Conformational analysis of an alpha-galactosyl trisaccharide epitope involved in hyperacute rejection upon xenotransplantation

alpha-Galactosyl epitopes are carbohydrate structures bearing an alpha-Gal- (1-->3)-Gal terminus (alpha-Gal epitopes). The interaction of these epitope s on the surface of animal cells with anti alpha-Gal antibodies in human se rum is believed to be the main cause in antibody-mediated hyperacute reject ion in xenotransplantation. In this paper, conformational analysis of an N- linked alpha-D-Galp-(1 --> 3)-beta-D-Galp-(1 --> 4)-beta-D-Glcp trisacchari de epitope was conducted in terms of each monosaccharide residue conformati on, primary hydroxymethyl group configuration, and interglycosidic conforma tions. Selective 2D J-delta INEPT experiments have been carried out at thre e different temperatures to evaluate three-bond, long-range C-13-H-1 coupli ng constants for the crucial alpha-(1 --> 3) linkage. The NMR experimental data were complemented by theoretical calculations. The flexibility and dyn amics of the trisaccharide have been studied by Metropolis Monte Carlo simu lations. Ensemble-averaged three-bond, long-range C-13-H-1 coupling constan ts and nuclear Overhauser effects were in good agreement with the experimen tal data. The alpha-(1 --> 3) glycosidic linkage has shown a restricted fle xibility as indicated by NMR spectroscopy and molecular modeling. (C) 1999 Elsevier Science Ltd. All rights reserved.