RRC ID 53858
Author Nakakita S, Itoh A, Nakakita Y, Nonaka Y, Ogawa T, Nakamura T, Nishi N.
Title Cooperative Interactions of Oligosaccharide and Peptide Moieties of a Glycopeptide Derived from IgE with Galectin-9.
Journal J Biol Chem
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.
Volume 291(2)
Pages 968-79
Published 2016-1-8
DOI 10.1074/jbc.M115.694448
PII S0021-9258(20)36227-X
PMID 26582205
PMC PMC4705414
MeSH Animals Antibodies, Monoclonal / metabolism Cell Degranulation Cell Line Chromatography, High Pressure Liquid Chromatography, Reverse-Phase Electrophoresis, Polyacrylamide Gel Enzyme Activation Galectins / metabolism* Glycopeptides / isolation & purification* Glycopeptides / metabolism Humans Immunoglobulin E / metabolism* Mice Mitogen-Activated Protein Kinases / metabolism Oligosaccharides / metabolism* Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase / metabolism Polysaccharides / chemistry Polysaccharides / metabolism Protein Binding Rats Serine Endopeptidases / metabolism Serum Albumin, Bovine / metabolism Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization Trypsin / metabolism beta-N-Acetylhexosaminidases / metabolism
IF 4.238
Times Cited 6
DNA material mIGEH (RDB15983) mIGEL (RDB15984)
Human and Animal Cells RBL 2H3(RCB2782)