COMPETITIVE PRODUCT INHIBITION OF AROMATASE BY NATURAL ESTROGENS

In order to better understand the function of aromatase, we carried out kinetic analyses to assess the ability of natural estrogens, estrone (E1), estradiol (E2), 16alpha-OHE1, and estriol (E3), to inhibit aromatization. Human placental microsomes (50 mug protein) were incubated for 5 min at 37-degrees-C with [1beta-H-3]testosterone (1.24 x 10(3) dpm H-3/ng, 35-150 nM) or [1beta-H-3,4-C-14]androstenedione (3.05 x 10(3) dpm H-3/ng, H-3/C-14 = 19.3, 7-65 nM) as substrate in the presence of NADPH, with and without natural estrogens as putative inhibitors. Aromatase activity was assessed by tritium released to water from the 1beta-position of the substrates. Natural estrogens showed competitive product inhibition against androgen aromatization. The K(i) of E1, E2, 16alpha-OHE1, and E3 for testosterone aromatization was 1.5, 2.2, 95, and 162 muM, respectively, where the K(m) of aromatase was 61.8 +/- 2.0 nM (n = 5) for testosterone. The K(i) of E1, E2, 16alpha-OHE1, and E3 for androstenedione aromatization was 10.6, 5.5, 252, and 1182 muM, respectively, where the K(m) of aromatase was 35.4 +/- 4.1 nM (n = 4) for androstenedione. These results show that estrogens inhibit the process of androgen aromatization and indicate that natural estrogens regulate their own synthesis by the product inhibition mechanism in vivo. Since natural estrogens bind to the active site of human placental aromatase P-450 complex as competitive inhibitors, natural estrogens might be further metabolized by aromatase. This suggests that human placental estrogen 2-hydroxylase activity is catalyzed by the active site of aromatase cytochrome P-450 and also agrees with the fact that the level of catecholestrogens in maternal plasma increases during pregnancy. The relative affinities and concentration of androgens and estrogens would control estrogen and catecholestrogen biosynthesis by aromatase.