title: Cooperative Interactions of Oligosaccharide and Peptide Moieties of a Glycopeptide Derived from IgE with Galectin-9.
abstract: We previously showed that galectin-9 suppresses degranulation of mast cells through protein-glycan interaction with IgE. To elucidate the mechanism of the interaction in detail, we focused on identification and structural analysis of IgE glycans responsible for the galectin-9-induced suppression using mouse monoclonal IgE (TIB-141). TIB-141 in combination with the antigen induced degranulation of RBL-2H3 cells, which was almost completely inhibited by human and mouse galectin-9. Sequential digestion of TIB-141 with lysyl endopeptidase and trypsin resulted in the identification of a glycopeptide (H-Lys13-Try3; 48 amino acid residues) with a single N-linked oligosaccharide near the N terminus capable of neutralizing the effect of galectin-9 and another glycopeptide with two N-linked oligosaccharides (H-Lys13-Try1; 16 amino acid residues) having lower activity. Enzymatic elimination of the oligosaccharide chain from H-Lys13-Try3 and H-Lys13-Try1 completely abolished the activity. Removal of the C-terminal 38 amino acid residues of H-Lys13-Try3 with glutamyl endopeptidase, however, also resulted in loss of the activity. We determined the structures of N-linked oligosaccharides of H-Lys13-Try1. The galectin-9-binding fraction of pyridylaminated oligosaccharides contained asialo- and monosialylated bi/tri-antennary complex type oligosaccharides with a core fucose residue. The structures of the oligosaccharides were consistent with the sugar-binding specificity of galectin-9, whereas the nonbinding fraction contained monosialylated and disialylated biantennary complex type oligosaccharides with a core fucose residue. Although the oligosaccharides linked to H-Lys13-Try3 could not be fully characterized, these results indicate the possibility that cooperative binding of oligosaccharide and neighboring polypeptide structures of TIB-141 to galectin-9 affects the overall affinity and specificity of the IgE-lectin interaction.
authors.affiliation: From the Division of Functional Glycomics and.
authors.forename: Shin-Ichi
authors.initials: S
authors.lastname: Nakakita
authors.affiliation: Division of Research Instrument and Equipment, Life Science Research Center, and.
authors.forename: Aiko
authors.initials: A
authors.lastname: Itoh
authors.affiliation: From the Division of Functional Glycomics and.
authors.forename: Yukari
authors.initials: Y
authors.lastname: Nakakita
authors.affiliation: the Department of Endocrinology, Faculty of Medicine, Kagawa University, 1750-1 Ikenobe, Miki-cho, Kita-gun, Kagawa 761-0793, Japan.
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Nonaka
authors.affiliation: the Department of Endocrinology, Faculty of Medicine, Kagawa University, 1750-1 Ikenobe, Miki-cho, Kita-gun, Kagawa 761-0793, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Ogawa
authors.affiliation: the Department of Endocrinology, Faculty of Medicine, Kagawa University, 1750-1 Ikenobe, Miki-cho, Kita-gun, Kagawa 761-0793, Japan.
authors.forename: Takanori
authors.initials: T
authors.lastname: Nakamura
authors.affiliation: Division of Research Instrument and Equipment, Life Science Research Center, and nnishi@med.kagawa-u.ac.jp.
authors.forename: Nozomu
authors.initials: N
authors.lastname: Nishi
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism; D015550:Cell Degranulation; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D056148:Chromatography, Reverse-Phase; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D037161:Galectins / Q000378:metabolism*; D006020:Glycopeptides / Q000302:isolation & purification* / Q000378:metabolism; D006801:Humans; D007073:Immunoglobulin E / Q000378:metabolism*; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009844:Oligosaccharides / Q000378:metabolism*; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase / Q000378:metabolism; D011134:Polysaccharides / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D051381:Rats; D012697:Serine Endopeptidases / Q000378:metabolism; D012710:Serum Albumin, Bovine / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014357:Trypsin / Q000378:metabolism; D001619:beta-N-Acetylhexosaminidases / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D037161:Galectins; D006020:Glycopeptides; C105032:LGALS9 protein, human; D009844:Oligosaccharides; D011134:Polysaccharides; C508186:galectin 9, mouse; C038601:trinitrophenyl-bovine serum albumin; D012710:Serum Albumin, Bovine; D007073:Immunoglobulin E; D020928:Mitogen-Activated Protein Kinases; D001619:beta-N-Acetylhexosaminidases; D012697:Serine Endopeptidases; C028344:glutamyl endopeptidase; D014357:Trypsin; C036813:lysyl endopeptidase; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
keywords: allergy; galectin; immunoglobulin E (IgE); mast cell; oligosaccharide