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 prostate and epididymis homogenates we encountered an extreme pH-dependency of the type II isozyme. The time-course of the metabolism of testosterone (T) to 17 beta-hydroxy-5 alpha-androstan-3-one (DHT) at acidic 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 conditions making a single time point measurement invalid. Assessing the pH-optimum of 5 alpha-reduction in both rat prostate and epididymis homogenates we found a strong substrate dependency: at high substrate concentrations a pH-optimum for the type II isozyme of pH 5.0 was found, whereas at lower concentrations pH 5.5 is optimal. Establishing Vmax (maximum velocities) and Km (affinity constants) for the 5 alpha-reduction of T at pH 4.5-8.0, the efficiency optimum Vmax/Km appeared to be pH 5.5 in both prostate and epididymis homogenates. Specifically at acidic pH these kinetic characteristics of the type II isozyme vary many-fold. Discrepancies in literature concerning 5 alpha-reductase characteristics can, at least in part, be attributed to the choice of optimal pH, or to pH shifts during the assay.
Span
P N
PN
Department of Medicine, University of Hospital, Nijmegen, The Netherlands.
Smals
A G
AG
Sweep
C G
CG
Benraad
T J
TJ
D000818:Animals; D042944:Cholestenone 5 alpha-Reductase; D004822:Epididymis / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D010088:Oxidoreductases / Q000378:metabolism*; D011467:Prostate / Q000201:enzymology*; D051381:Rats; D017208:Rats, Wistar
D007527:Isoenzymes; D010088:Oxidoreductases; D042944:Cholestenone 5 alpha-Reductase