Oligosaccharide structure of a functional unit RvH1-b of Rapana venosa hemocyanin using HPLC/electrospray ionization mass spectrometry |
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Authors: | Beck Alexander Hillen Nina Dolashki Alexandar Stevanovic Stefan Salvato Benedetto Voelter Wolfgang Dolashka-Angelova Pavlina |
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Institution: | 1. Klinisch-chemisches Zentrallaboratorium der Universitätskliniken, Abteilung Innere Medizin IV, Universität Tübingen, Otfried-Müller-Straße 10, D-72076 Tübingen, Germany;2. Interfakultäres Institut für Zellbiologie, Abteilung Immunologie, Universität Tübingen, Auf der Morgenstelle 15, D-72076 Tübingen, Germany;3. Interfakultäres Institut für Biochemie, Universität Tübingen, Hoppe-Seyler-Straße 4, D-72076 Tübingen, Germany;4. Department of Biology and CNR Institute for Biomedical Technologies, Section of Padova, University of Padova, Via Ugo Bassi 58/B, I-35131 Padova, Italy;5. Institute of Organic Chemistry, Bulgarian Academy of Sciences, G. Bonchev 9, Sofia 1113, Bulgaria |
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Abstract: | In the present study the structures of two glycopeptides (G1 and G1'), isolated from FU RvH(1)-b and two glycopeptides (G2 and G3), isolated from the structural subunit RvH(1) of Rapana venosa hemocyanin, were determined. To structurally characterize the site-specific carbohydrate heterogeneity and binding site of the N-linked glycopeptide(s), a combination of capillary reversed-phase chromatography and ion trap mass spectrometry was used. The amino acid sequences of glycopeptides G1 and G1' determined by Edman degradation and MS/MS sequencing demonstrated that the oligosaccharides are linked to N-glycosylation sites. Two peptides (a glycosylated (G1) and non-glycosylated one) were identified in this fraction and no linkage sites were observed in the latter one. Based on the sequencing of the glycosylated fractions G1, G1', G2 and G3, the carbohydrate structure Man(alpha1-6)Man(alpha1-3)Man(beta1-4)GlcNAc(beta1-4)Fuc(alpha1-6)]GlcNAc-R could be identified for glycopeptides G1 and G3, and only the typical core structure Man(alpha1-6)Man(alpha1-3)Man(beta1-4)GlcNAc(beta1-4)GlcNAc-R was found for G1' and G2. The Fuc residue found in glycopeptides G1 and G3 is attached to N-acetyl-glucosamine of the carbohydrate core, as often found in other glycoproteins. |
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Keywords: | Glycopeptides HPLC/electrospray ionization mass spectrometry Oligosaccharides Rapana venosa hemocyanin |
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