Hfq在RNA调控中行使功能的分子机制的结构生物学研究

项目名称： Hfq在RNA调控中行使功能的分子机制的结构生物学研究

项目编号： No.31270782

项目类型： 面上项目

立项/批准年度： 2013

项目学科： 生物科学

项目作者： 龚庆国

作者单位： 中国科学技术大学

项目金额： 80万元

中文摘要： 关于RNA调控目前被广泛接受的看法是：细菌对一些环境影响如氧化应激，渗透压休克，和温度变化的反应是通过一系列非编码的sRNAs来调节的。许多sRNA是通过直接的碱基配对和它们的目标mRNA来结合的，从而进一步影响mRNA的功能。DsrA是一个87个核苷酸的sRNA，它可以在低温下与rpoS基因发生相互作用，起到提高这个mRNA翻译水平的作用。rpoS编码RNA聚合酶的sigma亚基，因此被认为是大肠杆菌对应激响应的主调节者。具有RNA分子伴侣活性的Hfq蛋白被发现在DsrA 结合rpoS过程中对RNA相互作用起着重要的调节作用。我们计划使用综合X-ray晶体衍射、溶液核磁共振和顺磁共振等多种生物物理手段来研究Hfq及其RNA底物的多元复合物结构，以及Hfq的C末端在RNA调控中扮演的角色。通过这些结构研究，我们预期可以对Hfq在RNA调控中行使功能的结构基础和分子机制有个深入的了解。

中文关键词： 晶体结构；核磁共振；Hfq；非编码RNA；应激

英文摘要： The gene rpoS encodes sigma(S) subunit of RNA polymerase which is a global regulatory factor involved in many stress responses in Escherichia coli. It has been widely accepted that a variety of noncoding small RNAs (sRNAs) allow the precise regulation of rpoS mRNA translation by a multitude of environmental factors including oxidative stress, cell surface stress, and temperature change. Many sRNAs were reported to mediate the function of rpoS via basepairing-based interaction. The DsrA sRNA is a an 87-nucleotide, untranslated RNA that regulates expression of rpoS at low temperature. The DsrA secondary structure is conserved and is predicted to form a three-stem-loop structure. The first stem-loop is involved in the anti-antisense regulation of translation of the rpoS. On the other hand, RNA champeron protein Hfq was reported to play an indispensable role in this regulation. Escherichia coli Hfq consists of a conserved N-terminal Sm domain (amino acid residues 1～65) and a flexible C-terminal tail of 37 amino acid residues. Hfq interacts with regulatory sRNA and facilitates their antisense interaction with their targets. We are planning to employ multiple biophysical tools (NMR, EPR, and X-ray crystallization) to investigate the structure of Hfq complexed with RNAs, and the precise role of Hfq C-termius in the RNA

英文关键词： crystal structure；NMR；Hfq；noncoding RNA；stress