title: Clusterin binds by a multivalent mechanism to the Fc and Fab regions of IgG.
abstract: Clusterin was purified from human serum by IgG and monoclonal antibody affinity chromatography. SDS-PAGE and immunoblotting revealed no major differences between clusterin prepared in these two ways. An ELISA method for measuring the binding of clusterin to immunoglobulins was developed. Clusterin purified by IgG affinity chromatography bound to pooled human IgG with a similar affinity (S0.5 5.9 +/- 0.4 micrograms/ml) as clusterin purified by monoclonal antibody chromatography (S0.5 6.1 +/- 0.2 micrograms/ml). The apparent affinity of clusterin for IgG immobilised on ELISA plates increased with increasing concentrations of IgG in the coating solution. Aggregated IgG in solution was a more potent inhibitor of the binding of clusterin to immobilised IgG than was monomer IgG. Clusterin bound to all of the isotypes of human IgG, and to human IgA and IgM, with apparent affinities in the order IgG3 > IgG4 > IgM > IgG1 > IgG2, IgA. Clusterin bound to both the Fab and Fc fragments of human IgG. The clusterin binding site(s) on the Fc do not overlap with those for protein A and Clq.
authors.affiliation: Department of Biochemistry, University of Sydney, NSW, Australia.
authors.forename: M R
authors.initials: MR
authors.lastname: Wilson
authors.forename: S B
authors.initials: SB
authors.lastname: Easterbrook-Smith
mesh_terms: D000911:Antibodies, Monoclonal / Q000276:immunology; D001667:Binding, Competitive; D002846:Chromatography, Affinity; D051152:Clusterin; D015922:Complement C1q / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D006023:Glycoproteins / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D015151:Immunoblotting; D007140:Immunoglobulin Fab Fragments / Q000378:metabolism*; D007141:Immunoglobulin Fc Fragments / Q000378:metabolism*; D007074:Immunoglobulin G / Q000378:metabolism*; D018832:Molecular Chaperones; D013205:Staphylococcal Protein A / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; C495242:CLU protein, human; D051152:Clusterin; D006023:Glycoproteins; D007140:Immunoglobulin Fab Fragments; D007141:Immunoglobulin Fc Fragments; D007074:Immunoglobulin G; D018832:Molecular Chaperones; D013205:Staphylococcal Protein A; D015922:Complement C1q