Inactivation of human placental aromatase by 6 alpha- and 6 beta-hydroperoxyandrostenedione.

The epimeric 6 alpha- and 6 beta-hydroperoxy derivatives of androstendione caused irreversible inactivation of human placental aromatase. Microsomes from term-placentae were first preincubated in the presence of increasing concentrations of the hydroperoxides. The microsomes were then washed free of steroids and its residual aromatase activity was assayed by the tritium-exchange method to [3H]water. Aromatase activity decreased in a time-, and concentration-dependent manner; the axial, beta-hydroperoxy epimer was the slightly stronger inactivator. Less inactivation occurred when during the preincubation stage the natural aromatase substrate, androstenedione, or the anti-oxidant, dithiothreitol, was added. The sulfhydryl reagent, p-hydroxy-mercuribenzoate, decreased this protective effect. The inactivation is not dependent on the presence of NADPH. Both steroids induced a Type I difference spectrum with a Ks of 0.167 microM and 0.163 microM for the 6 alpha-, and the 6 beta-hydroperoxyandrostenedione, respectively. We suggest that these 6-hydroperoxyandrogens may function as active-site directed inhibitors and inactivators of estrogen synthetase through oxidation of cysteine residues.