title: Affinity polymers tailored for the protein A binding site of immunoglobulin G proteins.
abstract: Rational design in combination with a screening process was used to develop affinity polymers for a specific binding site on the surface of immunoglobulin G (IgG) proteins. The concept starts with the identification of critical amino acid residues on the protein interface and their topological arrangement. Appropriate binding monomers were subsequently synthesized. Together with a sugar monomer (2-5 equiv) for water solubility and a dansyl monomer (0.5 equiv) as a fluorescent label, they were subjected in aqueous solution to linear radical copolymerization in various compositions (e.g., azobisisobutyronitrile (AIBN), homogeneous water/DMF mixtures). After ultrafiltration and lyophilization, colorless dry water-soluble powders were obtained. NMR spectroscopic and gel permeation chromatography (GPC) characterization indicated molecular weights between 30 and 500 kD and confirmed retention of monomer composition as well as the absence of monomers. In a competitive enzyme-linked immunosorbent assay (ELISA) screen of the polymer libraries (20-50 members), few copolymers qualified as strong and selective binders for the protein A binding site on the Fc fragment of the antibody. Their monomer composition precisely reflected the critical amino acids found at the interface. The simple combination of a charged and a nonpolar binding monomer sufficed for selective submicromolar IgG recognition by the synthetic polymer. Affinities were confirmed by fluorescence titrations; they increased with decreasing salt load but remained largely unaltered at lowered pH. Other proteins, including those of similar size and isoelectric point (pI), were bound 10-1000 times less tightly. This example indicates that interaction domains in other proteins may also be targeted by synthetic polymers if their comonomer composition reflects the nature and arrangement of amino acid residues on the protein surface.
authors: Department of Chemistry, University of Duisburg-Essen, Universitätsstrasse 7, 45117 Essen (Germany), Fax: (+49) 201-183-4252.
authors: Patricia
authors: P
authors: Latza
authors: Patrick
authors: P
authors: Gilles
authors: Torsten
authors: T
authors: Schaller
authors: Thomas
authors: T
authors: Schrader
mesh_terms: D001665:Binding Sites; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry* / Q000378:metabolism*; D008958:Models, Molecular; D011108:Polymers / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D013205:Staphylococcal Protein A / Q000378:metabolism*
chemical_list: D007074:Immunoglobulin G; D011108:Polymers; D013205:Staphylococcal Protein A
keywords: antibodies; immunoglobulins; polymers; protein recognition; supramolecular chemistry