title: Thermodynamic investigation of Z33-antibody interaction leads to selective purification of human antibodies.
abstract: Antibodies, such as IgGs, are widely applied as detection probes, purification ligands and targeting moieties in research and medicine. Protein A from Staphylococcus aureus is capable of selectively binding to antibodies. Z33, a 33 amino acid peptide sequence derived from Protein A, is a minimized binding domain with comparable interaction potential. This peptide was fused to two different proteins without perturbing the properties of both the protein and the Z33-domain. The thermodynamic parameters for the interaction of the fusion proteins with antibodies from various species were determined by isothermal titration calorimetry. This showed that binding was enthalpically driven and entropically unfavorable. A difference in Z33 binding affinity of several orders of magnitude was observed between human and bovine antibodies. This selectivity toward human IgGs was utilized for the efficient and selective purification of human IgGs from mixtures containing bovine IgGs and other proteins by affinity precipitation employing a fusion protein of Z33 and a stimulus-responsive elastin-like polypeptide.
authors.affiliation: Radboud University Nijmegen, Institute for Molecules and Materials, Heyendaalseweg 135, 6525 AJ Nijmegen, The Netherlands; Dutch Polymer Institute, P.O. Box 902, 5600 AX Eindhoven, The Netherlands.
authors.forename: Mark B
authors.initials: MB
authors.lastname: van Eldijk
authors.affiliation: Radboud University Nijmegen, Institute for Molecules and Materials, Heyendaalseweg 135, 6525 AJ Nijmegen, The Netherlands.
authors.forename: Ferdinanda C M
authors.initials: FC
authors.lastname: Smits
authors.affiliation: DSM Research, P.O. Box 18, 6160 MB Geleen, The Netherlands.
authors.forename: Jens C
authors.initials: JC
authors.lastname: Thies
authors.affiliation: Radboud University Nijmegen, Institute for Molecules and Materials, Heyendaalseweg 135, 6525 AJ Nijmegen, The Netherlands.
authors.forename: Jasmin
authors.initials: J
authors.lastname: Mecinović
authors.affiliation: Radboud University Nijmegen, Institute for Molecules and Materials, Heyendaalseweg 135, 6525 AJ Nijmegen, The Netherlands. Electronic address: j.vanhest@science.ru.nl.
authors.forename: Jan C M
authors.initials: JC
authors.lastname: van Hest
mesh_terms: D000818:Animals; D001665:Binding Sites; D002417:Cattle; D004549:Elastin / Q000378:metabolism; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D008958:Models, Molecular; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013205:Staphylococcal Protein A / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism*; D013816:Thermodynamics
chemical_list: D007074:Immunoglobulin G; D011993:Recombinant Fusion Proteins; D013205:Staphylococcal Protein A; D004549:Elastin
keywords: Antibody purification; Elastin-like polypeptides; Isothermal titration calorimetry; Protein A; Protein engineering