Protein-sugar interactions: preparation, purification, and properties of rabbit antibodies against di-N-acetylchitobiose.
Antibodies against di-N-acetylchitobiose (CB) were raised in rabbits after injection of CB-bovine serum albumin conjugates and were fractionated by columns into two classes: the first bound to a column of Sepharose covalently coupled with N-acetylglucosamine (GlcNAc); the second bound to a column coupled with CB. Active antibodies were eluted by a moderate concentration of a chaotropic agent, but not by high ionic strength buffers or acidic buffers. The active fractions were identified as IgG by ultracentrifugation and immuno-electrophoresis. These antibodies gave precipitation bands with CB-protein conjugates and this reaction could be reversed in the presence of free CB. The GlcNAc-bound fraction quantitatively quenched the fluorescence of O-(4-methylumbelliferyl)-glucosides of GlcNAc and CB, while the CB-bound fraction quenched only the glycoside of CB; other O-(4-methylumbelliferyl)-glucosides were not quenched. Among eleven monosaccharides and oligosaccharides, only GlcNAc, CB, and tri-N-acetylchitotriose were able to inhibit the precipitation of antibodies against CB with CB-protein conjugate. These antibodies failed to agglutinate erythrocytes from various species but did agglutinate transformed cells and mouse lymphocytes. The binding of these antibodies on cell lembranes was reversed by free CB and by CB-protein conjugates. The properties of these antibodies are related to those of lectins with similar specificities and to the structure of glycoconjugates.
D000117:Acetylglucosamine / Q000276:immunology; D000818:Animals; D000906:Antibodies / Q000302:isolation & purification*; D000918:Antibody Specificity; D000937:Antigen-Antibody Reactions; D002846:Chromatography, Affinity; D004187:Disaccharides / Q000276:immunology*; D004912:Erythrocytes / Q000276:immunology; D006801:Humans; D008214:Lymphocytes / Q000276:immunology; D051379:Mice; D009374:Neoplasms, Experimental / Q000276:immunology; D010954:Plasmacytoma / Q000276:immunology; D011817:Rabbits
D000906:Antibodies; D004187:Disaccharides; D000117:Acetylglucosamine
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