The impact of second-shell nucleotides on ligand specificity in cyclic dinucleotide riboswitches

Cyclic dinucleotide riboswitches are specific for cyclic-di-GMP or cyclic-AMP-GMP. Specificity is derived from two binding site nucleotides. A new variant of the CDN riboswitch has been found to have a mutation in the binding site and is promiscuous for both ligands. The sequencing dataset presented here was collected by SMARTT (Sequencing-based Mutational Analysis of RNA Transcription Termination), a sequencing-based high-throughput assay that monitors the in vitro transcription termination efficiencies of individual mutant constructs as a function of ligand concentration. This dataset reveals the functional effects of over 7000 mutants of the promiscuous CDN riboswitch from Acholeplasma axanthum, both in the context of the wild-type construct and in the background of a binding site mutation that preferentially binds cyclic-di-GMP. This comprehensive mutational data identifies nucleotides involved in ligand specificity and promiscuity.