New Approaches for Protein Crystallization, Cryopreservation and Radiation Damage

描述（由申请人提供）：项目概述：美国国立卫生研究院已承诺大量资源用于传统和-通过其蛋白质结构倡议-蛋白质结构测定的高通量方法，欧洲和亚洲也做出了类似的承诺。对可溶性蛋白的研究结果的调查表明，获得高质量的晶体和获得精确结构的高质量x射线衍射数据是主要的瓶颈。该项目的长期目标是了解大分子晶体生长、蛋白质晶体冷冻保存和x射线数据收集期间辐射损伤的基础科学，并利用这种理解开发实用的方案和技术。这些研究利用了广泛的物理方法，从基于同步加速器的x射线衍射和成像到微加工。我们的具体目标是：(1)了解低温保存的基本过程，并基于超快速冷却，水混合物的动力学相图以及溶剂和晶体的热膨胀行为，开发更有效的方法来捕获和保存蛋白质和晶体结构；(2)。开发冷冻保存和长期储存蛋白质溶液的方法，以保持结晶行为；(3)量化和了解蛋白质晶体中的辐射损伤，并制定减少小晶体中这种损伤的方案；(4)探索与蛋白质结晶相关的蛋白质滴-表面相互作用和滴-动力学。相关性：蛋白质晶体学是现代结构基因组学和药物发现工作的核心组成部分，有望在21世纪彻底改变医学。拟议的研究将有助于更快速和准确地确定蛋白质结构，从而有助于了解疾病的潜在机制和开发新的治疗方法。

DESCRIPTION (provided by applicant): Project Summary: The NIH has committed substantial resources to both traditional and - through its Protein Structure Initiative - high-throughput approaches to protein structure determination, and similar commitments have been made in Europe and Asia. A survey of the results for soluble proteins shows that the main bottlenecks are obtaining high-quality crystals and obtaining high-quality X-ray diffraction data for accurate structures. The long-term goals of this project are to understand the underlying science involved in macromolecular crystal growth, in cryopreservation of protein crystals, and in radiation damage during X-ray data collection, and to use this understanding to develop practical protocols and technologies. These studies utilize a wide array of physical methods, from synchrotron-based X-ray diffraction and imaging to microfabrication. Our specific goals are: (1) To understand the fundamental processes in cryopreservation, and to develop more effective methods to capture and preserve protein and crystal structure based on ultra- fast cooling, the kinetic phase diagrams of aqueous mixtures, and the thermal expansion behavior of solvent and crystal; (2) To. develop methods for cryopreservation and long-term storage of protein solutions that preserve crystallization behavior; (3) To quantify and understand radiation damage in protein crystals, and to develop protocols for reducing this damage in small crystals; and (4) To explore protein drop-surface interactions and drop dynamics relevant to protein crystallization. Relevance: Protein crystallography is a central component of modern structural genomics and drug discovery efforts, which promise to revolutionize medicine in the 21st century. The proposed research will facilitate more rapid and accurate determination of protein structures, and thereby assist in understanding the mechanisms underlying disease and in developing new treatments.