Human glycophorin, the major sialoglycoprotein of erythrocyte membranes, was isolated from erythrocytes of healthy individuals and four patients with CDG syndrome. Sugar analysis revealed lower carbohydrate content in three out of four CDG-glycophorin samples. In order to characterize closer the glycosylation differences between glycophorin samples in health and disease, reaction with four biotinylated lectins was performed, using ELISA procedure on polystyrene microplates. Results obtained so far strongly suggest that both N- and O-glycans of glycophorin are affected in CDG syndrome.
Glycophorin A (GPA), the major sialoglycoprotein of the human erythrocyte membrane, was isolated from erythrocytes of healthy individuals of blood groups A, B and O using phenol-water extraction of erythrocyte membranes. Interaction of individual GPA samples with three lectins (Psathyrella velutina lectin, PVL; Triticum vulgaris lectin, WGA and Sambucus nigra I agglutinin SNA-I) was analyzed using a BIAcore biosensor equipped with a surface plasmon resonance (SPR) detector. The experiments showed no substantial differences in the interaction between native and desialylated GPA samples originating from erythrocytes of either blood group and each of the lectins. Desialylated samples reacted weaker than the native ones with all three lectins. PVL reacted about 50-fold more strongly than WGA which, similar to PVL, recognizes GlcNAc and Neu5Ac residues. SNA-I lectin, recognizing alpha2-6 linked Neu5Ac residues, showed relatively weak reaction with native and only residual reaction with desialylated GPA samples. The data obtained show that SPR is a valuable method to determine interaction of glycoproteins with lectins, which potentially can be used to detect differences in the carbohydrate moiety of individual glycoprotein samples.
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