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 1:1 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 1:1 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.

Keywords: 1:1 protein-ligand complexes; Drug optimization; Kinetic binding data; Measurement protocol; Thermodynamic binding data; kinITC.