INVESTIGATING STRUCTURAL BASIS OF SIGNAL TRANSDUCTION

我们正在研究信号转导的结构和功能 蛋白质，特别是异三聚体GTP结合蛋白(G 蛋白质)和G蛋白偶联受体(GPCRs)。当一种激动剂 如激素或神经递质结合其受体，交换 GDP的GTP与G蛋白结合是催化的。GTP绑定的Alpha G蛋白的亚基与β-伽马亚基分离 复杂，每一种都可以继续刺激下游 效应器。GTP被α亚基水解为GDP和 随后的异三聚体重组关闭了信号。而当 一些G蛋白亚基的结构已经在原子水平上被解决 受体的结构尚未确定。我们的特定 目标是通过设计来理解功能的结构基础 以及评估G蛋白和受体的突变形式。一 一类突变体将包含两个或更多组氨酸，以便 它们可以被金属离子连接起来。这样的联系可能会导致 依赖金属的蛋白质的激活或失活。在已知中 结构，这将增强我们对激活的知识 进程；在未知结构中，这也将产生有关 蛋白质各区域的接近程度。另一类突变体 来自随机突变。实验包括纯化的蛋白质 哺乳动物细胞系的功能分析和酵母菌的生长 “读出”是大量受体序列的化验 能够发出信号的。计算机图形学实验室设施 是许多步骤的关键 这项研究：观察结晶学或模型化结构以 设计突变体，使突变体的属性合理化，以 发展蛋白质-配体和蛋白质-蛋白质假说 相互作用，并发展构象变化的假说 参与激活和停用。

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 hterotrimer 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 will contain 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 information on the proximity of regions of the protein. Another category of mutants arises from random mutagenesis. Experiments include purified protein assays, functional assays in mammalian cell lines, and a yeast growth assay where the "readout" is a large number of receptor sequences capable of signalling. The Computer Graphics Laboratory facilities are crucial for many steps of this research: viewing crystallographic or modelled structures to design mutants, to rationalize the properties of the mutants, to develop hypotheses about protein-ligand and protein-protein interactions, and to develop hypotheses of conformational changes involved in activation and deactivation.