NMR SOLUTION STRUCTURES OF RNA

RNA可以催化化学反应的发现导致了 确定了许多新的生物学和生物化学作用的RNA， 细胞 然而，尽管作出了重大努力，目前几乎没有 RNA的三维结构。 这种情况 与蛋白质相反，蛋白质的X射线或NMR结构中有数百个 生物活性酶是已知的，并且这些数据被常规地使用 来帮助解开这些分子的分子作用机制。 在这个提议中，最先进的异相多维NMR 光谱学将用于探测结构和动力学的 各种生物学上重要的RNA和RNA结构基序。 部分 该项目需要开发和优化异质结 用于共振归属和结构的2D、3D和4D NMR技术 RNA的测定。 类似的技术已经彻底改变了 蛋白质的溶液结构测定，但它们还没有 应用于核酸。 方法的最新进展 产生毫克量的13 C和/或15 N标记的RNA， 现在可以进行更详细的结构测定， 更大的RNA 该项目的一个目标是确定和 RNA折叠基序的结构测定。 以下系统 将研究：i）RNA发夹属于经常发生的 GNRA（其中N是任何核苷酸并且R是A或G）和CUUG四环 ii）RNA三链体;和iii）天然存在的RNA双链体 含有内部环或“错配”碱基对。 研究 将为我们提供大量关于RNA构建模块的新信息 三级结构 研究RNA结构基序的重要性 在生物学功能中，由一个片段组成的蛋白质-RNA复合物 含有信号识别颗粒（SRP）RNA及其 位点特异性结合蛋白，SRP-19蛋白，将被研究， 匪r 该RNA结合位点含有GNRR四环，并且该系统 将用于确定这种19 kD蛋白的结合如何影响 RNA发夹的结构 这些结果将导致更好的 理解RNA结构基序在蛋白质-RNA中的作用 识别.

The discovery that RNA can catalyze chemical reactions has led to the identification of many new biological and biochemical roles for RNAs in cells. However despite significant effort, there is presently little information on the three-dimensional structures of RNA. This situation is in contrast to proteins where X-ray or NMR structures of hundreds of biologically active enzymes are known and these data are routinely used to help unravel the molecular mechanisms of action of these molecules. In this proposal state-of-the-art heteronuclear multi-dimensional NMR spectroscopy will be used to probe the structure and dynamics of a variety of biologically important RNAs and RNA structural motifs. Part of this project entails development and optimization of heteronuclear 2D, 3D and 4D NMR techniques for resonance assignment and structure determinations of RNAs. Similar techniques have revolutionized the solution structure determinations of proteins but they have yet to be applied to nucleic acids. Recent advances in the methodology for generating milligram quantities of 13C and/or 15N labelled RNAs, will now make it possible to perform more detailed structure determination of larger RNAs. One goal of this project focuses on identification and structure determination of RNA folding motifs. The following systems will be studied: i) RNA hairpins belonging to the frequently occurring GNRA (where N is any nucleotide and R is A or G) and CUUG tetraloop families; ii) RNA triplexes; and iii) naturally occurring RNA duplexes that contain internal loops or 'mismatched' base pairs. The studies will give us a wealth of new information on the building blocks of RNA tertiary structure. To investigate importance of RNA structural motifs in biological functions a protein-RNA complex consisting of a fragment of the signal recognition particle (SRP) RNA containing and its site-specific binding protein, the SRP-19 protein, will be studied by NMR. This RNA binding site contains a GNRR tetraloop and this system will be used to determine how the binding of this 19kD protein affects the structure of the RNA hairpin. These result will lead to a better understanding of the role of RNA structural motifs in protein-RNA recognition.