Ligand-induced Rearrangements of the GABAB Receptor Revealed by Fluorescence Resonance Energy Transfer
Ligand-induced Rearrangements of the GABAB Receptor Revealed by Fluorescence Resonance Energy Transfer
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DOI：
10.1074/jbc.m109.077990
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发表时间：
2010-04-02
影响因子：
4.8
通讯作者：
Tateyama, Michihiro
中科院分区：
文献类型：
--
作者：
Matsushita, Shinichi;Nakata, Hiroyasu;Tateyama, Michihiro
γ-氨基丁酸B型受体（GABA（B）R）是C家族G蛋白偶联受体成员之一，以由亚基GB 1和GB 2组成的异源二聚体形式存在。为了阐明GABA（B）R的配体诱导的活化机制，每个亚基在其细胞内环处与Cerulean或增强型黄色荧光蛋白融合，并监测激动剂应用后的荧光共振能量转移（FRET）变化。结果，在GB 1a环2和GB 2环2之间以及GB 1a环2和GB 2环1之间观察到FRET减少，表明在受体活化期间细胞内结构域解离。预期两个亚基间FRET对都忠实地捕获原始受体的激活，因为它们的药理学性质与野生型受体的药理学性质高度相似。然而，intrasubunit数据表明，受体激活不涉及每个亚基的跨膜结构域内的主要结构变化。结合两个不同水平的结果，可以得出结论，激动剂激活GABA（B）R与膜上的GB 1 a和GB 2的不对称亚基间重排有关。这种类型的激活模式，亚基间重排没有明显的螺旋内结构的变化，似乎共同的GABA（B）R和代谢型谷氨酸受体1 α，另一个家庭C G蛋白偶联受体先前由我们的小组研究。然而，细胞内结构域的运动方向和它的不对称性，在这里观察到突出了两个受体之间的质的差异。
The gamma-aminobutyric acid type B receptor (GABA(B)R), one of the family C G-protein-coupled receptor members, exists as a heterodimer comprised of subunits GB1 and GB2. To clarify the ligand-induced activation mechanism of the GABA(B)R, each subunit was fused with either Cerulean or enhanced yellow fluorescent protein at its intracellular loop, and fluorescence resonance energy transfer (FRET) changes upon agonist application were monitored. As a result, FRET decreases were observed between GB1a loop 2 and GB2 loop 2 and between GB1a loop 2 and GB2 loop 1, suggesting the dissociation of intracellular domains during the receptor activation. Both intersubunit FRET pairs were expected to faithfully capture the activation of the original receptor as their pharmacological properties were highly similar to that of the wild-type receptor. However, the intrasubunit data suggest that the receptor activation does not involve major structural changes within the transmembrane domain of each subunit. By combining the results obtained from two different levels, it was concluded that the GABA(B)R activation by agonist is associated with an asymmetrical intersubunit rearrangement of GB1a and GB2 on the membrane. This type of activation mode, an intersubunit rearrangement without apparent intrahelical structural changes, appears commonly shared by the GABA(B)R and the metabotropic glutamate receptor 1 alpha, another family C G-protein-coupled receptor previously studied by our group. Nevertheless, the directions of intracellular domain movements and its asymmetry observed here highlight the qualitative difference between the two receptors.