Molecular insight into protein conformational transition in hydrophobic charge induction chromatography: a molecular dynamics simulation.

Hydrophobic charge induction chromatography (HCIC) is an adsorption chromatography combining hydrophobic interaction in adsorption with electrostatic repulsion in elution. The method has been successfully applied in the separation and purification of antibodies and other proteins. However, little is understood about protein conformational transition and the dynamic process within adsorbent pores. In the present study, a pore model is established to represent the realistic porous adsorbent composed of matrix and immobilized HCIC ligands. Protein adsorption, desorption, and conformational transition in the HCIC pore and its implications to the separation performance are shown by a molecular dynamics simulation of a 46-bead beta-barrel coarse-grained model protein in the adsorbent pore. Repeated adjustment of both protein position and orientation is observed before reaching a stable adsorption. Once the protein is adsorbed, there is a dynamic equilibrium between unfolding and refolding. The effect of hydrophobic interaction strength between protein and ligands on adsorption phenomena is then examined. Strong hydrophobic interaction, representing the presence of high-concentration lyotropic salt in mobile phase, can speed up the adsorption but cause protein unfolding more significantly. On the contrary, weak hydrophobic interaction, representing the absence of a lyotropic salt or the presence of a chaotropic agent, can reserve native protein conformation but does not lead to stable adsorption. In the elution, protein unfolding occurs due to simultaneous hydrophobic adsorption and electrostatic repulsion in the opposite directions. When the protein has been desorbed, the conformational transition between unfolded and native protein is still observed due to the long-range nature of electrostatic interaction. The simulation has provided molecular insight into protein conformational transition in the whole HCIC process, and it would be beneficial to the rational design of ligands and parameter optimizations for high-performance HCIC.