Isothermal Titration Calorimetry

Thermodynamic characterization of the system provides us with important information about the stability, strength, specificity, and stoichiometry of interacting systems. The method of choice for the direct measurements of energetics of protein-ligand interactions is the isothermal titration calorimetry (ITC) technique. The use of ITC to measure the binding of a macromolecule (in general, a protein) to its ligand (ion, peptide, another protein, DNA, RNA, and so on) relies on the fact that such an interaction is accompanied by a heat effect. The heat absorbed (endothermic) or released (exothermic) upon the interaction Q is then used to obtain information about the binding constant, K a, and the enthalpy of binding ?H. The strength of ITC is that under proper experimental conditions, from one single titration both the binding constant and the enthalpy of binding can be obtained. Moreover, the temperature dependence of the enthalpy of binding allows one to calculate another important thermodynamic parameter: the heat capacity change of binding ?Cp. The ?Cp is calculated from the slope of ?H vs temperature, and it can be positive (hydrophobic interactions are disrupted upon binding) or negative (hydrophobic interactions are formed) ().