MOLECULAR STRUCTURE OF G PROTEIN COUPLED RECEPTORS

我的长期目标是阐明趋化因子受体的识别和激活的分子机制，趋化因子受体是G蛋白偶联受体的一个亚家族，在炎症、血管生成、肿瘤形成以及作为HIV-1进入的辅助受体方面发挥关键作用。该项目的目标是与巴塞尔大学生物中心的兰道博士合作，生产有序的G蛋白偶联受体的3-D晶体。我们计划结晶并解决头足类视紫红质的结构，这是一个原型(3蛋白偶联受体。这种受体在光感受器膜上高度表达，在洗涤剂中稳定，并在重组为脂泡时组装成二维晶体。为了形成三维晶体，我们将使用朗道博士开发的新程序，该程序包括将蛋白质插入到由脂质立方相提供的连续三维双层基质中。使用这种方法，兰道博士结晶了细菌视紫红质，一种光驱动的质子泵，并以2.5埃的分辨率解决了这种蛋白质的结构(Science 277,1676-1680,1997)。阐明o蛋白偶联受体的结构对于从分子角度解释许多关于这些蛋白的遗传和生物物理研究是至关重要的。我计划访问三次，每次四个月。结晶和X射线实验对申请者来说是新的研究领域，因此第一学期最有可能涉及学习蛋白质结晶和一些X射线分析。第二和第三个学期将涉及从不同头足类视紫红质中提取的视紫红质的结晶试验，以及一些初步的X射线实验。

DESCRIPTION My long term goal is to elucidate the molecular mechanisms of recognition and activation of chemokine receptors, a subfamily of G protein-coupled receptors, which plays a key role in inflammation, angiogenesis, tumorogenesis and as co-receptors for the entry of HIV-1. The aim of this project is to produce well ordered 3-D crystals of a G protein-coupled receptor in collaboration with Dr. Landau at the Biocenter in the University of Basel. We plan to crystallize and solve the structure of cephalopod rhodopsin, a prototype (3 protein-coupled receptor. This receptor is highly expressed in photoreceptor membranes, is stable in detergents and upon reconstitution into lipid vesicles assembles into two-dimensional crystals. For the formation of three- dimensional crystals we will employ the novel procedure developed by Dr. Landau, which involves insertion of the protein into a continuous three- dimensional bilayer matrix afforded by lipid cubic phases. Using this approach Dr. Landau crystallized bacteriorhodopsin, a-light-driven proton pump, and solved the structure of this protein at a resolution of 2.5 angstroms (Science 277, 1676-1680, 1997). Elucidation of the structure of a o protein-coupled receptor will be essential to interpret the many genetic and biophysical studies on these proteins in molecular terms. I am planning three visits, each of four months. Crystallization and X-ray experiments are new areas of research for the applicant, therefore the first term most likely will involve learning crystallization of proteins and some X-ray analysis. The second and third terms will involve crystallization trials with rhodopsins from different cephalopod rhodopsins, and some preliminary X-ray experiments.