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，特别是异源三聚体GTP结合蛋白（G 蛋白质）和G蛋白偶联受体（GPCR）。 当激动剂 如激素或神经递质结合其受体， 与G蛋白结合的GDP的GTP被催化。 GTP结合的alpha G蛋白的亚基与β-γ亚基分离 每一个都可以刺激下游 效应器 GTP通过α亚基水解为GDP， 随后的杂三聚体重组关闭了信号。 而 一些G蛋白亚基的结构已经在原子水平上得到了解决， 受体的结构尚未确定。 我们的具体 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 一类突变体将含有两个或更多个组氨酸， 它们可以通过金属离子桥接。 这种联系可能会导致 蛋白质的金属依赖性活化或失活。 以已知 结构，这将增强我们对激活的知识， 过程;在未知的结构中，这也将产生关于 蛋白质区域的邻近性。 另一类变种人 源于随机突变。 实验包括纯化蛋白质 测定、哺乳动物细胞系中的功能测定和酵母生长 其中“读出”是大量受体序列的测定 能够发出信号。 计算机图形实验室设施 对许多步骤至关重要， 这项研究：观察晶体学或模型结构， 设计突变体，使突变体的特性合理化， 提出蛋白质-配体和蛋白质-蛋白质假说 相互作用，并发展构象变化的假设 参与激活和去激活。

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.