Substrate and nucleotide specificity of placental microsomal 3β-hydroxysteroid dehydrogenase

Abstract

Recent kinetic studies on the placental microsomal 3β-hydroxysteroid dehydrogenase have shown that apparent Km values for 3β-hydroxy-5-androsten-17-one (dehydroepiandrosterone) and 3β-hydroxy-5-pregnen-20-one (pregnenolone) are 15nM and 40nM respectively, which are orders of magnitude lower than found in earlier studies. The purpose of this study was to investigate the substrate and nucleotide specificity of the 3β-hydroxysteroid dehydrogenase, and the ability of various steroids to inhibit the reaction at these lower steroid concentrations. Each steroid inhibited the metabolism of the other competitively, and the Ki values obtained were not significantly different from their respective Km values. The ability of various steroids to inhibit the reaction at concentrations of 100nM was usually less than that found at micromolar concentrations. However, certain steroids showed marked inhibition. For example, estrone and estradiol-17β inhibit the oxidation of both substrates competitively with Ki values of between 15 and 24nM. The Km values of dehydroepiandrosterone and pregnenolone with NADP+ as cofactor are higher than those with NAD+ as cofactor and the V values are much lower. These data indicate that in human placental microsomes a single 3β-hydroxysteroid dehydrogenase, essentially NAD+ specific, metabolizes dehydroepiandrosterone and pregnenolone.