Allosteric mechanism of the V. vulnificus adenine riboswitch resolved by four-dimensional chemical mapping.
Allosteric mechanism of the V. vulnificus adenine riboswitch resolved by four-dimensional chemical mapping.
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作者：
Tian S;Kladwang W;Das R
介导RNA分子的基因调控功能的结构相互转换可能不同于变构的经典模型，但即使在深入研究的系统中，相关的结构相关性仍然难以捉摸。在这里，我们提出了一个四维扩展的化学映射称为锁定突变地图救援（LM 2 R），它集成了多个层次的突变与核苷酸分辨率的化学映射。该技术解决了腺嘌呤响应性创伤弧菌添加核糖开关的核心机制，这是一个范式系统，已经提出了Monod-Wyman-Changeux（MWC）构象选择模型和非MWC替代方案。为了区分这些模型，我们通过设计的突变锁定了每个功能重要的螺旋，并通过化学作图评估了补偿性拯救来评估其他螺旋的形成或消耗。这些LM 2 R测量结果强烈支持MWC模型预测的预先存在的相关性，不赞成替代模型，并表明可能在无配体核糖开关中普遍存在的额外结构异质性。
The structural interconversions that mediate the gene regulatory functions of RNA molecules may be different from classic models of allostery, but the relevant structural correlations have remained elusive in even intensively studied systems. Here, we present a four-dimensional expansion of chemical mapping called lock-mutate-map-rescue (LM2R), which integrates multiple layers of mutation with nucleotide-resolution chemical mapping. This technique resolves the core mechanism of the adenine-responsive V. vulnificus add riboswitch, a paradigmatic system for which both Monod-Wyman-Changeux (MWC) conformational selection models and non-MWC alternatives have been proposed. To discriminate amongst these models, we locked each functionally important helix through designed mutations and assessed formation or depletion of other helices via compensatory rescue evaluated by chemical mapping. These LM2R measurements give strong support to the pre-existing correlations predicted by MWC models, disfavor alternative models, and suggest additional structural heterogeneities that may be general across ligand-free riboswitches.