Department of Biophysics, The University of Texas Southwestern Medical Center, Dallas, United States.
Molecular Biophysics Graduate Program, The University of Texas Southwestern Medical Center, Dallas, United States.
Elife. 2017 Oct 6;6:e28505. doi: 10.7554/eLife.28505.
GPCRs regulate all aspects of human physiology, and biophysical studies have deepened our understanding of GPCR conformational regulation by different ligands. Yet there is no experimental evidence for how sidechain dynamics control allosteric transitions between GPCR conformations. To address this deficit, we generated samples of a wild-type GPCR (AR) that are deuterated apart from H/C NMR probes at isoleucine δ1 methyl groups, which facilitated H/C methyl TROSY NMR measurements with opposing ligands. Our data indicate that low [Na] is required to allow large agonist-induced structural changes in AR, and that patterns of sidechain dynamics substantially differ between agonist (NECA) and inverse agonist (ZM241385) bound receptors, with the inverse agonist suppressing fast ps-ns timescale motions at the G protein binding site. Our approach to GPCR NMR creates a framework for exploring how different regions of a receptor respond to different ligands or signaling proteins through modulation of fast ps-ns sidechain dynamics.
G 蛋白偶联受体（GPCRs）调节着人类生理学的各个方面，生物物理研究加深了我们对不同配体调节 GPCR 构象的理解。然而，对于侧链动力学如何控制 GPCR 构象之间的变构转变，目前还没有实验证据。为了解决这一不足，我们生成了野生型 G 蛋白偶联受体（AR）的样本，这些样本除了 H/C NMR 探针外，在异亮氨酸 δ1 甲基处都被氘取代，这使得可以用相反的配体进行 H/C 甲基 TROSY NMR 测量。我们的数据表明，低 [Na] 是允许 AR 中发生大的激动剂诱导结构变化所必需的，并且在激动剂（NECA）和反向激动剂（ZM241385）结合的受体之间，侧链动力学模式有很大的不同，反向激动剂抑制了 G 蛋白结合部位的快速 ps-ns 时间尺度运动。我们的 GPCR NMR 方法为探索受体的不同区域如何通过调节快速 ps-ns 侧链动力学来响应不同的配体或信号蛋白提供了一个框架。
Zotero 插件
