Corrin Ring Modifications Reveal the Chemical and Spatial Requirements for the B 12 -btuB Riboswitch Interaction.

The btuB riboswitch is a regulatory RNA sequence controlling gene expression of the outer membrane B 12 transport protein BtuB by specifically binding coenzyme B 12 (AdoCbl) as its natural ligand. The B 12 sensing riboswitch class is known to accept various B 12 derivatives, leading to a division into two riboswitch subclasses, dependent on the size of the apical ligand. Here we focus on the role of side chains b and e on affinity and proper recognition, i. e. correct structural switch of the btuB RNA, which belongs to the AdoCbl-binding class I. Chemical modification of these side chains disturbs crucial hydrogen bonds and/or electrostatic interactions with the RNA, its effect on both affinity and switching being monitored by in-line probing. Chemical modifications at sidechain b of vitamin B 12 show larger effects indicating crucial B 12 -RNA interactions. When introducing the same modification to AdoCbl the influence of any side-chain modification tested is reduced. This renders the impact of the adenosyl-ligand for B 12 -btuB riboswitch recognition clearly beyond the known role in affinity.