BIOPHYSICAL CHEMISTRY OF A DEAD/H PROTEIN

死/H 蛋白的生物物理化学

• 批准号: 6387041
• 负责人: OLKE C UHLENBECK
• 金额: $ 21.84万
• 依托单位: UNIVERSITY OF COLORADO
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-06-01 至 2004-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6387041
• 关键词: Bacillus subtilis; Escherichia coli; RNA binding protein; adenosinetriphosphatase; bacterial proteins; chemical kinetics; crosslink; enzyme activity; helicase; photochemistry; protein engineering; protein structure function; ribosomal RNA; thermodynamics

DESCRIPTION: (From the Applicant's Abstract) The goal of this project is to obtain an understanding of the function of E. coli DbpA, a member of the large DEAD/H family of proteins that participate in many cellular pathways involving RNA. DEAD/H proteins couple the hydrolysis of ATP with RNA binding and are proposed to modify RNA secondary and tertiary structure. E. Coli DbpA and its B. subtilis homologue YxiN were chosen for detailed structural and mechanistic studies because, unlike nearly all other DEAD/H proteins, they bind tightly and specifically to a discrete region of 23S rRNA. The equilibrium and rate constants for the steps in the minimal kinetic scheme of the ATPase reaction will be determined to establish a framework for structure-function studies. The possibility that DbpA acts as a helicase in restructuring rRNA will be examined. RNA modification, photocrosslinking, and protein engineering experiments will test how different domains of DbpA interact with its cognate RNA, and whether the protein-RNA contacts change during the catalytic cycle. Finally, a DbpA disruption strain of E. coli will be used to search for the mechanism of action of DbpA in E. coli cells.

描述：（从申请人的摘要中）该项目的目标是 了解大肠杆菌DBPA的功能，该功能是大型成员 参与许多涉及的许多细胞途径的死亡/h蛋白家族 RNA。死亡/h蛋白将ATP的水解与RNA结合，为 提议修改RNA次级和三级结构。大肠杆菌DBPA及其 枯草芽孢杆菌同源物YXIN被选择用于详细的结构和机理 研究是因为，与几乎所有其他死亡/H蛋白不同，它们都紧密结合，并且 专门针对23S rRNA的离散区域。平衡和速率 ATPase反应最小动力学方案中步骤的常数 将确定为结构功能研究建立框架。这 DBPA在重组rRNA中充当解旋酶的可能性将是 检查。 RNA修饰，光叠链接和蛋白质工程 实验将测试不同的DBPA域与其同源的相互作用 RNA，以及蛋白-RNA接触在催化循环中是否发生变化。 最后，将使用大肠杆菌的DBPA破坏菌株搜索 DBPA在大肠杆菌细胞中的作用机理。