PROTEIN-FACILITATED RNA CATALYSIS

蛋白质促进的 RNA 催化

• 批准号: 6493847
• 负责人: Kevin M Weeks
• 金额: $ 2.91万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-05-01 至 2003-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6493847
• 关键词: DNA footprinting; RNA binding protein; RNA biosynthesis; RNA splicing; covalent bond; crosslink; enzyme activity; enzyme complex; helicase; introns; nucleic acid sequence; protein structure function; ribonucleoproteins; ribozymes

DESCRIPTION (adapted from applicant's abstract): Dr. Kevin Weeks requests four years of support to elucidate fundamental features of how RNA and protein molecules collaborate to perform RNA catalysis. The proposal focuses on a mode ribonucleoprotein catalyst formed by the bI5 group I intron, a catalytic RNA, and the CBP2 protein. Catalysis provides a sensitive proof of structure-function relationships in RNA-protein interactions. This system is simple enough to be understood in significant detail, yet it is sophisticated enough to illustrate principles fundamental to more complex RNA-protein machines like those responsible for the biologically and medically important processes of RNA biogenesis, polypeptide synthesis, and chromosome maintenance Specific Aims are as follows. (1) Obtain a detailed three dimensional view of the specific bI5 structures involved in interaction with CBP2. A novel method of covalent footprinting for mapping direct RNA-protein interactions in comple RNAs will be developed. RNA sequence and structural requirements for protein-facilitated catalysis will be examined using iterative in vitro selection. This work will also address compromises involved in requiring proteins to facilitate RNA catalysis. (2) Determine specific functional roles for the two structural domains that comprise the CBP2 splicing factor and assign functions to individual domains in the context of a specific, detailed, and comprehensive kinetic framework for protein-facilitated RNA splicing. The assignment of individual protein domains to specific activities will promote understanding of the general roles of splicing factors in RNA catalysis. (3) Establish a mechanistic framework for understanding how Suv3 protein functions to disassemble the CBP2-RNA complex. Facilitated recycling of CBP2 is essentia for additional rounds of splicing on a biologically relevant time scale. These studies will define principles governing reorganization of sophisticated RNA-protein complexes by proteins whose function is coupled to NTP hydrolysis.

描述（改编自申请人的摘要）：Kevin Weeks 博士请求 四年的支持来阐明 RNA 和 蛋白质分子协作进行 RNA 催化。 提案 专注于由 bI5 I 组形成的模式核糖核蛋白催化剂 内含子、催化RNA和CBP2蛋白。 催化提供了 RNA-蛋白质结构-功能关系的灵敏证明 互动。 这个系统足够简单，可以在重要的方面被理解 细节，但它足够复杂，可以说明基本原理 更复杂的RNA-蛋白质机器，例如那些负责 RNA 生物发生的生物学和医学重要过程， 多肽合成和染色体维护具体目标如下 接下来。 (1) 获取特定bI5的详细三维视图 参与与 CBP2 相互作用的结构。 一种新的共价方法 用于绘制复杂 RNA 中直接 RNA-蛋白质相互作用的足迹将 得到开发。 RNA序列和结构要求 将使用体外迭代来检查蛋白质促进的催化作用 选择。 这项工作还将解决涉及要求的妥协 促进RNA催化的蛋白质。 (2)确定具体功能 构成 CBP2 剪接因子的两个结构域的作用 并在特定的上下文中将功能分配给各个域， 蛋白质促进 RNA 的详细、全面的动力学框架 拼接。 将各个蛋白质结构域分配给特定的 活动将促进对剪接一般作用的理解 RNA催化的因素。 （三）建立机制框架 了解 Suv3 蛋白如何发挥分解 CBP2-RNA 的作用 复杂的。 促进 CBP2 的回收是额外几轮的必要条件 在生物学相关的时间尺度上进行剪接。 这些研究将定义 复杂 RNA-蛋白质复合物重组的原则 其功能与 NTP 水解耦合的蛋白质。