Structural dynamics underlying GPCR-G protein selectivity

摘要： 标题：GPCR-G蛋白选择性背后的结构动力学 在与不同效果的激动剂结合后，G蛋白偶联受体(GPCRs)介导 通过与不同亚型的G蛋白偶联，形成多条信号通路。某些激动剂表现出 它们对特定G蛋白信号通路的有效性的选择性。这样的配体提供了精确的 与当今GPCR靶向药物相比，治疗效果更好，副作用更少。那里 很少有选择性配体是已知的特定G蛋白信号通路，因为设计这样的 激动剂是一个令人望而生畏的实验性挑战。这是由于严重缺乏对如何 GPCRs与不同类型的G蛋白结合时调节其同源G蛋白的功能选择性 激动剂和细胞内。在过去的三年里，利用多尺度分子动力学的组合 (MD)方法和遗传编码的FRET传感器我们已经证明GPCR具有潜在的细胞内 可以重塑和连接不同G蛋白的空洞。通过重组G蛋白和GPCRs 我们已经确定了GPCR-G-蛋白界面上的热点残基，这些残基充当了 G蛋白通过GPCRs选择性偶联。尽管很明显，变构通讯来自于 G蛋白偶联位点的配体结合部位在G蛋白选择性中的作用目前知之甚少 关于这一沟通的机制和这一过程中涉及的残留物。在这里我们 建议(1)结合FRET使用本实验室开发的计算方法Alallsteer 传感器、NanoBRET分析和基于细胞的下游分析来描绘变构网络 GPCR残基参与调节9个不同Gs、Gi和Gq偶联A类的G蛋白选择性 在蜂窝条件下的GPCRs。第二个目标是了解部分激动剂在脑内的作用 与全激动剂相比，GPCR-G蛋白选择性偶联。我们会找出热点 与部分激动剂结合时G蛋白和GPCRs上的残基并比较其相似性和 与完全激动剂的区别。拟议工作的结果将提供变构的作用 G蛋白选择性中的残基网络和部分激动剂如何调节受体 构象。这将推动我们对GPCRs如何在生物学过程中的理解 在活细胞中识别它们的同源G蛋白。

Summary: Title: Structural dynamics underlying GPCR-G protein selectivity Upon binding to agonists with different efficacies, G protein-coupled receptors (GPCRs) mediate multiple signaling pathways by coupling to different subtypes of G proteins. Certain agonists exhibit selectivity in their efficacy to specific G-protein signaling pathways. Such ligands provide precise therapeutic benefits with fewer side effects as drugs compared to today's GPCR-targeted drugs. There are very few selective ligands known for specific G-protein signaling pathways because designing such agonists is a daunting experimental challenge. This is due to serious lack of understanding of how GPCRs modulate their functional selectivity for their cognate G-protein when bound to different type of agonists and in cells. In the past three years using a combination of multiscale molecular dynamics (MD) method and genetically coded FRET sensors we have shown that GPCRs have latent intracellular cavities that can reshape and couple to different G-proteins. By reengineering G-proteins and GPCRs we have identified the hotspot residues in the GPCR-G-protein interface that serve as the “QR code” for G-protein selective coupling by GPCRs. Although it is clear that allosteric communication from the ligand binding site to the G-protein coupling site plays a role in G-protein selectivity very little is known about the mechanism of this communication and the residues involved in this process. Here we propose to (1) use the computational method Allosteer developed in our lab in combination with FRET sensors, NanoBRET assays and cell based downstream assays to delineate the allosteric network of GPCR residues involved in regulating G-protein selectivity in 9 different Gs, Gi and Gq coupled class A GPCRs in cellular conditions. The second aim is to understand the effect of partial agonists in comparison to the full agonists, on the GPCR-G-protein selective coupling. We will identify the hotspot residues on the G-protein and GPCRs when bound to partial agonists and compare the similarities and differences with full agonists. The outcome of the proposed work will provide the role of allosteric network of residues in G-protein selectivity and how partial agonists modulate the receptor conformations. This will push forward our understanding of the biological process of how GPCRs recognize their cognate G-proteins in live cells.