Enhancing crystallization with binding partners, symmetry and diversity

描述(申请人提供)：X射线结晶学确定结构的瓶颈是结晶，大约70%的纯化蛋白质失败。蛋白质无法进入结晶态的两个主要原因是晶格接触太少，以及具有多个构象，这通常是缺少配对蛋白质的结果。这个项目的目的是克服结晶的障碍。该项目的关键总体思想是，可以通过(1)以高度适合结晶的形式呈现这种大分子和(2)产生多种不同形式的大分子来改善大分子或络合物的结晶。这些优化结晶性和变化性的主题是该项目所有三个组成部分的核心。我们建议开发的创新新方法将使用天然结合伙伴和结合模块的组合来改善目标大分子的结晶，方法是(1)创建多种不同形式的分子进行结晶，(2)寻找稳定和增强目标大分子结晶能力的天然伙伴大分子。许多形式的分子将使用一组对称形成模块来创建，这些模块可以连接到目标分子。这些模块将是基于抗体或绿色荧光蛋白的。结合新的生物信息学和实验方法，将会发现天然的结合伙伴。转化方法是可扩展的、协同的，通过提供一个分子工具包来多样化目标蛋白质的潜在结晶排列和对称性，从而提供了加速结构生物学和结构基因组学结果的潜力。该计划项目将由加州大学洛杉矶分校/洛斯阿拉莫斯大学团队作为三个紧密结合的子项目来实施，每个子项目都涉及来自加州大学洛杉矶分校和洛斯阿拉莫斯大学的研究人员。这项工作将导致将被结构生物界用来确定蛋白质结构的方法，这些方法将增加我们对人类健康的理解和我们治愈人类疾病的能力。

DESCRIPTION (provided by applicant): The bottleneck in structure determination by X-ray crystallography is crystallization, where roughly 70% of purified proteins fail. Two major reasons proteins fail to enter the crystalline state are having too few lattice contacts, and having multipe conformations, often the result of missing partner proteins. The purpose of this project is to overcome barriers to crystallization. The key overall ideas in this project are that crystallizatio of a macromolecule or complex can be improved by (1) presenting this macromolecule in a form that is highly suitable for crystallization and (2) creating many different forms of the macromolecule. These themes of optimizing crystallizability and variation are central to all three components of this projects. The innovation new methods we propose to develop will use a combination of natural binding partners and binding modules to improve crystallization of a target macromolecule by (1) creating many different forms of a molecule to crystallize and (2) finding a natural partner macromolecule that stabilizes and enhances the crystallizability of a target macromolecule. Many forms of a molecule will be created using a panel of symmetry-forming modules that can be linked to the target molecule. These modules will be antibody- or green fluorescent protein-based. Natural binding partners will be found with a combination of novel bioinformatics and experimental approaches. The transformative methods are scalable, synergistic, and offer the potential of accelerating results in both structural biology and structual genomics by providing a molecular toolkit for diversifying the potential crystallization arrangements and symmetries of targeted proteins. This program project will be carried out by our UCLA/ Los Alamos team as three tightly integrated subprojects, each involving researchers from both UCLA and Los Alamos. This work will lead to methods that will be used by the structural biology community to determine structures of proteins that will increase our understanding of human health and our ability to cure human disease.