INVESTIGATING STRUCTURAL BASES OF SIGNAL TRANSDUCTION: PROTEIN ENG

研究信号转导的结构基础：蛋白质工程

• 批准号: 6456752
• 负责人: ELAINE MENG
• 金额: $ 27.32万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-07-01 至 2003-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6456752
• 关键词: biological products; biomedical resource; enzymes

We are studying the structure and function of signal transduction proteins, specifically heterotrimeric GTP-binding proteins (G proteins) and G protein-coupled receptors (GPCRs). When an agonist such as a hormone or neurotransmitter binds its receptor, exchange of GTP for GDP bound to the G protein is catalyzed. The GTP-bound alpha subunit of the G protein separates from the beta-gamma subunit complex, and each of these can go on to stimulate downstream effectors. The hydrolysis of GTP to GDP by the alpha subunit and subsequent heterotrimer reassembly turns off the signal. While the structures of some G protein subunits have been solved, atomic-level structures have not been determined for the receptors. Our specific goals are to understand the structural basis of function by designing and evaluating mutant forms of the G proteins and receptors. One category of mutants contains two or more histidines placed so that they could be bridged by a metal ion. Such a link could cause metal-dependent activation or inactivation of the protein. In known structures, this would enhance our knowledge of the activation process; in unknown structures, this would also yield distance constraints. Another category of constructs are fusions to fluorescent proteins or mutants that contain a tetracysteine motif designed to bind the small fluorescent molecule FLASH. The fluorescence emitted by the resulting species will give us information on subcellular locations of the proteins, and fluorescence resonant energy transfer (FRET) between such moieties will show association and dissociation of these proteins during signal transduction cascades. The CGL facilities are crucial for many steps of this research: viewing crystallographic or modelled structures to design mutants, rationalizing the properties of the mutants, and to develop hypotheses about protein-ligand and protein-protein interactions, and about conformational changes involved in activation and deactivation.

我们正在研究信号转导的结构和功能