Probing the effects of surface hydrophobicity and tether orientation on antibody-antigen binding

Abstract

Antibody microarrays have the potential to revolutionize molecular detection for many applications, but their current use is limited by poor reliability, and efforts to change this have not yielded fruitful results. One difficulty which limits the rational engineering of next-generation devices is that little is known, at the molecular level, about the antibody-antigen binding process near solid surfaces. Atomic-level structural information is scant because typical experimental techniques (X-ray crystallography and NMR) cannot be used to image proteins bound to surfaces. To overcome this limitation, this study uses molecular simulation and an advanced, experimentally validated, coarse-grain, protein-surface model to compare fab-lysozyme binding in bulk solution and when the fab is tethered to hydrophobic and hydrophilic surfaces. The results show that the tether site in the fab, as well as the surface hydrophobicity, significantly impacts the binding process and suggests that the optimal design involves tethering fabs upright on a hydrophilic surface. The results offer an unprecedented, molecular-level picture of the binding process and give hope that the rational design of protein-microarrays is possible.

The Locational Impact of Site-Specific PEGylation: Streamlined Screening with Cell-Free Protein Expression and Coarse-Grain Simulation

“…We modified the coarse-grain simulation to include a conjugated PEG chain at a specified site. The methods used are those that have been outlined previously ,− with a few modifications to include PEG, as detailed in the Methods. While the mechanism behind PEG-based stabilization is not well understood, there are two primary theories: PEG stabilizes proteins through (1) direct interaction with the protein surface ,, or (2) entropic interactions with the solvent. , In this study we limited the simulation to evaluate only the entropic effect.…”

Investigating the Effect of Two-Point Surface Attachment on Enzyme Stability and Activity

“…This model employed the Karanicolas and Brooks Go-like protein model − and a corresponding surface potential that quantitatively describes surface hydrophobicity. Several studies have demonstrated the accuracy of predictions from this model for protein conformation and orientation while interacting with various sensor surfaces. ,,,,,− The detailed surface potential and various parameters were developed and published in earlier works , and are included in the Supporting Information for completeness. The simulation details are also described in the Supporting Information.…”

The effects of tether placement on antibody stability on surfaces

“…The model is transferable to peptides of any type and has been used to accurately model proteins of many sizes including enzymes and antibodies. 36,38,39,50 The model was parameterized with data at typical biological conditions (e.g., 6 pH 8, [Na] 200 mM) and is not valid far away from such. However, such limitation is of little importance to this work as antibody arrays are used at biological conditions.…”

“…As has been explained in other works, stabilization on the surface occurs because the protein can make the same favorable contacts in the folded state as in the bulk. 34,37,39,50,60 Specifically, surfaces always stabilize proteins entropically because the surface reduces the entropic cost of folding. [See especially Refs.…”