Structure/Function of Ribonuclease P

核糖核酸酶 P 的结构/功能

• 批准号: 6694050
• 负责人: MICHAEL E. HARRIS
• 金额: $ 33.66万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-01-01 至 2006-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6694050
• 关键词: Structure; Function; Ribonuclease

DESCRIPTION (provided by applicant): It is well established that enzymes carry out the chemical reactions that are essential to human cell function. We now know that several key enzymes, including the ribosome that carries out protein synthesis, contain essential RNA molecules that participate directly in catalysis. The long-term goal of our research program is to understand how an enzyme active site composed of RNA achieves specificity and chemical rate enhancement comparable to protein enzymes. To pursue this objective we have employed the catalytic RNA, or ribozyme, derived from bacterial ribonuclease P. This ribozyme is highly conserved and homologous to the enzyme found in humans. Thus, it provides an excellent experimental system for exploring fundamental aspects of RNA structure and function. RNase P uses metal ions to promote the attack of a water molecule to break a specific phosphodiester linkage in an RNA precursor to generate a functional tRNA molecule. The proposed research is aimed at providing an understanding of enzyme mechanism, the specific role of metal ions in catalysis, and a functional description of how non-covalent interactions contribute to specificity and rate enhancement. Insight into catalytic mechanism will be gained by examining the effect of substituting heavy isotopes for atoms involved in the chemical reaction. We will begin with an analysis or the attacking water and, in the long term, extend these studies to encompass additional atoms involved in the reaction. We will identify sites of catalytic metal ion binding by examining the effects of substituting specific atoms within the active site of the RNA on reaction rate and metal ion dependence. The role of non-covalent binding interactions will be explored by testing specific models for interactions by making compensatory mutations on the ribozyme and its substrate. Additionally, because enzymes conformation of is generally dynamic, we will extend these studies to determine how changes in conformation are linked to specificity and catalysis. The issues of mechanism addressed are common to all enzymes, thus we expect that the information gained will add to our understanding of biological catalysis. Furthermore, ribozymes, including RNase P, have been adapted to therapeutic purposes, and the results we will obtain should assist in applying catalytic RNAs to this goal.

描述（由申请人提供）：众所周知，酶执行对人类细胞功能必不可少的化学反应。我们现在知道，几种关键酶，包括进行蛋白质合成的核糖体，含有直接参与催化的必需RNA分子。我们的研究计划的长期目标是了解由RNA组成的酶活性位点如何实现与蛋白质酶相当的特异性和化学速率提高。为了实现这一目标，我们采用了源自细菌核糖核酸酶的催化​​RNA或核酶。该核酶高度保守，并且与人类发现的酶同源。因此，它为探索RNA结构和功能的基本方面提供了出色的实验系统。 RNase P使用金属离子来促进水分子的攻击，以打破RNA前体中特定的磷酸二键连接以产生功能性tRNA分子。拟议的研究旨在提供对酶机制的理解，金属离子在催化中的特定作用以及对非共价相互作用如何促进特异性和速率增强的功能描述。通过检查将重型同位素取代化学反应的原子的效果，可以获得对催化机制的见解。我们将从分析或攻击水开始，从长远来看，将这些研究扩展到涉及反应的其他原子。我们将通过检查RNA活跃位点内的特定原子对反应速率和金属离子依赖性的替代特定原子的影响来识别催化金属离子结合的位点。非共价结合相互作用的作用将通过在核酶及其底物上进行补偿性突变来测试特定模型来探索。此外，由于酶的构象通常是动态的，因此我们将扩展这些研究，以确定构象变化如何与特异性和催化有关。解决的机制问题对于所有酶都是常见的，因此我们希望获得的信息将增加我们对生物催化的理解。此外，包括RNase P在内的核酶已适应治疗目的，我们将获得的结果应有助于将催化RNA应用于此目标。