Protein-Chromatographic Ligand Interaction
For chromatographic separation, ligands are expected to interact with
different regions of the molecules based on their surface properties and
inherit affinity for a given region. To answer the question of preferred
protein-ligand interactions that lead to separation, different docking
studies were performed. Multiple single ligand docking assessments were
performed to the entire antibody surface, the Fab region, the CDR
region, and the Fc region. The contact sites for protein/ligand and
associated affinities were ranked based on interaction energy. Figure 1A
shows that the region with the highest (more negative ΔG) protein-ligand
interaction energies for the top three binding sites differed for the
three antibodies. There is a correlation between in silicobinding affinities (ΔG) and experimentally determined protein retention.
Previous studies have demonstrated that a single post translational
modifications (PTM) in the CDR region not only affect antibody potency
but can have significant effect on chromatographic separation (Kern,
Mende, Denefeld, Sackewitz, & Chelius, 2014). Thus, chromatographic
ligand binding to CDR region can elucidate potential chromatographic
separation (Figure 1B).