Traish Abdulmaged M, Huang Yue Hua, Min Kweonsik, Kim Noel N, Munarriz Ricardo, Goldstein Irwin
Department of Biochemistry, Boston University School of Medicine, 700 Albany Street, Room W607, Boston, MA 02118, USA.
Steroids. 2004 Jan;69(1):71-8. doi: 10.1016/j.steroids.2003.10.006.
In this study, we investigated the binding characteristics of [3H]Delta(5)-androstene-3beta,17beta-diol to rabbit vaginal cytosolic and nuclear extracts and in freshly excised intact tissue strips. [3H]delta(5)-Androstene-3beta,17beta-diol bound to a protein(s) in the vaginal nuclear extract with high affinity (K(d)=3-5 nM) and limited capacity (50-100 fmol/mg protein). No specific binding was detected in the cytoplasmic extracts. Competitive binding studies showed that binding of [3H]delta(5)-androstene-3beta,17beta-diol was effectively displaced with unlabeled delta(5)-androstene-3beta,17beta-diol but not with dehydroepiandrosterone, testosterone, dihydrotestosterone, triamcinolone acetonide, or progesterone. However, estradiol at high concentrations partially displaced bound [3H]delta(5)-androstene-3beta,17beta-diol. Incubation of freshly excised vaginal tissue strips with [3H]delta(5)-androstene-3beta,17beta-diol in the absence or presence of excess unlabeled delta(5)-androstene-3beta,17beta-diol for 1h at 37 degrees C resulted in specific binding to a soluble macromolecule in the nuclear KCl extracts. In addition, quantitative measurement of estrogen receptor, androgen receptor and delta(5)-androstene-3beta,17beta-diol binding protein was performed by equilibrium ligand binding assays using extracts of distal vaginal tissue from intact animals or ovariectomized animals treated for 2 weeks with vehicle, estradiol, testosterone, or estradiol plus testosterone. These changes in steroid hormone levels resulted in opposing trends between the estrogen receptor and delta(5)-androstene-3beta,17beta-diol binding protein, suggesting that delta(5)-androstene-3beta,17beta-diol binding protein is regulated differently by the hormonal milieu than the estrogen receptor. These data suggest that rabbit vaginal tissue expresses a novel binding protein which specifically binds delta(5)-androstene-3beta,17beta-diol and is distinct from the androgen and estrogen receptors.
在本研究中，我们研究了[3H]Δ⁵-雄烯-3β,17β-二醇与兔阴道胞质和核提取物以及新鲜切除的完整组织条带的结合特性。[3H]Δ⁵-雄烯-3β,17β-二醇以高亲和力（K(d)=3 - 5 nM）和有限容量（50 - 100 fmol/mg蛋白质）与阴道核提取物中的一种蛋白质结合。在胞质提取物中未检测到特异性结合。竞争性结合研究表明，[3H]Δ⁵-雄烯-3β,17β-二醇的结合可被未标记的Δ⁵-雄烯-3β,17β-二醇有效取代，但不能被脱氢表雄酮、睾酮、二氢睾酮、曲安奈德或孕酮取代。然而，高浓度的雌二醇可部分取代结合的[3H]Δ⁵-雄烯-3β,17β-二醇。在37℃下，将新鲜切除的阴道组织条带与[3H]Δ⁵-雄烯-3β,17β-二醇在不存在或存在过量未标记的Δ⁵-雄烯-3β,17β-二醇的情况下孵育1小时，导致其与核KCl提取物中的一种可溶性大分子发生特异性结合。此外，使用来自完整动物或用载体、雌二醇、睾酮或雌二醇加睾酮处理2周的去卵巢动物的远端阴道组织提取物，通过平衡配体结合测定法对雌激素受体、雄激素受体和Δ⁵-雄烯-3β,17β-二醇结合蛋白进行了定量测量。这些类固醇激素水平的变化导致雌激素受体和Δ⁵-雄烯-3β,17β-二醇结合蛋白呈现相反的趋势，表明Δ⁵-雄烯-3β,17β-二醇结合蛋白受激素环境的调节方式与雌激素受体不同。这些数据表明，兔阴道组织表达一种新型结合蛋白，该蛋白特异性结合Δ⁵-雄烯-3β,17β-二醇，且与雄激素和雌激素受体不同。
