title: Frontal affinity chromatography analysis of constructs of DC-SIGN, DC-SIGNR and LSECtin extend evidence for affinity to agalactosylated N-glycans.
abstract: Dendritic cell-specific intracellular adhesion molecule-3-grabbing nonintegrin (DC-SIGN) is a member of the C-type lectin family selectively expressed on immune-related cells. In the present study, we performed a systematic interaction analysis of DC-SIGN and its related receptors, DC-SIGN-related protein (DC-SIGNR) and liver and lymph node sinusoidal endothelial cell C-type lectin (LSECtin) using frontal affinity chromatography (FAC). Carbohydrate-recognition domains of the lectins, expressed as Fc-fusion chimeras, were immobilized to Protein A-Sepharose and subjected to quantitative FAC analysis using 157 pyridylaminated glycans. Both DC-SIGN-Fc and DC-SIGNR-Fc showed similar specificities for glycans containing terminal mannose and fucose, but great difference in affinity under the given experimental conditions. By contrast, LSECtin-Fc showed no affinity to these glycans. As a common feature, the DC-SIGN-related lectin-Fc chimeras, including LSECtin, exhibited binding affinity to mono- and/or bi-antennary agalactosylated N-glycans. The detailed FAC analysis further implied that the presence of terminal GlcNAc at the N-acetylglucosaminyltransferase I position is a key determinant for the binding of these lectins to agalactosylated N-glycans. By contrast, none of the lectins showed significant affinity to highly branched agalactosylated N-glycans. All of the lectins expressed on the cells were able to mediate cellular adhesion to agalactosylated cells and endocytosis of a model glycoprotein, agalactosylated α1-acid glycoprotein. In this context, we also identified three agalactosylated serum glycoproteins recognized by DC-SIGN-Fc (i.e. α-2-macroglobulin, serotransferrin and IgG heavy chain), by lectin blotting and MS analysis. Hence, we propose that 'agalactosylated N-glycans' are candidate ligands common to these lectins.
authors.affiliation: Research Center for Medical Glycoscience, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, Ibaraki, Japan.
authors.forename: Rikio
authors.initials: R
authors.lastname: Yabe
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Tateno
authors.forename: Jun
authors.initials: J
authors.lastname: Hirabayashi
mesh_terms: D000818:Animals; D016466:CHO Cells; D015815:Cell Adhesion Molecules / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D002846:Chromatography, Affinity / Q000379:methods; D002874:Chromosome Mapping; D002888:Chromosomes, Human, Pair 19; D006224:Cricetinae; D003412:Cricetulus; D003713:Dendritic Cells / Q000276:immunology / Q000502:physiology; D005260:Female; D006023:Glycoproteins / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D037102:Lectins / Q000276:immunology / Q000378:metabolism; D037181:Lectins, C-Type / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D008958:Models, Molecular; D010920:Placenta / Q000502:physiology; D011134:Polysaccharides / Q000737:chemistry / Q000378:metabolism*; D011247:Pregnancy; D011946:Receptors, Antigen / Q000737:chemistry / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity; D013601:T-Lymphocytes / Q000276:immunology; D000099215:DC-Specific ICAM-3 Grabbing Nonintegrin
chemical_list: D015815:Cell Adhesion Molecules; D006023:Glycoproteins; D037102:Lectins; D037181:Lectins, C-Type; D011134:Polysaccharides; D011946:Receptors, Antigen; D011956:Receptors, Cell Surface; D011993:Recombinant Fusion Proteins; D000099215:DC-Specific ICAM-3 Grabbing Nonintegrin