Boada L D, Fernández L, Zumbado M, Luzardo O P, Chirino R, Díaz-Chico B N
Departamento de Ciencias Clínicas, Universidad de Las Palmas de Gran Canaria, Spain.
J Pharmacol Exp Ther. 1996 Dec;279(3):1123-9.
Male rat liver microsomes contain a [3H]dexamethasone binding site, capable of binding glucocorticoids and progesterone. We have shown previously that the 17 alpha-alkylated androgen, stanozolol, can inhibit the [3H]dexamethasone binding to microsomes through a negative allosteric mechanism, which gives rise to the possibility of its interaction with a different binding site. In this study, the existence of a single-saturating binding site, capable of binding the radioactive steroid with a maximum number of the specific binding site of 49 +/- 2 pmol/mg of protein and a Kd of 37 +/- 1.3 nM was demonstrated by using [3H]stanozolol. In competition experiments, only stanozolol and danazol were able to compete with [3H]stanozolol for its binding to microsomes, among more than 60 steroids and other compounds tested. The binding of [3H]stanozolol was depressed after protease treatment of the microsomes, or after the administration of cycloheximide to adult male rats for 24 hr, which suggest its proteic nature. The [3H]stanozolol binding site was detected in many tissues of the rat, with the highest concentrations being found in the liver. It was detected from birth, increasing afterward in concentration and reaching a peak at 2 to 3 months of age. This is the first experimental verification of the existence in liver microsomes of a specific binding site for some 17 alpha-alkylate androgens, such as stanozolol and danazol, different from the androgen receptor or the [3H]dexamethasone binding site.
雄性大鼠肝脏微粒体含有一个[3H]地塞米松结合位点，能够结合糖皮质激素和孕酮。我们之前已经表明，17α-烷基化雄激素司坦唑醇可通过负别构机制抑制[3H]地塞米松与微粒体的结合，这增加了其与不同结合位点相互作用的可能性。在本研究中，通过使用[3H]司坦唑醇证明了存在一个单一饱和结合位点，该位点能够结合放射性类固醇，其特异性结合位点的最大数量为49±2 pmol/mg蛋白质，解离常数（Kd）为37±1.3 nM。在竞争实验中，在测试的60多种类固醇和其他化合物中，只有司坦唑醇和达那唑能够与[3H]司坦唑醇竞争其与微粒体的结合。用蛋白酶处理微粒体后，或给成年雄性大鼠注射环己酰亚胺24小时后，[3H]司坦唑醇的结合受到抑制，这表明其具有蛋白质性质。在大鼠的许多组织中都检测到了[3H]司坦唑醇结合位点，其中肝脏中的浓度最高。该位点在出生时即可检测到，随后浓度增加，并在2至3个月龄时达到峰值。这是首次通过实验证实肝脏微粒体中存在一些17α-烷基化雄激素（如司坦唑醇和达那唑）的特异性结合位点，该位点不同于雄激素受体或[3H]地塞米松结合位点。
