Thermodynamics and Kinetics of RNA Unfolding and Refolding

Abstract

Emergence of novel functions of non-coding RNAs imposes a new challenge for thermodynamics-based structural prediction of RNA. Here we review bulk and single-molecule techniques to measure the thermodynamics and kinetics of RNA folding and unfolding. RNA can be denatured by heat, chemicals, force, and by depletion of divalent cations. Various spectroscopic, calorimetric, chemical and biochemical methods have been used to study RNA structures. We empha- size single-molecule force unfolding as a new and powerful technique to study RNA structure and folding. Using optical tweezers, single RNA molecules can be stretched and relaxed; their changes in extension reflect structural rearrangements. We discuss determination of Gibb free energy of folding from mechanical work under both equilibrium and non-equilibrium conditions. Force can be applied to affect reaction rates as well as to manipulate molecular structure. Folding and unfolding kinetics can be monitored in real time.