PHYSICAL STUDIES OF RNA STRUCTURAL MOTIFS

大RNA分子和RNA-蛋白质复合体的许多方式 参与基因表达现在是已知的，并且结构 这些核糖核酸具有相当大的价值。的二级结构 几个大的RNA已经被很好地理解了，并且正在尝试 确定它们整体的三维折叠。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中保守。这些结构主题可以是 在小的合成RNA中复制，服从于详细的物理 学习。基序的弯曲、扭曲和氢键的测量 可以合并到一个大RNA的三维模型中。 主题的不同寻常的特性，例如增加的灵活性或 替代构象，可能为其提供功能基础 保护环境。 我们打算研究的RNA结构是螺旋片段，两者都 正则和非正则碱基对；凸起；富含嘌呤的内部 环；和伪结点。将使用几种不同的技术来 确定这些基序的结构：凝胶电泳法 在这个实验室中开发的可以测量与螺旋状相关联的扭曲 结构.旋转扩散的电双折射测量 可以准确测量RNAs的端到端长度，确定Repper 碱基对和折叠角.非辐射能量转移测量 可以估计凸起或环状结构的铰链式灵活性； 闭合环状RNA可以用来估计 凸起或环状结构；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.