RNA Dynamics by Solution and Solid State Deuterium NMR

本项目的目的是通过将溶液和固态核磁共振联合应用于范例系统HIV-1 TAR来研究核酸动力学。通过一个共同的理论框架，分析两种核磁共振方法获得的实验结果，将比迄今为止可能更全面地描述RNA动力学。两种光谱的同时使用将允许更广泛的时间尺度和地点范围，而不是单独使用任何一种技术，并且可以比较溶液和固体中的RNA动力学。该项目的核心假设是运动是核酸功能的一个重要方面，HIV TAR提供了这一前提的有力证明。其结构和分子相互作用已被广泛研究，但Tat蛋白识别TAR的分子基础尚不清楚，因为蛋白质和RNA中存在复杂的构象动力学。溶液核磁共振和固态核磁共振都是研究动力学的有力方法，但都有局限性。溶液弛豫数据对以一定速率发生的运动很不敏感，而固态氘线形分析需要单个氘化样品。因此，有必要开发新的方法来收集和分析实验结果，以定量详细地了解运动如何有助于核酸中的分子识别。通过相同的理论框架分析两组光谱结果，将有可能从定性和定量上获得TAR RNA内运动的本质的新见解，这些运动是Tat蛋白识别的基础，因此它在基因调控中的功能。本课题开发的研究核酸功能动力学的新工具将广泛适用于其他核酸甚至蛋白质。这个项目是高度跨学科的，跨越了物理化学、分子生物学、生物化学、分子药理学和分子病毒学等领域。因此，它为科学合作提供了许多机会，并使本科生和研究生接触跨学科研究。它为培养未来发展和应用核磁共振方法研究生物系统的领导者提供了肥沃的土壤。PI研究小组的一个重要方面是大多数女学生和一些代表性不足的少数民族的存在。每年夏天，其中一名pi通过华盛顿大学基因组和STAR项目监督一名来自弱势群体的高中生，该项目旨在增加招募弱势群体学生从事科学研究

The objective of this project is to investigate nucleic acids dynamics by applying solution and solid-state NMR conjointly to a paradigmatic system, the HIV-1 TAR. RNA dynamics will be described more comprehensively than it has been possible so far by analyzing the experimental results obtained by the two NMR methods through a common theoretical framework. The concomitant use of the two spectroscopies will allow a wider range of time scales and sites than would be possible by using either technique in isolation, and to compare dynamics of RNA in solution and the solid. The central hypothesis underlying this project is that motion is an important aspect of nucleic acid function and HIV TAR provides an eloquent demonstration of this premise. Its structure and molecular interactions have been studied extensively, yet the molecular basis for TAR recognition by Tat protein is poorly understood because of complex conformational dynamics in the protein and RNA. Both solution and solid state NMR are powerful methods to study dynamics, but each has limitations. Solution relaxation data are poorly sensitive to motions that occur at certain rates, while solid-state deuterium lineshape analysis requires singly deuterated samples. Therefore, it is necessary to develop new methods to collect and analyze the experimental results to understand in quantitative detail how motion contributes to molecular recognition in nucleic acids. By analyzing the two set of spectroscopic results through the same theoretical framework, it will be possible to gain qualitatively and quantitatively new insight into the nature of the motions within TAR RNA that underlie its recognition by Tat protein and therefore its function in gene regulation. The new tools to study functional dynamics in nucleic acids developed in this project will be broadly applicable to other nucleic acid and even to proteins.This project is highly interdisciplinary, crossing the areas of physical chemistry, molecular biology, biochemistry, molecular pharmacology and molecular virology. Therefore, it provides many opportunities for scientific collaborations and for exposing undergraduate and graduate students to interdisciplinary research. It provides fertile grounds for training future leaders in the development and application of NMR methods to study biological systems. An important aspect of the PI's research group is the presence of a majority of female students and a number of under-represented minorities. One of the PIs supervises every summer a high school student from an under-represented minority group through the University of Washington Genome and STAR programs aimed at increasing recruitment of under-represented minority students to the science