Calorimetry of Nucleic Acids. Issue 1 (1st December 2015) - Rozners, Eriks
Abstract: This unit describes the application of calorimetry to characterize the thermodynamics of nucleic acids, specifically, the two major calorimetric methodologies that are currently employed: differential scanning (DSC) and isothermal titration calorimetry (ITC). DSC is used to study thermally induced order‐disorder transitions in nucleic acids. A DSC instrument measures, as a function of temperature ( T ), the excess heat capacity ( C p ex ) of a nucleic acid solution relative to the same amount of buffer solution. From a single curve of C p ex versus T, one can derive the following information: the transition enthalpy (Δ H ), entropy (Δ S ), free energy (Δ G ), and heat capacity (Δ Cp ); the state of the transition (two‐state versus multistate); and the average size of the molecule that melts as a single thermodynamic entity (e.g., the duplex). ITC is used to study the hybridization of nucleic acid molecules at constant temperature. In an ITC experiment, small aliquots of a titrant nucleic acid solution (strand 1) are added to an analyte nucleic acid solution (strand 2), and the released heat is monitored. ITC yields the stoichiometry of the association reaction ( n ), the enthalpy of association (Δ H ), the equilibrium association constant ( K ), and thus the free energy of association (Δ G ). Once Δ H and Δ G are known, Δ S can also be derived. Repetition of the ITC experiment at a number of different temperatures yields the Δ C p for the association reaction from the temperature dependence of Δ H . © 2015 by John Wiley & Sons, Inc.