Functional Determinants in G Protein-Coupled Receptors

描述（由申请人提供）：这项工作的长期目标是合理操纵G蛋白偶联受体（gpcr）的激活，希望在这一主要的跨膜受体家族和药物靶点中开发新的治疗方法。我们的假设是，不同的gpcr具有其信号转导机制的共同要素。如果是这样，应该有可能比较和对比它们的序列，以揭示功能相关的氨基酸变异模式，其中一些是所有受体共有的，表明了共同的机制，而另一些是家族中某些分支所特有的，表明了配体特异性机制。这一原则导致了一种称为进化轨迹（ET）的通用算法，该算法将残基替换与进化分歧联系起来，从而对蛋白质残基的进化重要性进行排序。研究表明，在过去的资助期内，ET可以大规模地搜索蛋白质结构的功能位点及其特异性决定因素。在GPCRs和其他蛋白质的大量实验案例研究中，突变引导到ET的顶级残基（所谓的“微量残基”）反复阻断、分离、模仿或重新连接蛋白质相互作用。同时，高通量ET分析表明，微量残基在结构聚类、生物物理相互作用和功能特异性方面具有独特的、可量化的和蛋白质组范围的特性。本建议以这些观察结果为基础，力求实现三个具体目标：重定向生物胺受体中的配体结合。2. 重新设计代谢性谷氨酸受体的配体结合和二聚化。3. 大规模鉴定跨膜GPCR反应调节因子。该结果将揭示一个重要药物靶点家族信号传导分子基础的新方面。它还将把序列和结构基因组数据库与功能的分子基础和蛋白质相互作用的合理重新设计联系起来——这是操纵细胞途径的关键步骤。它将为合理的药物设计提供新的途径，并提高SNP分析和人类基因分型的诊断价值，从而造福人类健康。公共卫生相关性：迄今为止，药物成功靶向的少数G蛋白偶联受体仍然为近一半的现有药物提供了基础。制造针对它们的药物的困难在于我们对它们如何起作用的知识有限。这项提议试图揭示这些蛋白质的机制，这样我们就可以设计出新的药物来阻止它们在疾病中的作用。

DESCRIPTION (provided by applicant): The long-term goal of this work is to rationally manipulate G protein-coupled receptor (GPCRs) activation with the hope to develop novel therapeutic approaches in this major family of transmembrane receptors and drug targets. Our hypothesis is that different GPCRs share common elements of their signal transduction mechanism. If so, it should be possible to compare and contrast their sequences to reveal functionally relevant amino acid variation patterns, some that are common to all receptors and indicative of shared mechanisms, and others that are unique to some branches of the family and indicative of ligand-specific mechanisms. This principle led to a general algorithm, called the Evolutionary Trace (ET) that correlates residue substitutions with evolutionary divergences and thus ranks the evolutionary importance of a protein's residues. It was shown, in the past funding period that ET could search protein structure for functional sites and their specificity determinants on a large-scale. Mutations guided to ET's top-ranked residues (so-called "trace residues") repeatedly blocked, separated, mimicked, or rewired protein interactions in numerous experimental case studies in GPCRs and in other proteins. In parallel, high-throughput ET analyses suggested that trace residues have distinctive and quantifiable and proteome-wide properties in terms of structural clustering, biophysical interaction, and functional specificity. This proposal builds on these observations by pursuing three specific aims: 1. To redirect ligand binding in bioamine receptors. 2. To redesign ligand binding and dimerization in metabotropic glutamate receptors. 3. To identify transmembrane GPCR response modulators on a large scale. The outcome should reveal new aspects of the molecular basis of signaling in an important family of pharmaceutical targets. It will also link sequence and structure genomics databases to the molecular basis of function and to the rational re-design of protein interactions - key steps towards manipulating cellular pathways. It will benefit human health by providing new approaches to rational drug design and by enhancing the diagnostic value of SNP analysis and human genotyping. PUBLIC HEALTH RELEVANCE: The few G protein coupled receptors that have been successfully targeted by drugs so far still provide the basis for nearly half of all current medications. The difficulty in creating medications against them is that we have limited knowledge of how they work. This proposal attempts to uncover the mechanisms of these proteins so that we can design new drugs that block their role in diseases.