Bioinformatics, Institute of Computer Science, and Interdisciplinary Center for Bioinformatics, Leipzig University, Härtelstraße 16-18, 04107 Leipzig, Germany; University of Vienna, Faculty of Computer Science, Research Group Bioinformatics and Computational Biology, Währingerstraße 29, 1090 Vienna, Austria; University of Vienna, Faculty of Chemistry, Department of Theoretical Chemistry, Währingerstraße 17, 1090 Vienna, Austria.
Bioinformatics, Institute of Computer Science, and Interdisciplinary Center for Bioinformatics, Leipzig University, Härtelstraße 16-18, 04107 Leipzig, Germany; University of Vienna, Faculty of Computer Science, Research Group Bioinformatics and Computational Biology, Währingerstraße 29, 1090 Vienna, Austria; University of Vienna, Faculty of Chemistry, Department of Theoretical Chemistry, Währingerstraße 17, 1090 Vienna, Austria.
Methods. 2018 Jul 1;143:90-101. doi: 10.1016/j.ymeth.2018.04.003. Epub 2018 Apr 13.
This contribution sketches a work flow to design an RNA switch that is able to adapt two structural conformations in a ligand-dependent way. A well characterized RNA aptamer, i.e., knowing its K and adaptive structural features, is an essential ingredient of the described design process. We exemplify the principles using the well-known theophylline aptamer throughout this work. The aptamer in its ligand-binding competent structure represents one structural conformation of the switch while an alternative fold that disrupts the binding-competent structure forms the other conformation. To keep it simple we do not incorporate any regulatory mechanism to control transcription or translation. We elucidate a commonly used design process by explicitly dissecting and explaining the necessary steps in detail. We developed a novel objective function which specifies the mechanistics of this simple, ligand-triggered riboswitch and describe an extensive in silico analysis pipeline to evaluate important kinetic properties of the designed sequences. This protocol and the developed software can be easily extended or adapted to fit novel design scenarios and thus can serve as a template for future needs.
本研究概述了一种设计 RNA 开关的工作流程，该开关能够以配体依赖的方式适应两种结构构象。一个特征良好的 RNA 适体（即，了解其 K 值和适应性结构特征）是该设计过程的必要组成部分。在整个工作中，我们使用了众所周知的茶碱适体来说明这些原理。在配体结合能力结构中，适体代表开关的一种结构构象，而破坏结合能力结构的另一种折叠则形成另一种构象。为了简单起见，我们不纳入任何控制转录或翻译的调节机制。我们通过明确剖析和详细解释必要步骤，阐明了一种常用的设计流程。我们开发了一种新的目标函数，该函数指定了这种简单的、配体触发的核酶开关的机制，并描述了一个广泛的计算分析管道，以评估设计序列的重要动力学特性。该方案和开发的软件可以很容易地扩展或适应新的设计方案，因此可以作为未来需求的模板。
