Multiple Steps Determine the Overall Rate of the Reduction of 5α-Dihydrotestosterone Catalyzed by Human Type 3 3α-Hydroxysteroid Dehydrogenase: Implications for the Elimination of Androgens

Human type 3 3α-hydroxysteroid dehydrogenase, or aldo-keto reductase (AKR) 1C2, eliminates the androgen signal in human prostate by reducing 5α-dihydrotestosterone (DHT, potent androgen) to form 3α-androstanediol (inactive androgen), thereby depriving the androgen receptor of its ligand. The kcat for the NADPH-dependent reduction of DHT catalyzed by AKR1C2 is 0.033 s-1. We employed transient kinetics and kinetic isotope effects to dissect the contribution of discrete steps to this low kcat value. Stopped-flow experiments to measure the formation of the AKR1C2·NADP(H) binary complex indicated that two slow isomerization events occur to yield a tight complex. A small primary deuterium isotope effect on kcat (1.5) and a slightly larger effect on kcat/Km (2.1) were observed in the steady state. In the transient state, the maximum rate constant for the single turnover of DHT (ktrans) was determined to be 0.11 s-1 for the NADPH-dependent reaction, which was ∼4-fold greater than the corresponding kcat. ktrans wa...