CONTRIBUTION OF THE OLIGOSACCHARIDE MOIETIES TO THE HETEROGENEITY OF VIP RECEPTORS

In this study, we combined affinity labeling with glycosidase treatments in order to characterize the oligosaccharide moieties of VIP receptors from the human cell line IGR39, human and rat intestinal cells, and pig and rat liver. When using various non cleavable crosslinking reagents, BSOCOES was the most efficient in promoting VIP receptor labeling.-Apparent molecular weights of major I-125-VIP-labeled components were heterogenous, ranging from Mr=52,700 to 67,800. Treatment with peptide-N4-(N-acetyl-beta-D-glucosaminyl) asparagine amidase (PNGase F), which hydrolyses the glycosamine linkage of N-glycoproteins, resulted in a strong decrease of heterogeneity. Molecular weights of deglycosylated complexes ranged from Mr=40,000 to 43,800, thus reducing the maximal Mr difference from 15,100 to only 3,800 after enzyme digestion. Moreover, neuraminidase treatment of VIP receptor revealed interesting differential sensitivity to this enzyme according to the tissue studied. These results indicated that the VIP receptors from the membranes used in this study possess protein backbones with similar size, whatever the species (human, rat, pig) and the tissue (melanoma, intestine, liver). Therefore, differences in the amount of asparagine-linked glycans, as well as in the composition of the oligosaccharide chains, can account for VIP receptor heterogeneity among tissues and species.