PHYSICAL STUDIES OF RNA STRUCTURAL MOTIFS

大的RNA分子和RNA-蛋白质复合物 参与基因表达的蛋白质的结构是已知的， 这些RNA具有相当大的意义。 的二级结构 几种大的RNA是很好理解的，并且正在尝试 决定了它们的整体三维折叠。 作为一种方法 这个问题，我们注意到，一些小的结构单元， 在不同的RNA中是保守的。 这些结构基序可以是 在小的合成RNA中复制， 问题研究 基序的弯曲、扭曲和氢键的测量 可以整合到一个大RNA的三维模型中。 图案的不寻常特性，例如增加的灵活性或 替代构象，可能提供了一个功能的理由， 节约 我们打算研究的RNA结构是螺旋片段， 典型和非典型碱基对;凸起;富含嘌呤的内部 循环;和伪结。 将使用几种不同的技术来 确定这些基序的结构：凝胶电泳方法 在这个实验室开发的可以测量螺旋状的扭曲 结构;旋转扩散的电双折射测量 可以精确测量RNA的端到端长度， 碱基对和弯曲角.非辐射能量传递测量 可以估计凸起或环结构的铰链状柔性; 闭合环状RNA可用于估计 凸起或环结构;熔化实验可以测量 不寻常结构的稳定性

Many ways in which large RNA molecules and RNA-protein complexes participate in gene expression are now known, and the structures of these RNA are of considerable interest. The secondary structures of several large RNAs are well-understood, and attempts are being made to determine their overall three dimensional folding. As one approach to this problem, we notice that a number of small structural units are conserved among different RNAs. These structural motifs can be reproduced in small, synthetic RNAs amenable to detailed physical studies. Measurement of bend, twist, and hydrogen bonding of a motif could be incorporated into a three dimensional model of a large RNA. Unusual properties of a motif, such as an increased flexibility or alternate conformations, might provide a functional rationale for its conservation. The RNA structures we intend to study are helical segments, with both canonical and non-canonical base pairs; bulges; purine-rich internal loops; and pseudoknots. Several different techniques will be used to determine the structures of these motifs: Gel electrophoresis methods developed in this lab can measure the twist associated with a helix-like structure; Electric birefringence measurements of rotational diffusion can accurately measure end-to-end length of RNAs, determining rise per base pair and bend angles; Non-radiative energy transfer measurements can estimate the hinge-like flexibility of a bulge or loop structure; Closed circular RNAs can be used to estimate the torsional stiffness of bulge or loop structures; Melting experiments can measure the stabilities of unusual structures.