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Formation of a Ligand-Assisted Complex of Two RNA Hairpin Loops

机译：两个RNA发夹环的配体辅助复合物的形成

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The hairpin structure is one of the most common secondary structures in RNA and holds a central position in the stream of RNA folding from a non-structured RNA to structurally complex and functional ribonucleoproteins. Since the RNA secondary structure is strongly correlated to the function and can be modulated by the binding of small molecules, we have investigated the modulation of RNA folding by a ligand-assisted formation of loop-loop complexes of two RNA hairpin loops. With a ligand (NCT6), designed based on the ligand binding to the G-G mismatches in double-stranded DNA, we successfully demonstrated the formation of both inter- and intra-molecular NCT6-assisted complex of two RNA hairpin loops. NCT6 selectively bound to the two hairpin loops containing (CGG)3 in the loop region. Native polyacrylamide gel electrophoresis analysis of two doubly-labeled RNA hairpin loops clearly showed the formation of intermolecular NCT6-assisted loop-loop complex. Forster resonance energy-transfer studies of RNA constructs containing two hairpin loops, in which each hairpin was labeled with Alexa488 and Cy3 fluorophores, showed the conformational change of the RNA constructs upon binding of NCT6. These experimental data showed that NCT6 simultaneously bound to two hairpin RNAs at the loop region, and can induce the conformational change of the RNA molecule. These data strongly support that NCT6 functions as molecular glue for two hairpin RNAs.
机译：发夹结构是RNA中最常见的二级结构之一，在从非结构RNA折叠到结构复杂和功能性核糖核蛋白的RNA折叠流中占据中心位置。由于RNA二级结构与功能密切相关，并且可以通过小分子的结合进行调节，因此我们研究了配体辅助形成两个RNA发夹环的环-环复合物对RNA折叠的调控。使用基于配体结合双链DNA中G-G错配的配体（NCT6），我们成功地证明了两个RNA发夹环的分子间和分子内NCT6辅助复合物的形成。 NCT6有选择地结合到在环区域中包含（CGG）3的两个发夹环上。对两个双标记的RNA发夹环的天然聚丙烯酰胺凝胶电泳分析清楚地表明了分子间NCT6辅助环-环复合物的形成。对含有两个发夹环的RNA构建体进行Forster共振能量转移研究，其中每个发夹都被Alexa488和Cy3荧光团标记，表明RNA构筑体在NCT6结合后的构象变化。这些实验数据表明，NCT6在环区同时与两个发夹RNA结合，并可以诱导RNA分子的构象变化。这些数据强烈支持NCT6充当两个发夹RNA的分子胶。

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