Molecular mimicry in the recognition of glycosphingolipids by Galα3Galβ4GlcNAcβ-binding Clostridium difficile toxin A, human natural anti α-galactosyl IgG and the monoclonal antibody Gal-13: characterization of a binding-active human glycosphingolipid, non-identical with the animal receptor

• Published in: Glycobiology (Oxford) Vol. 6; no. 6; pp. 599 - 609
• Main Authors: Teneberg, Susann, Lönnroth, Ivar, López, Javier F. Torres, Galili, Uri, Halvarsson, Maria Ölwegård, Ångström, Jonas, Karlsson, Karl-Anders
• Format: Journal Article
• Language: English
• Published: Oxford University Press 1996
• Subjects: Clostridium difficile toxin A; Gal-13 monoclonal antibody; Galα3Galβ4GlcNAcβ epitope; human anti α-galactosyl IgG; x2 glycosphingolipid
• ISSN: 1460-2423; 0959-6658; 1460-2423
• DOI: 10.1093/glycob/6.6.599

Glycoconjugates with terminal Galα3Galβ4GlcNAcβ sequences have been shown to be recognized by three carbohydrate-binding proteins: toxin A of Clostridium difficile, human natural anti α-galactosyl IgG and the monoclonal antibody Gal-13. However, the biological significance of this binding specificity in humans is unclear, since unsubstituted Galα3Galβ4GlcNAcβ sequences are not found in human tissues, due to suppression of the gene coding for the enzyme Galα3-transferase. To explore this inconsistency, the binding of toxin A, human natural anti α-galactosyl IgG, and the Gal-13 monoclonal antibody to various glycosphingolipids was examined using the thin-layer chromatogram binding assay. The binding to Galα3Galβ4GlcNAcβ-terminated glycosphingolipids of rabbit erythrocytes was confirmed. A minor binding-active compound was also detected in the non-acid glycosphingolipid fraction of human erythrocytes. This glycosphingolipid was isolated and characterized by EI mass spectrometry, gas chromatography-EI mass spectrometry after degradation, and proton NMR spectroscopy, as GalNAcβ3Galβ4GlcNAcβ3Galβ4Glcβ1Cer, corresponding to the x2 glycosphingolipid isolated before from this source. Two additional binding-active glycosphingolipids were found. One was GalNAcα3Galβ4GlcNAcβ4Glcβ1Cer produced from blood group A-active GalNAcα3(Fucβ2)Galβ4GlcNAcβ3Galβ4Glcβ1Cer by acid-induced defucosylation. The other was GlcNAcα3Galβ4GlcNAcβ4Glcβ1Cer, generated from NeuGcαGalβ4GlcNAcβ3Galfβ4GlcNAcβ1Cer by enzymatic hydrolysis. A number of other glycosphingolipid sequences, including the human Lex, Ley and I blood group determinants suggested to act as receptors for toxin A, were not recognized by the three ligands. Despite the different terminal substituents and anomerity of the binding-active glycosphingolipids, calculated minimum energy conformations demonstrated topographical similarities in the spatial orientation of the terminal trisaccharides, possibly accounting for the crossreactivity.