High resolution proton NMR studies of gangliosides. Structure of two types of GD3 lactones and their reactivity with monoclonal antibody R24.

Ganglioside GD3 was converted at room temperature to two stable lactones, denoted as GD3 lactones I and II. The reaction sequence was presumed to be GD3----GD3 lactone I----GD3 lactone II based on the time course of their production. Lactone I behaved as a monosialoganglioside and lactone II as a neutral species. The two lactones were isolated by DEAE-Sephadex column chromatography. The positions of the inner ester linkages were investigated by two-dimensional J-correlated proton NMR spectroscopy. An ester linkage was most likely formed between the carboxyl group of the external sialic acid residue and C9-OH of the internal sialic acid residue in lactone I. In addition to this ester linkage, a second ester linkage between the carboxyl group of the internal sialic acid and C2-OH of the galactose residue was likely formed in lactone II. The structural changes induced by lactonization were further examined by their reactivity with the monoclonal antibody R24 (Puckel, C. S., Lloyd, K. O., Travassos, L. R., Dippold, W. G., Oettgen, H. F., and Old, L. J. (1982) J. Exp. Med. 155, 1133-1147), which reacted with GD3. R24 was found to bind weakly to GD3 lactone I, but not to GD3 lactone II. The results suggest that the monoclonal antibody requires both sialic acid residues for high affinity binding, and the complete lactonization results in a loss of negative charges and/or a change in the overall conformation of the oligosaccharide moiety which may account for the loss of binding.