STRUCTURE AND CATALYSIS OF AN RNA ENZYME

描述：这里提出的研究的全球目标是 理解一个基本的问题，即一种称为 锤头状核酶有效。结合使用传统和 时间分辨X射线结晶学实验与合成 修改的RNA，最近阐明的基态的假设 锤头状核酶的结构与催化相关 测试过。在这样做的过程中，对催化之间的关系的理解 通过对结构的研究将获得RNA的结构和功能 锤头状核酶在RNA上的不同重要位置 裂解反应坐标。中描述的计划的具体目标 本研究的建议是：(1)捕捉晶体结构 锤头状RNA裂解中间体；(2)获得晶体结构 切割后的锤头RNA的结构；(3)捕获 裂解锤头状核酶连接过程中的限速物种 底物；(4)获得锤头RNA裂解反应的结构 过渡态模拟；(5)理解的结构基础 催化增强锤头状RNA修饰；以及(6)选择 与锤头RNA结合并锁定的蛋白质 活性构象，并确定该化合物的晶体结构 RNA-蛋白质复合体。这六个具体目标都是为了探索， 独立地，这种小RNA酶的切割机制。 锤头状核酶作为靶向治疗药物的潜在用途 RNA病毒(如HIV)和病理性mRNAs(如癌基因 成绩单)是目前医学领域中一个活跃的研究领域 社区。锤头状核酶作为潜在的药物 特别感兴趣，因为它们可能被证明对治疗疾病有用 事实证明，这些问题在其他方面都很棘手。这次袭击的主要动机之一是 在这里提出的研究，除了回答一个基本的问题 科学性，就是向科学和技术人员提供实用信息 医学界将使更有效的基于核糖体的药物 被设计好。

DESCRIPTION: The global objective of the research proposed here is to understand the fundamental question of how a small RNA enzyme, called the hammerhead ribozyme, works. Using a combination of conventional and time-resolved X-ray crystallography experiments, together with synthetically modified RNA, the hypothesis that the recently elucidated ground-state structure of the hammerhead ribozyme is catalytically relevant will be tested. In doing so, an understanding of the relationship between catalytic RNA structure and function will be obtained by investigating the structures of the hammerhead ribozyme at various important points along the RNA cleavage reaction coordinate. The specific aims of the program described in this research proposal are: (1) to capture the crystal structures of hammerhead RNA cleavage intermediates; (2) to obtain the crystal structure of the hammerhead RNA after cleavage; (3) to capture structures of rate-limiting species during ligation of the cleaved hammerhead ribozyme substrate; (4) to obtain the structure of a hammerhead RNA cleavage reaction transition-state analog; (5) to understand the structural basis of catalysis-enhancing hammerhead RNA modifications; and (6) to select for a protein that binds to, and locks, the hammerhead RNA in a catalytically active conformation, and to determine the crystal structure of this RNA-protein complex. These six specific aims are each designed to probe, independently, the cleavage mechanism of this small RNA enzyme. The potential use of hammerhead ribozymes as therapeutic agents that target RNA viruses (such as HIV) and pathological mRNAs (such as oncogene transcripts) is currently an active area of research in the medical community. Hammerhead ribozymes as potential pharmaceuticals are of particular interest because they may prove usful for treating conditions that have proven otherwise intractable. One of the primary motives for the research proposed here, in addition to answering questions of a basic scientific nature, is to provide practical information to the scientific and medical communities to enable more potent ribosome-based pharmaceuticals to be designed.