Rational Re-engineering of a Transcriptional Silencing PreQ1 Riboswitch

• DOI: 10.1021/jacs.5b03405
• Publication Date: 2015-06-24T18:55:20Z
• Authors: Ming-Cheng Wu; Phillip T. Lowe; Christopher J. Ro...; Helen A. Vincent; Neil Dixon; James Leigh; Jason Micklefield
• Keywords: Riboswitch; Synthetic Biology; Rational design

Re-engineered riboswitches that no longer respond to cellular metabolites, but instead can be controlled by synthetic molecules, are potentially useful gene regulatory tools for use in biology and biotechnology fields. Previously, extensive genetic selection screening approaches were employed re-engineer a natural adenine riboswitch create orthogonal ON-switches, enabling translational control of target expression response ligands. Here, we describe how rational targeted approach was used the PreQ1 from Bacillus subtilis into an OFF-switch. In this case, evaluation just six compounds with seven mutants led identification riboswitch–ligand pairing effectively repressed transcription selected genes B. subtilis. The streamlining re-engineering approach, its extension second class riboswitches, provides methodological platform creation new components biotechnological applications including functional analysis antimicrobial validation screening.