Probing the Structural Basis of Innate G Protein Specificity in G Protein-Coupled Receptor Signaling

探讨G蛋白偶联中天然G蛋白特异性的结构基础 受体信号传导 摘要 G蛋白偶联受体(GPCRs)是一种完整的膜蛋白，直接与 它们的信号伙伴如G蛋白、GPCRK(GRKs)和Drastins，以响应 细胞外信号，并可触发控制范围的多个细胞内信号级联 重要的生物过程。细胞已经进化出GPCR介导的信号机制，其中 配体和受体发挥着独特的功能作用。每个受体都进化为主要激活 G蛋白的一个亚家族，它触发一系列特定的信号级联反应。配基是更重要的 功能强大，因为它可以激活多种受体亚型，进而激活多组 比起单独的受体，信号级联会产生更深远的细胞影响。平凡的 配体和受体功能的特征是配体诱导的天然G蛋白的特异性 感受器。这项建议旨在探讨受体亚型的G蛋白特异性 由同一配体激活(使用毒鼠碱型乙酰胆碱受体和多巴胺受体 作为例子)，由计算生物物理方法的发展将探索 观察到的G蛋白选择性背后的热力学和动力学因素。这些方法将 为多巴胺D1和D2受体中G蛋白特异性提供结构基础，这将是 用于设计具有不同于野生型G蛋白特异性的受体突变体 感受器。这些方法将通过使用这些突变的基于细胞的生化分析来验证 受体，部分通过复制先前观察到的G蛋白选择性开关 毒碱受体突变体。这种机械的理解将对以下方面产生更广泛的影响 许多GPCR亚家族，并将为针对特定治疗的药物奠定基础 有益的受体-G蛋白相互作用，以减少靶上和靶外副作用。

Probing the Structural Basis of Innate G Protein Specificity in G Protein-Coupled Receptor Signaling Abstract G protein-coupled receptors (GPCRs) are integral membrane proteins that directly couple with their signaling partners like G proteins, GPCR kinases (GRKs), and arrestins, in response to extracellular signals and can trigger multiple intracellular signaling cascades that control a range of vital biological processes. Cells have evolved GPCR-mediated signaling mechanisms, where ligands and receptors play unique functional roles. Each receptor has evolved to activate mainly one subfamily of G proteins, which triggers a specific set of signaling cascades. A ligand is more powerful as it can activate multiple receptor subtypes, which in turn activate multiple set of signaling cascades that causes far reaching cellular effects than a receptor alone. The common feature of ligand and receptor function is the ligand-induced innate G protein specificity of the receptors. This proposal is aimed at probing the G protein specificity of receptor subtypes activated by the same ligand (using muscarinic acetylcholine receptors and dopamine receptors as examples), by the development of computational biophysical methods that will explore thermodynamic and kinetic factors behind the observed G protein selectivities. The methods will provide a structural basis for G protein specificity in dopamine D1 and D2 receptors, which will be used to design receptor mutants with G protein specificity different from that of the wild-type receptors. The methods will be validated by cell based biochemical assays using these mutant receptors and in part by reproducing previously observed switches in G protein selectivity of muscarinic receptor mutants. This mechanistic understanding will have broader implications for many GPCR subfamilies and will lay the foundation for drugs targeting specific therapeutically beneficial receptor – G protein interactions to reduce on-target and off-target side-effects.