Complex Carbohydrate Composition of Large Dense‐Cored Vesicles from Sympathetic Nerve

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Abstract
Highly purified noradrenergic, large, dense‐cored vesicles were isolated from bovine sympathetic nerve endings by sucrose‐D2O density gradient centrifugation. Their concentration of glycoprotein hexosamine and sialic acid was 6.6 and 3.9 μmol/100 mg lipid‐free dry weight, respectivly. values which are similar to those previously found in vine chromaffin granules. However, whereas chromaffin granule glycoproteins are characterized by their high proportion of N‐acetylgalactosamine‐containing O‐glycosidically‐linked oligosaccharides (present in the chromo‐granins), such oligosaccharides accounted for only 17% of those in noradrenergic synaptic vesicle glycoproteins. Fractionation of N‐3H‐acetylated glycopeptides by sequential lectin affinity chromatography demonstrated that approxi‐mtely two‐thirds of the oligosaccharides were of the triand tetraantennary complex type, accompanied by 14% biantennary oligosaccharides and 3% high‐mannose oligosaccharides. The vesicles had a relatively low concentration of chondroitin sulfate (less than 5% of that in chromaffin granules) but significant amounts of heparan sulfate (0.4 μmol N‐acetylglucosamine/100 mg lipid‐free dry weight). No hyaluronic acid was detected. The concentration of ganglioside sialic acid in the noradrenergic vesicles was approximately 1 μmol/100 mg lipid‐free dry weight, which is significantly higher than that of a crude membrane mixture from which the vesicles were prepared; the ratio of N‐acetyl‐to N‐glycolylneuraminic acid was 0.8. Several molecular species of gangliosides were detected by thin‐layer chromatography, but most of these did not exactly comigrate with bovine brain gangliosides. Cholera toxin binding indicated that approximately half or less of the gangliosides belong to the gangliotetraose series.