Carbohydrate structure of a thrombin‐like serine protease from Agkistrodon rhodostoma

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Abstract
The carbohydrate side chains of the thrombin-like serine protease ancrod from the venom of the Malayan pit viper Agkistrodon rhodostoma were liberated from tryptic glycopeptides by treatment with peptide-N4-(N-acetyl-β-glucosaminyl)asparagine amidase F and fractionated by high-performance liquid chromatography. Glycans obtained were characterized by digestion with exoglycosidases, methylation analysis and, in part, by liquid secondary-ion mass spectrometry and 1H-NMR spectroscopy. The results reveal that this snake venom glycoprotein contains partially truncated di-, tri- and tetraantennary complex type N-glycans carrying Fuc(α1-6) residues at the innermost N- acetylglucosamine and solely (α2-3)-linked sialic acid substituents. As a characteristic feature, ancrod oligosaccharides comprise mainly sialylated Galβ3GlcNAcβ lactosamine antennae. Furthermore, a small proportion of the sugar chains were found to carry a NeuAcα3GalNAcβ4GlcNAcβ antenna exclusively linked to C-2 of Man(α1-3) residues of the pentasaccharide core. Thus, many of the glycans found represent novel glycoprotein-N-glycan structures.