ABSTRACT
The binding affinity of the oligosaccharide moiety of a neutral glycosphingolipid, asialoGM1, towards Ricinus communis agglutinin (RCAI) was determined for the first time by fluorescence resonance energy transfer (RET). The asialoGM1 was incorporated into a phospholipid (DMPC) vesicle doped with dansylated DPPE and then titrated with an increasing amount of the galactose specific RCAI. The efficiency of RET was determined by a saturable increase in the quenching of 'donor' fluorescence, i.e. the 'trp' residue of RCAI, due to the energy transfer from the 'acceptor' dansyl group on the surface of the vesicle. The apparent binding constant was found to be in the range of 10(5)-10(6) M-1 at 27 degrees C.
Subject(s)
Carbohydrates/analysis , Dimyristoylphosphatidylcholine , Energy Transfer , Galactose/analysis , Liposomes , Oligosaccharides/analysis , Phosphatidylethanolamines , Ricin , Spectrometry, FluorescenceABSTRACT
To examine how surface potential controls the reactivity of glycoconjugates at cell surface, the interaction of galactose-specific lectins e.g. peanut agglutinin, Ricinus cummunis agglutinin with liposomes bearing asialo GM1 were studied in the presence of varying amount of ganglioside mixture, GMn. The presence of 5% GMn causes complete slowing down of precipitin reaction and thereby make carbohydrate moiety of asialo GM1 completely inaccessible i.e. ‘cryptic’. In contrast the presence of 1–2% GMn enhances the apparent rate and amplitude of the precipitin reaction as surface potential becomes more negative. The relevance of the findings has been discussed in relation to the expression and involvement of the cell-surface sialic acid residues during development and differentiation.