The Structure and Reaction Dynamics of Three Catalytic RNAs

这个项目的主要焦点是生物大分子的功能是如何由它们的结构控制的问题。该研究计划将重点放在RNA酶（或核酶）背景下的这个问题上，希望发现生物催化的基本特性，这些特性是所有酶（无论是RNA还是蛋白质）所共有的。这项提议的总体目标是发展对生物催化剂如何发挥作用的基本理解，无论是蛋白质还是RNA。两种催化RNA系统将研究使用静态和时间分辨x射线晶体学阐明其结构性质和反应机制。这些，按照复杂性增加的顺序，包括发夹核酶和RNase p。锤头核酶的结构已经解决，时间分辨的晶体学实验已经开始。更复杂的核酶已经结晶，并将用于静态和时间分辨晶体学，以努力阐明其反应机制。本研究的主要目的是：(1)确定发夹核酶的三维结构，并利用锤头核酶的技术开始确定发夹核酶裂解和连接反应中间体的结构；(2)开始确定RNase P的三维结构。这项研究的主要目标是发展对催化原理的基本理解，使RNA分子成为酶。这一知识将有助于理解RNA酶的三维结构如何决定其催化性能。通过了解RNA酶的工作原理，该项目将发现哪些特性对生物催化剂至关重要，以至于它们对RNA和蛋白质酶都是不变的。

The primary focus of this project is upon the problem of how the functions of biological macromolecules are governed by their structures. The research plan focuses upon this problem in the context of RNA enzymes (or ribozymes), with the hope of discovering the properties of biological catalysis that are so fundamental that they are shared by all enzymes, whether composed of RNA or protein. The overall goal of this proposal is to develop a fundamental understanding of how biological catalysts function, be they proteins or RNA.Two catalytic RNA systems will be investigated using both static and time-resolved X-ray crystallography to elucidate their structural properties and reaction mechanisms. These, in order of increasing complexity, include the hairpin ribozyme, and RNase P. The hammerhead ribozyme structure has been solved and time-resolved crystallographic experiments have been initiated. The more complex ribozymes have been crystallized and will be used in static and time-resolved crystallography in an effort to elucidate their reaction mechanisms. The principle objectives of this research are: (1) to determine the three-dimensional structure of the hairpin ribozyme, and to initiate determination of the structures of reaction intermediates of hairpin ribozyme cleavage and ligation using the techniques developed for the hammerhead ribozyme; and (2) to initiate determination of the three-dimensional structure of RNase P.. The primary goal of this research is to develop a fundamental understanding of the principles of catalysis that enable RNA molecules to be enzymes. This knowledge will lead to the understanding of how the three-dimensional structure of an RNA enzyme determines its catalytic properties. By learning how RNA enzymes work, this project will discover what properties are so fundamental to biological catalysts that they are invariant with respect to both RNA and protein enzymes.