Structure and Function of Group II Intron Ribozyme

描述（由申请人提供）：II族内含子是具有多种生物学功能的核酶：它们催化前体RNA的剪接并转位到新的基因组位置。至少三分之一的人类基因组可能起源于祖先II组内含子，它们可能是真核内含子、剪接体和主要类型的反转录转座子的进化前身。II类内含子在现代生物的代谢和进化中继续发挥着重要作用，由于它们能够通过转位反应改变特定的DNA和RNA序列，因此它们是生物技术和分子医学的有前途的工具。在更基本的层面上，II组内含子具有不同寻常的结构，提供了关于RNA折叠、三级结构和RNA-蛋白质相互作用的丰富信息。为了将II族内含子应用于生物医学和基础研究，并了解真核RNA剪接的分子机制，有必要对它们的三维结构进行可视化。为此，我们提出了一系列的实验来阐明在内含子折叠和剪接的多个阶段II族内含子的高分辨率结构。具体来说，我们正在使用晶体学和生化方法来建立内含子的折叠结构状态，并可视化活性位点的分子细节。这些研究是针对分离的内含子rna和II组内含子及其天然伴侣蛋白的复合物进行的。该项目旨在对RNA分子折叠成三级结构、催化化学反应和与蛋白质合作以进行细胞基本代谢的机制产生基本的新见解。II组内含子的高分辨率结构信息将帮助我们理解并潜在地操纵控制基因表达模式的RNA剪接反应的分子机制。它还将促进二类内含子作为生物技术和基因治疗应用工具的发展。

DESCRIPTION (provided by applicant): Group II introns are ribozymes with a diversity of biological functions: They catalyze their own splicing from precursor RNA and transposition into new genomic locations. At least one-third of the human genome may have originally derived from ancestral group II introns, which are the likely evolutionary predecessors of eukaryotic introns, the spliceosome and major classes of retrotransposons. Group II introns continue to play a major role in the metabolism and evolution of modern organisms and, given their ability to alter specific DNA and RNA sequences through transposition reactions, they are promising tools for biotechnology and molecular medicine. At a more basic level, group II introns have an unusual architecture that is providing a wealth of information on RNA folding, tertiary structure, and RNA-protein interactions. In order to apply group II introns in biomedical and basic research, and to understand molecular mechanisms for eukaryotic RNA splicing, it is essential that we visualize their three-dimensional structures. To this end, we propose a series of experiments to elucidate the high-resolution structures of group II introns during the multiple stages of intron folding and splicing. Specifically, we are employing crystallographic and biochemical methods to establish the folded architectural states of the intron and to visualize molecular details of the active-site. These studies are being conducted on isolated intron RNAs and on group II introns in complex with their natural partner proteins. This project is designed to yield fundamental new insights into the mechanisms by which RNA molecules fold into tertiary structures, catalyze chemical reactions and collaborate with proteins in order to carry out the basic metabolism of the cell. High-resolution structural information on group II introns will help us to understand, and potentially manipulate, molecular mechanisms for the RNA splicing reactions that control our patterns of gene expression. It will also facilitate the development of group II introns as tools for application in biotechnology and gene therapy.