STRUCTURE DETERMINATION OF AN INTACT BACTERIAL

本子项目是利用由NIH/NCRR资助的中心赠款提供的资源的众多研究子项目之一。子项目和研究者（PI）可能已经从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。列出的机构是中心的，不一定是研究者的机构。天然存在的催化rna或核酶有几类。了解这类生物催化剂如何促进其反应的重要性随着最近对50S核糖体的结构测定而变得明显，这表明所有生物体内的蛋白质合成都发生在RNA活性位点内。核酶是最有可能成为现代生物催化剂祖先的分子，了解它们如何促进它们的反应提供了对酶学功能的关键见解。本项目重点研究I族自剪接内含子的结构，这是第一类发现的催化rna。结构信息对于理解和解释这类生物催化剂的生物学功能是必要的。我们正朝着三个目标努力。1. 本项目的主要目的是确定完整细菌I族自剪接内含子的高分辨率x射线晶体结构。如果达到这一目标，将进行另外两个项目：重金属离子和替代内含子抑制剂将用于从结构上研究金属离子如何参与I族内含子折叠和催化。3. 磷转移反应的过渡态将在结构上探索使用稳定的模拟过渡态几何。这些研究将为20多年的生物化学研究提供结构基础。它还将提供对RNA催化性质的化学见解，特别是RNA金属酶，其中一个尚未进行结构分析的例子。鉴于RNA在包括肽键形成和mrna前剪接在内的几个自然过程中的关键作用，RNA催化是一个特别感兴趣的问题。这些坐标将提供给科学界进行分析和实验设计。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. There are several classes of naturally occurring catalytic RNAs or ribozymes. The importance of understanding how this class of biocatalysts promotes their reactions became evident with the recent structure determination of the 50S ribosome, which demonstrated that protein synthesis in all living organisms occurs within an RNA active site. Ribozymes are the molecules most likely to be the progenitors of modern biological catalysts and understanding how they promote their reactions provides critical insight into enzymological function. This project focuses on the structure of the group I self-splicing intron, the first class of catalytic RNAs discovered. Structural information is necessary to understand and interpret the biological function of this class of biocatalyst. We are working toward three goals. 1. The primary aim of this project is to determine the high resolution x-ray crystal structure of an intact bacterial group I self-splicing intron. If this aim is achieved, two additional projects will be undertaken: 2. Heavy metal ions and alternative intron inhibitors will be used to structurally investigate how metal ions contribute to group I intron folding and catalysis. 3. The transition state of the phosphotransfer reaction will be structurally explored using stable mimics of the transition state geometry. These studies will provide a structural basis for more than 20 years of biochemistry. It will also provide chemical insights into the nature of RNA catalysis, particularly for an RNA metalloenzyme, an example of which has not yet been structurally analyzed. RNA catalysis is an issue of particular interest given the critical of role of RNA in several natural processes including peptide bond formation and pre-mRNA splicing. The coordinates will be made available to the scientific community for analysis and experimental design.