Characterization of steroid hormone ester hydrolyzing enzymes in liver microsomes

Abstract

To clarify the characteristics of steroid hormone ester hydrolyzing enzymes, optimum pH, species differences, tissue distribution, hepatic subcellular distribution, and the effects of several enzyme inhibitors were determined with steroid hemisuccinate and acetate esters as charged and uncharged substrates respectively. Kinetic experiments were also performed. The optimum pH for charged and uncharged esters, respectively, was pH 5.5 and 8.0. Diethyl- p-nitrophenylphosphate (E-600) and diisopropylfluorophosphate (DFP) (10 −5 M) completely inhibited esterase activities for all substrates. Bis- p-nitrophenylphosphate (BNPP) and phenylmethylsulfonyl fluoride (PMSF) (10 −5 M) inhibited 80 and 95 per cent, respectively of hemisuccinate esterase activities, but only 20 per cent or less of acetate esterase activities. HgCl 2 and p-chloromercuribenzoate (PCMB) (10 −4 M) inhibited approximately 80 per cent or more of steroid acetate esterase activities. The K m values for acetate and hemisuccinate esterases were almost the same (0.3 to 0.7 mM), but three different V max values were obtained. These results indicate the presence of two or more steroid hormone ester hydrolyzing enzymes.