Membrane Proteins. Crystallogenesis and X-ray Structure

描述（由申请人提供）：为了理解膜蛋白的功能而解决结构仍然是所有生物学中的重大挑战之一。鉴于人类基因组的三分之一编码膜蛋白，其中许多蛋白起信号传导、结构和运输作用，是疾病病原体和治疗剂的目标，迎接这一挑战的健康后果是巨大的。一个多方面的方法将采取生产用于大分子晶体学的结构级晶体，以促进我们对膜蛋白功能的理解。重点放在结晶在介晶中间相什么是所谓的'在介'或立方相的方法。这被证明是一个普遍有用的方法膜蛋白结构的测定。用这种方法，结晶发生在膜环境中，这可能是有利的目标膜蛋白。我们已经建立了一个独特的，国家的最先进的机器人，在介晶高通量的方式进行，需要微量的蛋白质，脂质和沉淀剂。它将在当前的应用中用于生产3-D晶体，用于选择一组膜蛋白的高分辨率结构测定。目标组涵盖广泛的膜蛋白类型，包括真核生物和原核生物、整体和外周、单体和多聚体，以及蛋白质-蛋白质和蛋白质-肽复合物。一些目标蛋白质将在内部生产，一些将由个人合作者提供，而另一些将通过NIH结构蛋白质组学计划提供。 与NIH结构生物学路线图一致，并与拟议的结构研究平行，将致力于改善晶体发生和扩大膜蛋白靶点的范围，以确定结构。这将通过实施合成程序来实现，由此产生具有期望的物理化学特性的脂质以用于结晶试验。晶体成核和生长的分子机制也将被研究，以期更合理和成功的晶体发生。成功获得晶体，并最终获得膜蛋白的原子结构，将增强我们对人类健康所不可或缺的细胞功能的一些最基本过程的理解。

DESCRIPTION (provided by applicant): Solving the structure with a view to understanding the function of membrane proteins remains one of the grand challenges in all of Biology. Given that a third of the human genome codes for membrane proteins, many of which serve signal transducing, structural and transport roles and are targets of disease causative and treatment agents, the health consequences of meeting the challenge are great. A multifaceted approach will be taken to advance our understanding of membrane protein function by producing structure grade crystals for use in macromolecular crystallography. Emphasis is placed on crystallization in lipidic mesophases by what is referred to as the 'in meso' or cubic phase method. This is proving to be a generally useful approach for membrane protein structure determination. With this method crystallization takes place in a membrane environment that is likely to be favored by the target membrane protein. We have built a unique, state-of-the-art robot that performs in meso crystallization in high-throughput fashion and that requires miniscule amounts of protein, lipid and precipitant. It will be used in the current application to produce 3-D crystals for the high-resolution structure determination of a select group of membrane proteins. The target group covers a broad range of membrane protein types including eukaryote and prokaryote, integral and peripheral, monomeric and multimeric, as well as protein-protein and protein-peptide complexes. Some of the target proteins will be produced in-house, some will be provided by individual collaborators, while others will be supplied through the NIH Structural Proteomics Initiative. In line with the NIH Structural Biology Road Map, and in parallel with the proposed structure study, effort will be devoted to improving crystallogenesis and to broadening the range of membrane protein targets that yield to structure determination. This will be achieved by implementing a synthesis program whereby lipids with desirable physico-chemical characteristics are produced for use in crystallization trials. The molecular mechanism of crystal nucleation and growth will be studied too with a view to more rational and successful crystallogenesis. Success in obtaining crystals, and ultimately the atomic structure of membrane proteins, will enhance our understanding of some of the most fundamental processes underlying cellular function that are integral to human health.