Molecular Insight into the Ligand–IgG Interactions for 4-Mercaptoethyl-pyridine Based Hydrophobic Charge-Induction Chromatography

Abstract

Hydrophobic charge-induction chromatography (HCIC) with 4-mercaptoethyl-pyridine (MEP) as the ligand is a novel technology for antibody purification. In the present work, the molecular simulation methods were used to investigate the interactions between MEP ligand and Fc fragment of IgG (Fc-A). Six ligands with different structures of spacer arm were studied with molecular docking and dynamics simulation at neutral and acidic pH. The binding modes and the interaction energies were analyzed. The results indicated that all ligands tested could bind into the selected pocket on the C(H2) domain of Fc-A at neutral pH. The pyridine ring on the top of MEP ligands acts as a major role to provide the hydrophobic association and hydrogen bond for the ligand-IgG binding; meanwhile, the sulfone group on the spacer arm might form the additional hydrogen bond and enhance the binding of ligand onto the surface of IgG. The replacements of thioether sulfur atom on the spacer arm with either nitrogen or oxygen atom seem to have little influence on the binding. The influences of pH on the ligand-IgG interactions were also studied with the molecular dynamics simulation. It was found that MEP ligands would departed from the surface of Fc-A at low pH due to the electrostatic repulsion. The ligands with a sulfone group on the spacer arm would weaken the electrostatic repulsion and need more acidic conditions for the departing of ligand. The molecular simulation results were in agreement with some experimental observations, which would be useful to elucidate the molecular mechanism of HCIC and design a novel ligand to improve the efficiency of antibody separation.

Preparation and evaluation of dextran-grafted agarose resin for hydrophobic charge-induction chromatography

“…Combining several molecular interactions with specially designed ligands [8][9][10], HCIC resins usually show high selectivity on the adsorption of target protein compared to traditional ion-exchange chromatography (IEC) and hydrophobic interaction chromatography (HIC) [11]. However, due to the relatively weak hydrophobic interactions between target protein and ligand, the dynamic binding capacities (DBC) of HCIC resins are limited, especially for low retention time, which strongly impedes the large-scale application of HCIC for protein production.…”

Protein adsorption behavior and immunoglobulin separation with a mixed‐mode resin based on p‐aminohippuric acid

“…For example, 4‐mercaptoethyl‐pyridine (MEP) is a typical MMC ligand with a product name of MEP HyperCel. The heterocyclic group can provide hydrophobic interactions with target proteins and the nitrogen atom of pyridine can form hydrogen bonds with certain residues on protein surface . The multifunctional interactions can improve binding selectivity between ligands and proteins, which provides higher possibilities to optimize operation conditions .…”

Hydrophobic charge‐induction resin with 5‐aminobenzimidazol as the functional ligand: preparation, protein adsorption and immunoglobulin G purification

“…Molecular simulation was used to study interactions between HCIC ligands and the Fc fraction of IgG molecule [18,19,[20][21][22]. It was also used to analyze the recognition mode between IgG-Fc and natural ligands (e.g.…”