A loop–loop interaction and a K-turn motif located in the lysine aptamer domain are important for the riboswitch gene regulation control

Abstract

The lysine riboswitch is associated to the lysC gene in Bacillus subtilis, and the binding of lysine modulates the RNA structure to allow the formation of an intrinsic terminator presumably involved in transcription attenuation. The complex secondary structure of the lysine riboswitch aptamer is organized around a five-way junction that undergoes structural changes upon ligand binding. Using single-round transcription assays, we show that a loop–loop interaction is important for lysine-induced termination of transcription. Moreover, upon close inspection of the secondary structure, we find that an unconventional kink-turn motif is present in one of the stems participating in the loop–loop interaction. We show that the K-turn adopts a pronounced kink and that it binds the K-turn-binding protein L7Ae of Archaeoglobus fulgidus in the low nanomolar range. The functional importance of this K-turn motif is revealed from single-round transcription assays, which show its importance for efficient transcription termination. This motif is essential for the loop–loop interaction, and consequently, for lysine binding. Taken together, our results depict for the first time the importance of a K-turn-dependent loop–loop interaction for the transcription regulation of a lysine riboswitch.