Automated physics-based design of synthetic riboswitches from diverse RNA aptamers.
Automated physics-based design of synthetic riboswitches from diverse RNA aptamers.
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DOI：
10.1093/nar/gkv1289
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发表时间：
2016-01-08
影响因子：
14.9
通讯作者：
Salis HM
中科院分区：
文献类型：
--
作者：
Espah Borujeni A;Mishler DM;Wang J;Huso W;Salis HM
核糖开关是形状改变的调节RNA，其结合化学物质并调节基因表达，直接将传感耦合到细胞致动。然而，目前还不清楚它们的序列如何控制核糖开关开关和激活的物理过程，特别是当改变配体结合适体结构域时。我们报告了一个统计热力学模型的发展，预测序列-结构-功能的关系，激活基因表达，其特征在于细胞内和细胞内的无细胞转录-翻译测定的抑制调节核糖开关。使用该模型，我们进行了62个合成核糖开关的自动计算设计，这些核糖开关使用六种不同的RNA适体来检测不同的化学物质（茶碱，四甲基玫瑰胺，氟化物，多巴胺，甲状腺素，2，4-二硝基甲苯），并激活基因表达高达383倍。该模型解释了如何适体结构，配体亲和力，开关自由能和大分子拥挤共同控制核糖开关激活。我们的基于模型的方法工程核糖开关定量证实了几个物理机制，配体诱导的RNA形状变化，并使无细胞和细菌传感器的发展，为不同的应用。
Riboswitches are shape-changing regulatory RNAs that bind chemicals and regulate gene expression, directly coupling sensing to cellular actuation. However, it remains unclear how their sequence controls the physics of riboswitch switching and activation, particularly when changing the ligand-binding aptamer domain. We report the development of a statistical thermodynamic model that predicts the sequence-structure-function relationship for translation-regulating riboswitches that activate gene expression, characterized inside cells and within cell-free transcription–translation assays. Using the model, we carried out automated computational design of 62 synthetic riboswitches that used six different RNA aptamers to sense diverse chemicals (theophylline, tetramethylrosamine, fluoride, dopamine, thyroxine, 2,4-dinitrotoluene) and activated gene expression by up to 383-fold. The model explains how aptamer structure, ligand affinity, switching free energy and macromolecular crowding collectively control riboswitch activation. Our model-based approach for engineering riboswitches quantitatively confirms several physical mechanisms governing ligand-induced RNA shape-change and enables the development of cell-free and bacterial sensors for diverse applications.