Deciphering GPCR signal transduction through NMR structure and dynamics studies

通过 NMR 结构和动力学研究破译 GPCR 信号转导

基本信息

批准号：
9135502
负责人：
Joshua James Ziarek
金额：
$ 9万
依托单位：
HARVARD MEDICAL SCHOOL
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-09-01 至 2017-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9135502
关键词：
Adrenergic Receptor Agonist Amino Acids Applied Research Area Arrestins Basic Science Binding Biochemical Biochemistry Boston Cells Chemicals Chemistry Chimeric Proteins Codon Nucleotides Communities Complex Comprehension Data Analyses Data Collection Development Diphosphates Directed Molecular Evolution Discipline Disease Environment Escherichia coli Faculty Funding G Protein-Coupled Receptor Signaling G protein coupled receptor kinase G-Protein-Coupled Receptors G-substrate GTP-Binding Proteins Gatekeeping Gene Silencing Guanosine Triphosphate Health Heterotrimeric GTP-Binding Proteins Human Insecta Institutes Isotope Labeling Label Laboratories Laboratory Research Lead Length Ligand Binding Ligands Marketing Mass Spectrum Analysis Measures Membrane Proteins Mentors Methods Molecular Biology Monitor Motion Muramidase Mutation NMR Spectroscopy Neurotensin Neurotensin Receptors Nucleotides Outcome Pattern Pharmaceutical Preparations Phase Phosphopeptides Phosphorylation Phosphotransferases Physics Protein Biochemistry Protein Family Protein Kinase C Proteins Proteome Public Health Reaction Receptor Activation Research Role Scheme Scientist Serine Signal Pathway Signal Transduction Social Sciences Specificity Structure Surface System Techniques Technology Testing Threonine Training Translating United States National Institutes of Health Universities Work X-Ray Crystallography analog arrestin 1 arrestin 2 base drug discovery empowered extracellular flexibility human disease insight interest macromolecule medical schools member milligram millisecond molecular recognition nanobodies nanodisk nanosecond professor programs protein complex receptor receptor structure function receptor-mediated signaling research study sortase structural biology theories

项目摘要

DESCRIPTION (provided by applicant): G protein-coupled receptors (GPCRs) are critical eukaryotic signal transduction gatekeepers and represent the largest protein family in the human proteome, with more than 800 members. In recent years, X-ray crystallography has yielded many G protein-coupled receptor structures but the mechanism of allosteric signaling remains unknown. X-ray crystallography of GPCRs requires multiple forms of receptor stabilization that, subsequently, limit conformational dynamics. RESEARCH: Directed evolution of GPCRs enables milligram quantities of functional, isotopically-labeled protein to be produced from prokaryotic expression systems. This technology opens the field for NMR studies of GPCRs with various ligands and in complex with G proteins and arrestins. Focusing on the neurotensin receptor, the PI will use NMR spectroscopy to probe the structure and conformational dynamics of GPCR activation in solution. TRAINING: The proposed training in Professor Wagner's laboratory at Harvard Medical School will solidify the PI's comprehension of NMR theory and implementation. In addition to advanced NMR techniques, the PI will acquire a thorough understanding of membrane protein biochemistry, nanodiscs, and directed evolution. Together, these techniques will empower the PI to establish an independent, NIH-funded structural biology research program. ENVIRONMENT: Professor Gerhard Wagner's laboratory at Harvard University Medical School presents the optimal environment for this project. His work has been integral to the development of multidimensional NMR experiments for biomolecule characterization; in particular, Dr. Wagner is a leader in data collection and analysis of large macromolecules. Harvard Medical School has nearly 8,000 faculty and 17 affiliated facilities. At the core of the Medical School are its educational and research programs. The Medical School has nine departments in basic and social science disciplines. The wider Boston/Cambridge area contains numerous research laboratories including MIT, Whitehead, and Broad Institutes with interests in structural biology, biochemistry, molecular biology, chemistry and physics, with scientists interested in the general themes related to this proposal. IMPACT ON PUBLIC HEALTH: GPCR-mediated signaling pathways have been related to numerous human diseases, and GPCRs are the targets of an estimated 30-40% of all drugs currently on the market. In view of their fundamental roles in health and disease, a detailed understanding of GPCR structure and function is of value to the basic science community interested in cell signaling and molecular recognition, as well as to the applied science community interested in drug discovery.

描述（由应用提供）：G蛋白偶联受体（GPCR）是关键的真核信号转导守门者，代表了人类蛋白质中最大的蛋白质家族，拥有800多名成员。近年来，X射线晶体学产生了许多G蛋白偶联受体结构，但变构信号传导的机制仍然未知。 GPCR的X射线晶体学需要多种形式的受体稳定，随后限制了会议动力学。研究：GPCR的定向演变使毫克量的功能性，同位素标记的蛋白可以由原核表达系统产生。这项技术为使用各种配体的GPCR进行NMR研究开放，并与G蛋白和逮捕蛋白复杂化。 PI专注于神经素受体，将使用NMR光谱探测溶液中GPCR激活的结构和构象动力学。培训：哈佛医学院瓦格纳教授实验室的拟议培训将巩固PI对NMR理论和实施的理解。除了先进的NMR技术外，PI还将对膜蛋白生物化学，纳米盘和定向进化有深入的了解。这些技术共同使PI能够建立独立的，NIH资助的结构生物学研究计划。环境：哈佛大学医学院的Gerhard Wagner教授实验室展示了该项目的最佳环境。他的工作一直是生物分子表征多维NMR实验的开发不可或缺的一部分。特别是，瓦格纳博士是数据收集和分析大型大分子的领导者。哈佛医学院拥有近8,000名教职员工和17个会员设施。医学院的核心是其教育和研究计划。医学院拥有基础和社会科学学科的九个部门。更广泛的波士顿/剑桥地区包含许多研究实验室，包括麻省理工学院，怀特海和广泛的机构，对结构生物学，生物化学，分子生物学，化学和物理学有兴趣，以及对与该建议相关的一般主题感兴趣的科学家。对公共卫生的影响：GPCR介导的信号通路与许多人类疾病有关，GPCR是目前市场上所有药物的30-40％的目标。鉴于它们在健康和疾病中的基本作用，对GPCR结构和功能的详细了解对对细胞信号传导和分子识别感兴趣的基础科学界以及对药物发现感兴趣的应用科学界有价值。