Entropy-driven folding of an RNA helical junction: An isothermal titration calorimetric (ITC) analysis of the hammerhead ribozyme

RNA helical junctions are common in RNA structures, including ribozymes, mRNA untranslated regions, rRNAs, and regulatory RNAs.?RNA junctions are also in complex with proteins.?However, little was known about the thermodynamic forces that drive the folding of RNA junctions, and it is difficult to dissect these forces for several reasons, including non-two-state folding, secondary and tertiary folding components are superimposable, and opposing enthalpic and entropic contributions to RNA folding.?The authors used Isothermal Titration Calorimetry (ITC) to study the folding of hammerhead ribozyme, a three-way RNA helical junction.?In their work, they had RNA in both the ITC cell and syringe, and during titration, both RNA stands annealed to form complete folded bimolecular ribozyme. This approach was different than other published ITC research on hammerhead ribozymes, which titrated concentrated MgCl2 into unimolecular hammerhead ribozyme, and measured Mg2+ binding (Hammann, C., Cooper, A., Lilley, D.M., Biochemistry 40, 1423-1429, 2001).