Toward Rational Design of Selective Molecularly Imprinted Polymers (MIPs) for Proteins: Computational and Experimental Studies of Acrylamide Based Polymers for Myoglobin

Abstract

Molecularly imprinted polymers (MIPs) have potential as alternatives to antibodies in the diagnosis and treatment of disease. However, atomistic level knowledge of the prepolymerization process is limited that would facilitate rational design of more efficient MIPs. Accordingly, we have investigated using computation and experiment the proteinmonomer binding interactions that may influence the desired specificity. Myoglobin was used as the target protein and five different acrylamide-based monomers considered. Protein binding sites were predicted using SiteMap and binding free energies of monomers at each site calculated using MM-GBSA. Statistical thermodynamic analysis and study of atomistic interactions facilitated rationalization of monomer performance in MIP rebinding studies (% rebind; imprinting factors). CD spectroscopy was used to determine monomer effects on myoglobin secondary structure, with all monomers except the smallest monomer (acrylamide) causing significant changes. A complex interplay between different protein-monomer binding effects and MIP efficacy was observed. Validation of hypotheses for key binding features was achieved by rational selection of two different co-monomer MIP combinations that produced experimental results in agreement with predictions. The co-monomer studies revealed that uniform, non-competitive binding of monomers around a target protein is favourable. This study represents a step towards future rational in silico design of MIPs for proteins.

Evaluation of acrylamide-based molecularly imprinted polymer thin-sheets for specific protein capture—a myoglobin model

“…We produced a series of acrylamide-based (AAm, NHMAm, NHEAm, DMAm, and TrisNHMAm) hydrogel MIPs with selective recognition for the target protein myoglobin [ 30 ]. By comparing the MIPs with their corresponding non-imprinted polymer (NIP) controls, an imprinting factor (IF) was calculated, using equation ( 1 ) and used to assess performance.…”

“…Monomers interacting with proteins could disrupt the secondary structure within the protein, especially if the monomer interactions are with functional groups along the protein backbone [ 50 ]. Indeed, Sullivan et al [ 30 ] showed that the hydroxyl groups in TrisNHMAm were able to strongly hydrogen bond to helical backbone residues within the potential binding sites of myoglobin leading to changes in this target’s secondary structure. In addition to this, TrisNHMAm has a low IF value of 1.10 suggesting that the MIP behaves similarly to the NIP and is therefore deemed unacceptable for use as a MIP.…”

“…However, since the bulk MIP approach is more easily scalable and uses low cost monomer reagents, such losses can be acceptable. For example, 3–5g of useable bulk MIP microparticles can be produced per batch within 24 h [ 10 , 30 ].…”

Hybrid Aptamer‐Molecularly Imprinted Polymer (aptaMIP) Nanoparticles from Protein Recognition—A Trypsin Model

“…Removal of the template molecule from the polymer results in the formation of specific binding cavities, complementary in terms of structure and functionality to the template molecule. [ 13–15 ] For several years MIPs were limited by high levels of heterogeneity and by synthetic methods, but significant advances in molecular modeling and understanding of polymer chemistry have countered these issues.…”

Evaluation of Molecularly Imprinted Polymers for Point-of-Care Testing for Cardiovascular Disease

“…Hence, there are several factors that significantly influence the generation of specific binding sites. A study by Sullivan et al demonstrated that acrylamide-based monomers that were smaller in size and that formed comparatively less hydrogen bonds with the side chains than the protein backbone, resulted in less secondary structural changes within the protein template prior to polymerisation [91]. Notably, if the conformational structure of the template protein is disturbed before or during polymerisation, the PIP will consist of binding sites that will inhibit rebinding.…”