光镊技术研究rpoS RNA非编码区局域结构的稳定性

It is well known that the function of no-coding RNA is defined by its structure. However, up to date, it is difficult to understand RNA 3-D conformation of molecules longer than 1000 nt and different software often give inconsistent prediction..Recently, we found the conformation of rpoS RNA untranslated region is very diverse under AFM but contain stem loops with similar shape. Stretching experiment done by optical tweezers revealed the unzipping force of self-inhibition stem loop is significant bigger than normal RNA hairpin..Therefore, we hypothesize that the no-coding region of rpoS mRNA may be constituted by multiple stem loops with strong internal interaction. Stem loops may connect together diversely. In this proposal, we plan to measure unzipping force, length and free energy of different stem loops, study their dynamic with optical tweezers. Thus, we will able to investigate the stability of each potential unit among rpoS mRNA untranslated region, relate these characteristics with mechanism of rpoS gene regulation and test our hypothesis. .We believe this study will facilitate prediction of RNA conformation by providing a new quantitative method. This work will be very meaningful for people to understand the mechanism of RNA folding and their biological functions.

RNA的生物学功能与其结构密切相关。但是对于长度超过1000nt的RNA三维结构了解非常有限，而且不同结构预测软件给出的结果也不一致。长链RNA结构研究是结构生物学的难点。.我们通过原子力显微镜发现rpoS信使RNA非编码区的卷曲方式多样，但存在一些外形相似的小茎环；光镊实验也发现其自抑制茎环内部结合强度大于一般RNA发卡结构的结合强度。.由此我们提出rpoS信使RNA非编码区域可能由一些内部结合力较强的片段组成，各片段再以不同形态相连接。我们拟通过光镊拉伸RNA非编码区不同的片段或茎环结构，研究打开的动力学过程，测量打开所需要的外力、长度、自由能等物理参数，讨论各个片段的稳定性以及其与该RNA生物学功能之间的关系，验证假说。.研究结果将提供一种辅助长链RNA空间结构预测新的定量研究方法，用于帮助人们理解RNA形成三维结构的动态过程以及其结构与生物学功能之间的关系，具有重要的生物学意义。