Structural basis for the resistance of Tay-Sachs ganglioside GM2 to enzymatic degradation

To understand the reason why, in the absence of GM2 activator protein, the GaLNAc and the NeuAc in GM2 (GalNAc beta 1-->4(NeuAc alpha 2-->3)Gal beta 1 -->4Glc beta 1-1'Cer) are refractory to beta-hexosaminidase A and sialidase , respectively, we have recently synthesized a linkage analogue of GM2 name d 6'GM2 (GalNAc beta 1-->6(NeuAc alpha 2-->3)Gal beta 1-->4Glc beta 1-1'Cer ). While GM2 has GalNAc beta 1-->4Gal linkage, 6'-GM2 has GalNAc beta 1-->6 Gal linkage (Ishida, H., Ito, Y,, Tanahashi, E., Li, Y.-T., Kiso, M., and H asegawa, A (1997) Carbohydr. Res. 302, 223-227). We have studied the enzyma tic susceptibilities of GM2 and 6'GM2, as well as that of the oligosacchari des derived from GM2, asialo-GM2 (GalNAc beta 1-->4Gal beta 1-->4Glc beta 1 -1'Cer) and 6'GM2. In addition, the conformational properties of both GM2 a nd 6'GM2 were analyzed using NMR spectroscopy and molecular mechanics compu tation. In sharp contrast to GM2, the GalNAc and the Neu5Ac of 6'GM2 were r eadily hydrolyzed by beta-hexosaminidase A and sialidase, respectively, wit hout GM2 activator. Among the oligosaccharides derived from GM2, asialo-GM2 , and 6'GM2, only the oligosaccharide from GM2 was resistant to p-hexosamin idase A. Conformational analyses revealed that while GM2 has a compact and rigid oligosaccharide head group, 6'GM2 has an open spatial arrangement of the sugar units, with the GalNAc and the Neu5Ac freely accessible to extern al interactions. These results strongly indicate that the resistance of GM2 to enzymatic hydrolysis is because of the specific rigid conformation of t he GM2 oligosaccharide.