Jul 01 2018 In silico design of ligand triggered RNA switches

In the world of synthetic biology, the design of RNA switches holds immense promise for various applications, ranging from diagnostics to therapeutics. This paper presents a comprehensive workflow for designing RNA switches that can dynamically alter their structural conformations in response to specific ligands

The research paper outlines a step-by-step RNA design process, shedding light on each crucial stage with intricate detail. From selecting a well-characterized RNA aptamer, such as the theophylline aptamer, to delineating the structural features essential for ligand binding, every aspect of the design process is elucidated.

We introduce a novel, probability-based objective function which allows researchers to quantitatively assess the performance of their designed RNA switches, making it easier to identify the most effective candidates for their specific application.

Building on these advancements, we establish key criteria to ensure robust analysis of RNA switch behavior. This focus on rigorous methodology strengthens the foundation for reliable results in the design process.

Finally, we present a method to visualize the kinetic properties of RNA switch sequences. This visualization tool provides valuable insights into the speed of structural changes within the switch, enabling researchers to efficiently rank and filter designed sequences and pinpoint the optimal candidates for further investigation.

Abstract

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 Kd 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.

Citation

In silico design of ligand triggered RNA switches
Sven Findeiß, Stefan Hammer, Michael T. Wolfinger, Felix Kühnl, Christoph Flamm, Ivo L.Hofacker
Methods 143:90-101 (2018) | doi: 10.1016/j.ymeth.2018.04.003 | Preprint PDF