Carbohydrate-antibody interactions by NMR for a13C-labelled disaccharide ligand

Abstract

Incorporation of a13C label into a carbohydrate ligand, methyl 3-O-(3,6-dideoxy-α-D-xylohexopyranosyl)-2-O-methyl-α-D-mannopyranoside permitted by NMR spectroscopy the study of its binding to the Fab from a monoclonal antibody, Se 155-4. The signals of the free and bound form were observed in the13C spectrum of the carbohydrate-protein complex. The dissociation rate constants were consequently determined by full lineshape analysis of the13C spectrum. Comparison with simplified analyses relying only on the linewidth of the1H and13C signals of the free ligand were made and the justifications of underlying assumptions used in these analyses were discussed. For1H NMR, the protein resonances were purged with a13C filter to observe only the ligand resonances and NOEs between the ligand and the protein.Key words: carbohydrate, binding, NMR, C-13 label, chemical exchange.