BIOPHYSICAL CHEMISTRY OF A DEAD/H PROTEIN

描述：（来自申请人的摘要）本项目的目标是 了解E.大肠杆菌DbpA，一个大的成员 DEAD/H蛋白家族参与许多细胞途径，包括 核糖核酸DEAD/H蛋白将ATP水解与RNA结合偶联， 提出修改RNA二级和三级结构。E.大肠杆菌DbpA及其 B。选择枯草杆菌同源物YxiN进行详细的结构和机制分析， 因为与几乎所有其他DEAD/H蛋白不同，它们紧密结合， 特别是23 S rRNA的离散区域。均衡和速率 ATP酶反应最小动力学方案中各步骤的常数 将决心建立一个结构-功能研究的框架。的 DbpA在重组rRNA中作为解旋酶的可能性将是 考察RNA修饰、光交联和蛋白质工程 实验将测试DbpA的不同结构域如何与其同源物相互作用 RNA，以及蛋白质-RNA接触是否在催化循环期间发生变化。 最后，获得了一株DbpA破坏菌E.大肠杆菌将用于搜索 DbpA在E. coli细胞。

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.