Residues in the seventh membrane-spanning segment of the dopamine D2 receptor accessible in the binding-site crevice
Residues in the seventh membrane-spanning segment of the dopamine D2 receptor accessible in the binding-site crevice
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DOI：
10.1021/bi960928x
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发表时间：
1996-09-03
期刊：
影响因子：
2.9
通讯作者：
Javitch, JA
中科院分区：
文献类型：
--
作者：
Fu, DY;Ballesteros, JA;Javitch, JA
多巴胺D2受体的结合位点，像其他同源G蛋白偶联受体一样，包含在其七个跨膜片段之间形成的水可进入的缝隙中。使用取代的半胱氨酸可及性方法，我们以前映射的残基，形成的结合位点裂缝的表面在第三和第五跨膜段（M3和M5）。我们现在已经突变为半胱氨酸，一次一个，第七跨膜片段（M7）中和侧翼的26个连续残基，并在HEK 293细胞中表达突变体受体。这些突变体中的9个与带电的、亲水的、疏脂的、巯基特异性的试剂反应，加入细胞外，并通过可逆的多巴胺拮抗剂舒必利保护反应。因此，我们推断这些残基的侧链在结合位点裂缝的水可接近表面中。半胱氨酸取代突变体的可及性模式与M7是扭结的α-螺旋一致。
The binding site of the dopamine D2 receptor, like that of other homologous G-protein-coupled receptors, is contained within a water-accessible crevice formed among its seven membrane-spanning segments. Using the substituted-cysteine accessibility method, we previously mapped the residues that form the surface of the binding-site crevice in the third and fifth membrane-spanning segments (M3 and M5). We have now mutated to cysteine, one at a time, 26 consecutive residues in and flanking the seventh membrane-spanning segment (M7) and expressed the mutant receptors in HEK 293 cells. Nine of these mutants reacted with charged, hydrophilic, lipophobic, sulfhydryl-specific reagents, added extracellularly, and were protected from reaction by a reversible dopamine antagonist, sulpiride. Thus, we infer that the side chains of these residues are in the water-accessible surface of the binding-site crevice. The pattern of accessibility of the cysteine-substitution mutants is consistent with M7 being a kinked alpha-helix.