Abiotic Stimuli-Responsive Protein Affinity Reagent for IgG.

Abstract

We describe an approach for the discovery of protein affinity reagents (PARs). Abiotic synthetic hydrogel copolymers can be "tuned" for selective protein capture by the type and ratios of functional monomers included in their polymerization and by the polymerization conditions (i.e., pH). By screening libraries of hydrogel nanoparticles (NPs) containing charged and hydrophobic groups against a protein target (IgG), a stimuli-responsive PAR is selected. The robust carbon backbone synthetic copolymer is rapidly synthesized in the chemistry laboratory from readily available monomers. The production of the PAR does not require living cells and is free from biological contamination. The capture and release of the protein by the copolymer NP is reversible. IgG is sequestered from human serum at pH 6.5 and following a wash step, the purified protein is released by elevating the pH to 7.3. The binding and release of the protein occur without denaturation. The abiotic material functions as a selective PAR for the F(ab')2 domain of IgG for pull-down and immunoprecipitation experiments and for isolation and purification of proteins from complex biological mixtures.