L-Proline Destabilization of RNA Duplexes is Temperature Dependent

Abstract

L-proline is an osmoprotectant and nonessential amino acid that destabilizes nucleic acid secondary and tertiary structures and, at high concentrations, denatures nucleic acids. In this work the interactions between L-proline and the functional groups on the solvent accessible nucleic acid surface area that would be exposed by unfolding (ΔASA) of eleven RNA dodecamer duplexes with guanine-cytosine (GC) content ranging from 17-100% were quantified using thermal and titration denaturation monitored by uv-absorbance. L-proline destabilized GC-rich duplexes to a greater degree during thermal denaturation than low GC-content duplexes due to greater accumulation of L-proline in the vicinity of the RNA ΔASA. This accumulation was temperature dependent; L-proline-RNA interactions quantified using titration denaturation at 25 C were nearly independent of GC-content. Thus, the greater destabilization of RNA GC-rich duplexes observed during thermal denaturation was due to greater L-proline interactions with the RNA ΔASA at the higher transition temperatures of GC-rich duplexes. Our results suggest L-proline interactions are entropically-driven, potentially due to dehydration of L-proline when interacting with the RNA ΔASA. In addition, L-proline destabilized the RNA duplexes to a greater degree than glycine betaine and urea. This suggests glycine betaine, L-proline, and urea have the potential to form a triad of cosolutes to probe changes in solvent accessible surface area during biochemical reactions.