X-RAY STUDIES OF CHEY--CHEMOTAXIS PROTEIN FROM E. COLI

这项研究计划的基本目标是 研究一种主要趋化性的结构和功能 对照蛋白E.大肠杆菌中，在分子水平上 具体地说， 目的是确定三维分子结构 的CheY蛋白的X-射线衍射方法，并涉及 其正常和突变形式的结构，以其功能， 细菌趋化性的控制系统。 细菌趋化系统是研究细菌趋化性的理想模型 细胞行为反应的分子控制。 详细 对系统组织的理解现在存在于 基因水平;解释控制系统的趋化性上 分子基础，三维结构的知识， 其蛋白质成分是必不可少的。 cheY基因产物， 由于其在趋化性信号处理中的核心作用， 系统，已被选为第一个组件进行分析， x射线衍射 在解决这三个问题的过程中， Chey Y的空间结构在于它的主序列 与其他七种蛋白质同源，所有调节剂在各种 细胞功能。 这些同源性使CheY具有代表性 一种新型的细菌调节域 proteins. CheY已被分离和测序， 也已经被测序和表型表征。 用于获得本发明的大单晶的可再现条件 已经建立了野生型CheY蛋白。 这些晶体 布拉格间距超过2.7 A。 一个潜在的重型 还制备了CheY晶体的原子衍生物。 在拟议的项目期间， CheY蛋白质的结构将使用 传统的多个同晶置换方法和 非晶体学对称平均技术。 的 CheY的分子结构将与现有的 基因结果来研究结构-功能关系， 形成趋化性中信号处理的基础。

The fundamental objective of this research proposal is to investigate the structure and function of a principal chemotaxis control protein from E. coli, on a molecular level. Specifically, the aim is to determine the three-dimensional molecular structure of the CheY protein by x-ray diffraction methods, and to relate the structure of its normal and mutant forms to its function in the control system of bacterial chemotaxis. The bacterial chemotaxis system is an ideal model of studying the molecular control of a cellular behavioral response. A detailed understanding of the system's organization now exists on the genetic level; to explain the control system of chemotaxis on a molecular basis, knowledge of the three-dimensional structures of its protein components is essential. The cheY gene product, because of its central role in the chemotaxis signal processing system, has been chosen as the first component to be analyzed by x-ray diffraction. An additional importance in solving the three- dimensional structure of CheY lies in its primary sequence homology with seven other proteins, all regulators in a variety of cellular functions. These homologies make CheY representative of a new type of regulatory domain for an entire class of bacterial proteins. CheY has been isolated and sequenced, and inactive point mutants have also been sequenced, and phenotypically characterized. Reproducible conditions for obtaining large, single crystals of the wild type CheY protein have been established. These crystals diffract to a Bragg spacing beyond 2.7 A. A potential heavy atom derivative of the CheY crystals has also been prepared. Within the proposed project period, the three-dimensional structure of the CheY protein will be solved using both the conventional multiple isomorphous replacement methods and the techniques of non-crystallographic symmetry averaging. The molecular structure of CheY will be used with the available genetic results to study the structure-function relationships that form the basis of signal processing in chemotaxis.