Simultaneous determination of thermodynamic and kinetic data by isothermal titration calorimetry.

ABSTRACT

BACKGROUND: Thermodynamic and binding kinetic data increasingly support and guide the drug optimization process. METHODS: Because ITC thermograms contain binding thermodynamic and kinetic information, an efficient protocol for the simultaneous extraction of thermodynamic and kinetic data for 11 protein ligand reactions from AFFINImeter kinITC in one single experiment are presented. RESULTS: The effort to apply this protocol requires the same time as for the standard protocol but increases the precision of both thermodynamic and kinetic data. CONCLUSIONS: The protocol enables reliable extraction of both thermodynamic and kinetic data for 11 protein-ligand binding reactions with improved precision compared to the 'standard protocol'. GENERAL SIGNIFICANCE: Thermodynamic and kinetic data are recorded under exactly the same conditions in solution without any labeling or immobilization from a protein sample that is not 100% active and would otherwise render the extraction of kinetic parameters impossible.