The asparagine-linked carbohydrate of honeybee venom hyaluronidase

Hyaluronidase from the venom of the honeybee (Apis mellifera) has been purified by gelpermeation and cation exchange chromatography. Its asparagine-linked carbohydrate chains were released from tryptic glycopeptides with N-glycosidase A and reductively aminated with 2-aminopyridine. Separation of th...

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Bibliographic Details
Published in:Glycoconjugate journal 1995-02, Vol.12 (1), p.77-83
Main Authors: Kubelka, V, Altmann, F, März, L
Format: Article
Language:English
Subjects:
Aminopyridines - chemistry
Animals
Asparagine - analysis
Asparagine - chemistry
Bee Venoms - enzymology
Carbohydrate Sequence
Cattle
Chemical Fractionation
Chromatography, High Pressure Liquid
Chromatography, Ion Exchange - methods
Hyaluronoglucosaminidase - chemistry
Hyaluronoglucosaminidase - isolation & purification
Hyaluronoglucosaminidase - metabolism
Mannosidases - chemistry
Mannosidases - metabolism
Molecular Sequence Data
Molecular Structure
Oligosaccharides - chemistry
Oligosaccharides - metabolism
Polysaccharides - chemistry
Sialic Acids - chemistry
Trypsin - chemistry
Trypsin - metabolism
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Summary:Hyaluronidase from the venom of the honeybee (Apis mellifera) has been purified by gelpermeation and cation exchange chromatography. Its asparagine-linked carbohydrate chains were released from tryptic glycopeptides with N-glycosidase A and reductively aminated with 2-aminopyridine. Separation of the fluorescent derivatives by size-fractionation and reversed-phase HPLC afforded eighteen fractions which were analysed by two-dimensional HPLC mapping combined with exoglycosidase digestions. The bulk of the N-linked glycans of hyaluronidase consisted of small oligosaccharides (Man1-3GlcNAc2), most of which were either alpha 1,3-monofucosylated or alpha 1,3-(alpha 1,6-)difucosylated at the innermost GlcNAc residue. High-mannose type structures constituted the minor fractions, together making up about 5% of the oligosaccharide pool from hyaluronidase. Four fractions, making up 8% of the N-linked glycans, contained the terminal trisaccharide GalNAc beta 1-4[Fuc alpha 1-3]GlcNAc beta 1- in beta 1,2-linkage to the core alpha 1,3-mannosyl residue. No evidence for the presence of O-glycans or sialic acids could be found.
ISSN:0282-0080
1573-4986
DOI:10.1007/bf00731872