Yan Dong, White Michael M
Department of Pharmacology and Physiology, Drexel University College of Medicine, 245 N. 15th Street, Philadelphia, PA 19102, USA.
Mol Pharmacol. 2005 Aug;68(2):365-71. doi: 10.1124/mol.105.011957. Epub 2005 May 24.
The serotonin type 3 receptor (5-HT(3)R) is a member of the cys-loop ligand-gated ion channel (LGIC) superfamily. Like almost all membrane proteins, high-resolution structural data are unavailable for this class of receptors. We have taken advantage of the high degree of homology between LGICs and the acetylcholine binding protein (AChBP) from the freshwater snail Lymnea stagnalis, for which high-resolution structural data are available, to create a structural model for the extracellular (i.e., ligand-binding) domain of the 5-HT(3)R and to perform a series of ligand docking experiments to delineate the architecture of the ligand-binding site. Structural models were created using homology modeling with the AChBP as a template. Docking of the antagonist granisetron was carried out using a Lamarckian genetic algorithm to produce models of ligand-receptor complexes. Two energetically similar conformations of granisetron in the binding site were obtained from the docking simulations. In one model, the indazole ring of granisetron is near Trp90 and the tropane ring is near Arg92; in the other, the orientation is reversed. We used double-mutant cycle analysis to determine which of the two orientations is consistent with experimental data and found that the data are consistent with the model in which the indazole ring of granisetron interacts with Arg92 and the tropane ring interacts with Trp90. The combination of molecular modeling with double-mutant cycle analysis offers a powerful approach for the delineation of the architecture of the ligand-binding site.
5-羟色胺3型受体（5-HT(3)R）是半胱氨酸环配体门控离子通道（LGIC）超家族的成员。与几乎所有膜蛋白一样，这类受体尚无高分辨率结构数据。我们利用LGIC与淡水蜗牛椎实螺的乙酰胆碱结合蛋白（AChBP）之间的高度同源性（已有AChBP的高分辨率结构数据），构建了5-HT(3)R细胞外（即配体结合）结构域的结构模型，并进行了一系列配体对接实验以描绘配体结合位点的结构。以AChBP为模板，通过同源建模构建结构模型。使用拉马克遗传算法对拮抗剂格拉司琼进行对接，以生成配体-受体复合物模型。对接模拟得到了格拉司琼在结合位点的两种能量相似的构象。在一种模型中，格拉司琼的吲唑环靠近色氨酸90，托烷环靠近精氨酸92；在另一种模型中，方向相反。我们使用双突变循环分析来确定这两种方向中的哪一种与实验数据一致，结果发现数据与格拉司琼的吲唑环与精氨酸92相互作用、托烷环与色氨酸90相互作用的模型一致。分子建模与双突变循环分析相结合，为描绘配体结合位点的结构提供了一种强大的方法。
