Single-molecule investigations of RNA dissociation

Abstract

Given the essential cellular roles for ribonucleic acids (RNAs) it is important to understand the stability of three-dimensional structures formed by these molecules. This study aims to investigate the dissociation energy landscape for simple RNA structures via atomic-force-microscopy-based single-molecule force-spectroscopy measurements. This approach provides details on the locations and relative heights of the energy barriers to dissociation, and thus information upon the relative kinetic stabilities of the formed complexes. Our results indicate that a simple dodecamer RNA helix undergoes a forced dissociation process similar to that previously observed for DNA oligonucleotides. Incorporating a UCU bulge motif is found to introduce an additional energy barrier closer to the bound state, and also to destabilize the duplex. In the absence of magnesium ions a duplex containing this UCU bulge is destabilized and a single, shorter duplex is formed. These results reveal that a bulge motif impacts upon the forced dissociation of RNA and produces an energy landscape sensitive to the presence of magnesium ions. Interestingly, the obtained data compare well with previously reported ensemble measurements, illustrating the potential of this approach to improve our understanding of RNA stability and dissociation kinetics.