多巴胺D_2/血管紧张素AT_1嵌合受体:D_2受体第三细胞内环影响受体与配基结合以及与G蛋白偶联的特性(英文)

目的:研究受体第三细胞内环(IL_3)的长度对受体与配基结合及与G蛋白偶联特性的影响.方法:用目前已知的G蛋白偶联受体中IL_3最短的血管紧张素Ⅱ AT_1受体的IL_3替换野生型D_2受体较长的IL_3,组成D_2/AT,嵌合受体.结果:与野生型D_2受体相比,D_2/AT_1嵌合受体与拮抗剂的亲和性均降低,与激动剂的亲和性有的增高,有的降低.嵌合受体失去与G蛋白偶联的能力,也不能产生磷酸肌醇水解.结论:受体的IL_3对受体配基结合位点和空间构象有一定影响;受体与G蛋白的偶联不仅与IL_3有关,而且还受非IL_3区域的影响,而IL_3的长度是决定这两方面影响的因素之一.

Chimeric dopamine D_2/ angiotensin AT_1 receptors: role of the length of third intracellular loop of D_2 receptors in conferring specificity of receptor binding and G-protein coupling

AIM: To define roles of the third intracellular loop (IL_3) length of G-protein coupled receptors in conferring the specificity for receptor binding and G-protein coupling. METHODS: By polymerase chain reaction (PCR), the IL_3 of D_2 receptor was replaced with the counter part of AT_1 receptor which has the shortest loop among all G-protein coupled receptors. D_2/AT_1 receptor cDNA was then stably transfected into Chinese hamster ovary cells and a clone with high level expression was obtained for receptor binding and agonist-induced phosphatidylinositols (PI) turnover experiments. RESULTS: Comparing to the D_2 receptor, D_2/AT_1 chimeric receptor had lower affinities for all D_2 receptor antagonists tested (spiperone, haloperidol, ( )-butaclamol, chlopromazine, clozapine, trifluo-perdazine) and different affinity profiles to agonists ( apomorphine, dopamine, quinpirole, bromo-criptine). But the chimeric receptor failed to couple to G-protein and subsequent stimulation of PI turnover. CONCLUSION: The length of IL_3 of D_2 receptor participates defining receptor binding sites conformation, and structure beyond IL_3 may affect receptor G-protein coupling.