RAT STEROID 5-ALPHA-REDUCTASE KINETIC CHARACTERISTICS - EXTREME PH DEPENDENCY OF THE TYPE-II ISOZYME IN PROSTATE AND EPIDIDYMIS HOMOGENATES

Reevaluating the assay for rat steroid 5 alpha-reductase isozymes in p rostate and epididymis homogenates we encountered an extreme pH-depend ency of the type II isozyme. The time-course of the metabolism of test osterone (T) to 17 alpha-hydroxy-5 alpha-androstan-3-one (DHT) at acid ic pH shows an initial burst when the homogenate is not brought to pH before the start of the incubation. Therefore, the rat type II 5 alpha -reductase isozyme does not follow Michaelian law under these conditio ns making a single time point measurement invalid. Assessing the pH-op timum of 5 alpha-reduction in both rat prostate and epididymis homogen ates we found a strong substrate- dependency: at high substrate concen trations a pH-optimum for the type II isozyme of pH 5.0 was found, whe reas at lower concentrations pH 5.5 is optimal. Establishing V-max (ma ximum velocities) and K-m (affinity constants) for the 5 alpha-reducti on of T at pH 4.5-8.0, the efficiency optimum V-max/K-m appeared to be pH 5.5 in both prostate and epididymis homogenates. Specifically at a cidic pH these kinetic characteristics of the type II isozyme vary man y-fold. Discrepancies in literature concerning 5 alpha-reductase chara cteristics can, at least in part, be attributed to the choice of optim al pH, or to pH shifts during the assay.