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Structural Studies in Rhodopsin and G Protein Activation

视紫红质和 G 蛋白激活的结构研究

基本信息

批准号：
8134295
负责人：
DAVID THOMAS LODOWSKI
金额：
$ 9万
依托单位：
CASE WESTERN RESERVE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-09-01 至 2012-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8134295
关键词：
Agonist Binding Binding Proteins Biochemical Biological Models Blood Pressure Cell membrane Complex Couples Crystallization Crystallography Data Disease Event Fluorescence Funding G-Protein-Coupled Receptors GTP-Binding Protein alpha Subunits, Gs GTP-Binding Proteins Goals Heart Rate Human Genome Imagery Individual Knowledge Ligands Light Membrane Methodology Methods Modeling Molecular Molecular Probes Movement Neutron Diffraction Nucleotides Pathology Pathway interactions Peptides Physiological Processes Play Process Protein Binding Protein Subunits Proteins Protons Publishing Receptor Activation Regulation Research Resolution Rhodopsin Roentgen Rays Role Side Signal Transduction Smell Perception Structure System Taste Perception Techniques Therapeutic Transducin Vision Visual system structure Water Work X-Ray Crystallography base crosslink design electron crystallography extracellular heart metabolism member particle protein activation protein complex protonation public health relevance reconstruction tool

项目摘要

DESCRIPTION (provided by applicant): G protein coupled receptors (GPCRs) are a universally conserved signalling mechanism by which extracellular signals can be transduced across the plasma membrane. They make up ~3% of the human genome and ~50% of all non-antimicrobial therapeutics act upon GPCRs or their pathways. Upon binding of agonist, structural changes activate the GPCR, and allow the GPCR to bind and induce nucleotide exchange upon the alpha subunit of the heterotrimeric G protein complex. Rhodopsin represents perhaps the best understood GPCR for both GPCR activation by ligand and for the consequent binding and induction of nucleotide exchange upon the G alpha subunit of its cognate G protein, transducin. Furthermore, rhodopsin and the rhodopsin: transducin complex has been used as a prototypical GPCR to model the process of activation in general among all GPCRs. While structures of both the ground state and several photo intermediate states of rhodopsin have been solved through a variety of structural techniques, a significant gap exists in our knowledge. In order to fully understand the process of activation in rhodopsin as well we need to determine the precise molecular interactions that are responsible for (1) the process of rhodopsin going from the ground (dark) state to the fully active (Meta II) state and (2) the structural determinants for transducin binding and nucleotide exchange and activation. A high resolution structure of the Meta II activated state is needed to fully understand the sequence of events that comprises rhodopsin activation. Similarly, a large volume of biochemical and biophysical studies on transducin and G protein activation have been conducted, but without a structure of this complex, it is difficult to understand the underlying molecular basis for binding and nucleotide exchange. This proposal seeks to determine the fundamental changes in rhodopsin structure that accompany its transition to the activated Meta II state. Furthermore, we will determine the structure of the complex between rhodopsin and transducin, which will elucidate the molecular mechanisms of G protein binding and activation. PUBLIC HEALTH RELEVANCE: Structural studies of the molecular interactions that allow the sensing of light by the protein, rhodopsin, will greatly increase our knowledge of how this system functions as well as the underlying pathologies of some blinding diseases. Furthermore, as rhodopsin is a member of the G protein coupled receptor superfamily, it will serve to explain how important physiological processes, such as sight, smell, taste; heart rate and metabolism are governed.

描述（由申请人提供）：G蛋白偶联受体（gpcr）是一种普遍保守的信号传导机制，细胞外信号可以通过其传导穿过质膜。它们占人类基因组的约3%，所有非抗菌药物的约50%作用于gpcr或其途径。在与激动剂结合后，结构变化激活GPCR，并允许GPCR结合并诱导异三聚体G蛋白复合物α亚基上的核苷酸交换。视紫红质可能是最被理解的GPCR，既可以通过配体激活GPCR，也可以随后结合并诱导其同源G蛋白的G α亚基上的核苷酸交换。此外，视紫红质和视紫红质：转导蛋白复合物已被用作GPCR的原型来模拟所有GPCR的激活过程。虽然通过各种结构技术已经解决了紫红质基态和几个光中间态的结构，但在我们的知识中存在显着的差距。为了充分了解视紫红质的激活过程，我们需要确定精确的分子相互作用，这些相互作用负责(1)视紫红质从基态（暗态）到完全活性（Meta II）状态的过程，以及(2)转导蛋白结合和核苷酸交换和激活的结构决定因素。需要一个高分辨率的Meta II激活状态结构，以充分了解包括视紫红质激活的事件序列。同样，对转导蛋白和G蛋白活化进行了大量的生化和生物物理研究，但如果没有这种复合物的结构，就很难理解其结合和核苷酸交换的潜在分子基础。该提案旨在确定视紫红质结构伴随其过渡到活化的Meta II状态的基本变化。此外，我们将确定视紫红质和转导蛋白之间复合物的结构，这将阐明G蛋白结合和激活的分子机制。