rat liver microsomal NADH-5 alpha-reductase with a nucleoside analogue

A time dependent irreversible loss of rat liver microsomal NADH-5 alpha-reductase activity is caused by incubation of microsomes with the nucleoside 5'-p-fluorosulfonylbenzoyladenosine (FSA). The decrease of activity is dependent on FSA concentration and shows first order kinetics. Presence of NADH partially stabilizes the NADH-5 alpha-reductase. Thioglycerol present before incubation prevents loss of activity, and stops decrease of activity when added during incubation. NADPH-5 alpha-reductase (E.C. 1.3.1.4) and NADPH-cytochrome c reductase (E.C. 1.6.2.4) are not influenced while NADH-cytochrome c reductase (E.C. 1.6.99.3) is inhibited by FSA. Evidently FSA causes inactivation of the enzymes by binding to the NADH-binding site. Affinity labeling by FSA thus clearly distinguishes between NADH- and NADPH-dependent 5 alpha-reductases from rat liver microsomes.