Measurement of chemical exchange between RNA conformers by 19F NMR

Many noncoding RNA molecules adopt alternative secondary and tertiary conformations that are critical for their roles in gene expression. Although many of these rearrangements are mediated by other biomolecular components, it is important to evaluate the equilibrium relationship of the conformers. To measure the spontaneous interconversion in a bi-stable RNA stem loop sequence into which a single 19F-uridine label was incorporated, a 19F-19F EXSY experiment was employed. The kinetic exchange rate measured from EXSY experiments for this system was 37.3 ± 2.8 s−1. The advantage of this approach is that exchange kinetics can be monitored in any RNA sequence into which a single 19F nucleotide is incorporated by commercial synthesis. This method is therefore suitable for application to biologically significant systems in which dynamic conformational rearrangement is important for function and may therefore facilitate studies of RNA structure-function relationships.