Isothermal Titration Calorimetry as a Method for Analyzing Protein Adsorption in Ion Exchange Chromatography

Abstract

Ion exchange chromatography is a widely used method for purification of all types of biomolecules in current biotechnological downstream processes. Knowledge on the binding behavior of proteins provides valuable insight for understanding the molecular mechanisms of protein interactions in a biological context. However, thermodynamic parameters such as enthalpy and entropy changes that characterize protein adsorption are still unknown. Isothermal titration calorimetry applications in biosciences has gained its merit to study binding of soluble molecules, protein inhibition, conformational changes, reaction kinetics, and protein adsorption. However, in the case of protein adsorption, a lot of complications arise since the usual models used to study protein interactions in solution are no longer valid. This work explains a detailed methodology for the obtention of adsorption enthalpy, entropy and Gibbs energy of protein adsorption, by using ITC together with equilibrium adsorption isotherms.