Formation of Ligand Clusters on Multimodal Chromatographic Surfaces

Abstract

Multimodal chromatography is a powerful tool which uses multiple modes of interaction, such as charge and hydrophobicity, to purify protein-based therapeutics. In this work, we performed molecular dynamics simulations of a series of multimodal cation-exchange ligands immobilized on a hydrophilic self-assembled monolayer surface at the commercially relevant surface density (1 ligand/nm2). We found that ligands that were flexible and terminated in a hydrophobic group had a propensity to aggregate on the surface, while less flexible ligands containing a hydrophobic group closer to the surface did not aggregate. For aggregating ligands, this resulted in the formation of a surface pattern that contained relatively large patches of hydrophobicity and charge whose sizes exceeded the length scale of the individual ligands. On the other hand, lowering the surface density to 1 ligand/3 nm2 reduced or eliminated this aggregation behavior. In addition, the introduction of a flexible linker (corresponding to the commercially available ligand) enhanced cluster formation and allowed aggregation to occur at lower surface densities. Further, the use of flexible linkers enabled hydrophobic groups to collapse to the surface, reducing their accessibility. Finally, we developed an approach for quantifying differences in the observed surface patterns by calculating distributions of the patch size and patch length. This clustering phenomenon is likely to play a key role in governing protein–surface interactions in multimodal chromatography. This new understanding of multimodal surfaces has important implications for developing improved predictive models and designing new classes of multimodal separation materials.

Insight into molecular characteristics of SARS-CoV-2 spike protein following D614G point mutation, a molecular dynamics study

“…Position restraint process was used in NVT and NPT ensemble steps. In addition, other algorithms such as Lennard-Jones potentials (Bilodeau et al, 2019) and PME (Collier et al, 2009) were employed to handle van der Waals and electrostatic interactions. The structures obtained at this stage were used to perform the desired analyzes.…”

Probing IgG1 FC–Multimodal Nanoparticle Interactions: A Combined Nuclear Magnetic Resonance and Molecular Dynamics Simulations Approach

“…Ligand density has been shown to have an important impact on the behavior of MM chromatographic systems. ,, Further, recent simulation work in our group has shown that the Capto ligand at sufficiently high-ligand surface densities tends to self-associate, resulting in the formation of hydrophobic clusters on the surface . Importantly, this behavior did not occur in simulations at lower “Capto ligand” densities or with “Nuvia ligand”-functionalized surfaces.…”

“…Finally, we employ MD simulations to provide further insights into the adsorption of the F C molecule in these different MM systems. MM ligand-immobilized SAM surfaces have been shown to present patterned surfaces with patches of charge and hydrophobicity in the absence of a protein . Here, we examine the impact of this ligand self-association phenomenon on protein adsorption by examining the intermolecular interactions and mechanisms occurring during binding of the F C to these different MM ligand surfaces.…”

A framework for calculating orthogonal selectivities in multimodal systems directly from cell culture fluid

“…At pH 7.0 (Figure 3b), this orthogonality was apparent for both resins, while at pH 6.0 (Figure 3a), only Capto MMC ImpRes displayed this behavior. This difference in pH behavior is likely due to ligand density effects and perhaps ligand clustering (Bilodeau, Lau, et al, 2020;Bilodeau et al, 2019). These surprising results further demonstrate the importance of having a quantitative method to determine orthogonality which can identify counterintuitive orthogonality trends and opportunities.…”