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：
8950825
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/8950825
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 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 Solutions 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 directed evolution drug discovery empowered extracellular flexibility human disease insight interest macromolecule medical schools member milligram millisecond molecular recognition nanobodies nanodisk nanosecond professor programs protein complex public health relevance 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（由申请人提供）：G蛋白偶联受体（GPCR）是关键的真核信号转导看门人，代表人类蛋白质组中最大的蛋白质家族，拥有800多个成员。近年来，X射线晶体学已经产生了许多G蛋白。 GPCR 的 X 射线晶体学需要多种形式的受体稳定，从而限制构象。研究：GPCR 的定向进化使得原核表达系统能够产生毫克量的功能性同位素标记蛋白质，该技术为 GPCR 与各种配体以及与 G 蛋白和抑制蛋白复合物的 NMR 研究开辟了领域。对于神经降压素受体，PI 将使用 NMR 光谱来探测溶液中 GPCR 激活的结构和构象动力学。培训：建议在哈佛医学院 Wagner 教授的实验室进行培训。学校将巩固 PI 对 NMR 理论和实施的理解，除了先进的 NMR 技术外，PI 还将获得对膜蛋白生物化学、纳米圆盘和定向进化的透彻理解，这些技术将使 PI 能够建立独立的、美国国立卫生研究院 (NIH) 资助的结构生物学研究项目。环境：哈佛大学医学院的格哈德·瓦格纳 (Gerhard Wagner) 教授的实验室为该项目提供了最佳环境，他的工作对于生物分子多维核磁共振实验的发展至关重要。哈佛医学院拥有近 8,000 名教职员工和 17 个附属设施，其教育和研究项目是该学院的核心。波士顿/剑桥地区有九个基础和社会科学学科系，拥有众多研究实验室，包括麻省理工学院、怀特海德研究所和博德研究所，对结构生物学、生物化学、分子生物学、化学和物理学感兴趣，科学家们对这些领域感兴趣。与该提案相关的一般主题：GPCR 介导的信号通路与多种人类疾病有关，鉴于其基本原理，GPCR 是目前市场上估计 30-40% 的药物的靶标。由于 GPCR 在健康和疾病中的作用，详细了解 GPCR 结构和功能对于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都很有价值。