pH dependence of the kinetic parameters for 3-oxo-delta 5-steroid isomerase. Substrate catalysis and inhibition by (3S)-spiro[5 alpha-androstane-3,2'-oxiran]-17-one.
The pH-rate profiles for kcatobsd and (kcat/KM)obsd at 25.0 degrees C have been measured for 3-oxo-delta 5-steroid isomerase by using 5-androstene-3,17-dione (2), 5-pregnene-3,20-dione (3), and 5(10)-estrene-3,17-dione (4) as substrates. Results from the nonsticky substrate 4 suggest values for the pK of a catalytically important group on the free enzyme (pKE) of 4.57 and the pK of the same group in the enzyme-substrate complex of 4.74. For the sticky substrates 2 and 3, pKES is ca. 4.75 and 5.5, respectively. Analysis of the (kcat/KM)obsd vs. pH profile for 2 reveals that the intermediate E X S complex decomposes to products at a rate similar to its reversion to E + S. The pH-rate profile for inhibition of the isomerase by (3S)-spiro-[5 alpha-androstane-3,2'-oxiran]-17-one (7 beta) shows values for pKE of 4.75 and pKEI of 4.90. The similarity of the pH-rate profiles for isomerization of 4 and inhibition by 7 beta suggests that both reactions may be governed by the ionization state of the same carboxyl group of the enzyme.
R M
RM
Pollack
S
S
Bantia
P L
PL
Bounds
B M
BM
Koffman
D000731:Androstanes / Q000494:pharmacology*; D004355:Drug Stability; D006863:Hydrogen-Ion Concentration; D007535:Isomerases / Q000378:metabolism*; D007700:Kinetics; D008433:Mathematics; D011485:Protein Binding; D013251:Steroid Isomerases / Q000378:metabolism*
D000731:Androstanes; C049257:spiro(androstan-3,2'-oxiran)-17-one; D007535:Isomerases; D013251:Steroid Isomerases; C022660:steroid delta-isomerase