STRUCTURE-FUNCTION RELATIONSHIPS OF RNA POLYMERASES

RNA聚合酶的结构-功能关系

• 批准号: 3300436
• 负责人: BI-CHENG WANG
• 金额: $ 21.79万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-05-01 至 1994-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3300436
• 关键词: DNA; DNA directed RNA polymerase; X ray crystallography; bacteriophage T7; crystallization; electron density; enzyme structure; genetic promoter element; genetic transcription; nucleoside triphosphate; oligonucleotides; protein structure function

The objective of this research is to provide, through crystallographic studies, information which will help to understand the structure-function relationship of RNA polymerase at the atomic level. A large amount of T7 RNA polymerase has recently been isolated from an overexpression system, and experimental conditions have been established under which diffraction quality single crystals can be grown. The crystals, which belong to space group P21 with cell constants a = 114.5, b = 138.6, c = 125.7 angstrom, and beta = 98.1 degree, grow to a size of 1.5 x 0.2 x 0.1 mm, and diffract to at least 3.5 angstrom resolution. A systematic crystallographic study is proposed on a series of RNA polymerases and their DNA complexes. Specific aims are as follows: (1) to determine the crystal structure of the T7 RNA polymerase crystals presently in hand, and to extend the diffraction resolution of these crystals and analyze their structural details, using synchrotron radiation data. (2) to grow and study the T7 RNA polymerase-DNA co-crystals containing the class II and class III T7 promoters and nucleoside triphosphates. (3) to grow native crystals of bacteriophage T3 RNA polymerase, and its DNA complexes, with the aim of solving their crystal structures. Success in obtaining diffraction quality single crystals of T7 RNA polymerase represents the first significant step toward a complete crystallographic study of the enzyme. The proposed research is expected to provide new information and knowledge necessary for the understanding of the structure-function relationship of RNA polymerase and its involvement in DNA transcription.

这项研究的目的是通过 晶体学研究，将有助于理解的信息 RNA聚合酶在原子上的结构功能关系 等级。 最近已经有大量的T7 RNA聚合酶 从过表达系统和实验条件中分离出来 已经确定了哪个衍射质量单。 可以种植晶体。 属于太空群的晶体 P21具有单元常数A = 114.5，B = 138.6，C = 125.7 Angstrom， β= 98.1度，生长到1.5 x 0.2 x 0.1 mm， 并至少衍射为3.5 Angstrom分辨率。 提出了一系列RNA的系统晶体学研究 聚合酶及其DNA复合物。 具体目的如下： （1）确定T7 RNA聚合酶的晶体结构 晶体目前握在手中，并延长衍射 分辨这些晶体并分析它们的结构细节， 使用同步加速器辐射数据。 （2）生长和研究T7 RNA聚合酶-DNA共晶 包含II类和III类T7启动子和核苷 三磷酸盐。 （3）生长噬菌体T3 RNA聚合酶的天然晶体， 及其DNA复合物，目的是解决其晶体 结构。 获得T7 RNA的衍射质量单晶的成功 聚合酶代表了迈向完整的第一步 酶的晶体学研究。 拟议的研究是 期望提供新的信息和知识 理解RNA的结构功能关系 聚合酶及其参与DNA转录。