Dighe R R, Murthy G S, Kurkalli B S, Moudgal N R
Department of Biochemistry, Indian Institute of Science, Bangalore.
Mol Cell Endocrinol. 1990 Jul 30;72(1):63-70. doi: 10.1016/0303-7207(90)90240-9.
The conformation of the common alpha-subunit of human glycoprotein hormones, luteinizing hormone (hLH), follicle-stimulating hormone (hFSH), thyroid-stimulating hormone (hTSH) and chorionic gonadotropin (hCG) was probed using a highly specific polyclonal antiserum against the alpha-subunit of hCG and several monoclonal antibodies (MAbs) produced against hCG which recognized the alpha-subunit in free and combined form. The alpha-subunit was found to be conformationally altered (compared to its conformation in the isolated state) when it was in combination with various beta-subunits as indicated by shifts in the displacement curves of binding of [125I]hCG alpha to the polyclonal antiserum. The extent of the change was dependent on the beta-subunit present with minimum change being observed with hLH beta, intermediate with hCG beta and maximum change with hFSH and TSH beta-subunits. However, the affinity constants of this antiserum for all four hormones were nearly similar. Further, it was also found that binding of any one of the glycoprotein hormones to this antibody could be completely inhibited by any other hormone suggesting that the conformation of the alpha-subunit in all the four hormones is probably very similar. This was further investigated using five hCG MAbs capable of recognizing the alpha-subunit, but with different epitope specificities. All these MAbs could recognize all the four hormones suggesting the presence of the epitopes in these proteins. These epitopes were conformation specific since the MAbs did not bind reduced and carboxymethylated alpha-subunit. Displacement analysis using [125I]hCG as the tracer showed that two epitopes have nearly the same conformation in all the four hormones, while two were partially modified depending on the beta-subunit present. Based on these results, it is concluded that the alpha-subunit of glycoprotein hormones has nearly the same conformation, though subtle differences do exist.
使用针对人绒毛膜促性腺激素（hCG）α亚基的高度特异性多克隆抗血清以及几种针对hCG产生的单克隆抗体（MAb），对人糖蛋白激素促黄体生成素（hLH）、促卵泡激素（hFSH）、促甲状腺激素（hTSH）和绒毛膜促性腺激素（hCG）的共同α亚基的构象进行了探究，这些单克隆抗体可识别游离和结合形式的α亚基。当α亚基与各种β亚基结合时，发现其构象发生了改变（与分离状态下的构象相比），这通过[125I]hCGα与多克隆抗血清结合的置换曲线的位移表明。变化程度取决于存在的β亚基，与hLHβ结合时变化最小，与hCGβ结合时变化中等，与hFSH和TSHβ亚基结合时变化最大。然而，该抗血清对所有四种激素的亲和常数几乎相似。此外，还发现任何一种糖蛋白激素与该抗体的结合都可被任何其他激素完全抑制，这表明所有四种激素中α亚基的构象可能非常相似。使用五种能够识别α亚基但具有不同表位特异性的hCG单克隆抗体对此进行了进一步研究。所有这些单克隆抗体都能识别所有四种激素，表明这些蛋白质中存在表位。这些表位具有构象特异性，因为单克隆抗体不与还原和羧甲基化的α亚基结合。使用[125I]hCG作为示踪剂的置换分析表明，所有四种激素中有两个表位具有几乎相同的构象，而另外两个表位则根据存在的β亚基而部分修饰。基于这些结果，可以得出结论，糖蛋白激素的α亚基具有几乎相同的构象，尽管确实存在细微差异。
