Structure of the K-turn U4 RNA: a combined NMR and SANS study.
Structure of the K-turn U4 RNA: a combined NMR and SANS study.
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DOI：
10.1093/nar/gkq380
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发表时间：
2010-10
影响因子：
14.9
通讯作者：
Carlomagno T
中科院分区：
文献类型：
--
作者：
Falb M;Amata I;Gabel F;Simon B;Carlomagno T
K-turn基序是通用的RNA结构元件，在几种细胞环境中为蛋白质提供结合平台。它们的特征是磷酸骨架中的一个尖锐的扭结，使蛋白质结合RNA的两个螺旋茎成60°角。然而，到目前为止，没有高分辨率的结构的裸K-转折基序是可用的。在这里，我们提出的第一个结构调查在原子分辨率的一个未绑定的K-turn RNA（剪接体U4-Kt RNA）相结合的NMR和小角中子散射数据。通过这项研究，我们希望解决的问题，是否K-转角结构基序假设在蛋白质结合剂和二价阳离子的情况下，急剧扭结的构象。以前的研究已经解决了这个问题，通过荧光共振能量转移，生化分析和分子动力学模拟，这表明K-turn RNA存在于一个扭结的构象，这是有能力的蛋白质结合，和一个更扩展的构象之间的平衡，与人口分布取决于二价阳离子的浓度。我们的数据表明，U4-Kt RNA主要假设在蛋白质和二价阳离子的情况下，更扩展的构象。内环区域结构良好，但采用与蛋白质复合物中观察到的不同构象。我们的数据表明，K-转角共有序列本身不编码扭结构象;相反，尖锐的骨架扭结需要通过蛋白质结合剂来稳定。
K-turn motifs are universal RNA structural elements providing a binding platform for proteins in several cellular contexts. Their characteristic is a sharp kink in the phosphate backbone that puts the two helical stems of the protein-bound RNA at an angle of 60°. However, to date no high-resolution structure of a naked K-turn motif is available. Here, we present the first structural investigation at atomic resolution of an unbound K-turn RNA (the spliceosomal U4-Kt RNA) by a combination of NMR and small-angle neutron scattering data. With this study, we wish to address the question whether the K-turn structural motif assumes the sharply kinked conformation in the absence of protein binders and divalent cations. Previous studies have addressed this question by fluorescence resonance energy transfer, biochemical assays and molecular dynamics simulations, suggesting that the K-turn RNAs exist in equilibrium between a kinked conformation, which is competent for protein binding, and a more extended conformation, with the population distribution depending on the concentration of divalent cations. Our data shows that the U4-Kt RNA predominantly assumes the more extended conformation in the absence of proteins and divalent cations. The internal loop region is well structured but adopts a different conformation from the one observed in complex with proteins. Our data suggests that the K-turn consensus sequence does not per se code for the kinked conformation; instead the sharp backbone kink requires to be stabilized by protein binders.
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