Binding performance of pepsin surface-imprinted polymer particles in protein mixtures

Surface-imprinted polymer particles facilitate the accessibility of synthetic selective binding sites for proteins. Given their volume-to-surface ratio, submicron particles offer a potentially large surface area facilitating fast rebinding kinetics and high binding capacities, as investigated herein by batch rebinding experiments. Polymer particles were prepared with (3-acrylamidopropyl) trimethylammonium chloride as functional monomer, and ethylene glycol dimethacrylate as cross-linker in the presence of pepsin as template molecule via miniemulsion polymerization. The obtained polymer particles had an average particle diameter of 623 nm, and a specific surface area of 50 m(2) g(-1). The dissociation constant and maximum binding capacity were obtained by fitting the Langmuir equation to the corresponding binding isotherm. The dissociation constant was 7.94 mu M, thereby indicating a high affinity; the binding capacity was 0.72 mu mol m(-2). The binding process was remarkably fast, as equilibrium binding was observed after just 1 min of incubation. The previously determined selectivity of the molecularly imprinted polymer for pepsin was for the first time confirmed during competitive binding studies with pepsin, bovine serum albumin, and beta-lactoglobulin. Since pepsin has an exceptionally high content in acidic amino acids enabling strong interactions with positively charged quaternary ammonium groups of the functional monomers, another competitive protein, i.e., alpha 1-acid glycoprotein, was furthermore introduced. This protein has a similarly high content in acidic amino acids, and was used for demonstrating the implications of ionic interactions on the achieved selectivity.