A comparative study of the asparagine-linked oligosaccharides on siglec-5, siglec-7 and siglec-8, expressed in a CHO cell line, and their contribution to ligand recognition.
The siglecs (sialic acid-binding immunoglobulin-like lectins) mediate sialic acid-dependent cellular interactions and may in some cases signal through SH2-binding domains. In addition to the previously characterized siglecs, sialoadhesin, CD22, CD33 and myelin-associated glycoprotein, several new ones, siglec-5, siglec-7 and siglec-8, have recently been cloned. Although these novel receptors have generated considerable interest as therapeutic targets because of their expression pattern on immune cells, very little is known about how their lectin activity is regulated. Previous studies with sialoadhesin, CD22 and CD33 have shown that siglec glycosylation has significant effects on binding. To determine any differences in the glycan composition of siglec-5, siglec-7 and siglec-8 that may modify their function, we released and characterized the N-linked oligosaccharide distribution in these three glycoproteins. The glycan pools from siglec-5 and siglec-7 contained a larger proportion of sialylated and core-fucosylated biantennary, triantennary and tetra-antennary oligosaccharides, whereas the carbohydrate mixture released from siglec-8 is noticeably less sialylated and is more abundant in 'high-mannose'-type glycans. In addition, we show that, in contrast with CD22 and CD33, mutating the conserved potentially N-linked glycosylation site in the first domain has no effect on binding mediated by siglec-5 or siglec-7.
Freeman
S
S
University of Oxford, Institute of Molecular Medicine, John Radcliffe Hospital, Headington, Oxford, UK.
Birrell
H C
HC
D'Alessio
K
K
Erickson-Miller
C
C
Kikly
K
K
Camilleri
P
P
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D015703:Antigens, CD / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000944:Antigens, Differentiation, B-Lymphocyte / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015214:Antigens, Differentiation, Myelomonocytic / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001216:Asparagine / Q000378:metabolism*; D016466:CHO Cells; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002460:Cell Line; D006224:Cricetinae; D004912:Erythrocytes / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D037102:Lectins; D008024:Ligands; D013058:Mass Spectrometry; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed / Q000235:genetics; D019158:N-Acetylneuraminic Acid / Q000378:metabolism; D009439:Neuraminidase / Q000378:metabolism; D009844:Oligosaccharides / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D016415:Sequence Alignment; D005644:alpha-L-Fucosidase / Q000378:metabolism
D015703:Antigens, CD; D000944:Antigens, Differentiation, B-Lymphocyte; D015214:Antigens, Differentiation, Myelomonocytic; D037102:Lectins; D008024:Ligands; D008562:Membrane Glycoproteins; D009844:Oligosaccharides; C114531:SIGLEC5 protein, human; C402206:SIGLEC7 protein, human; C403967:SIGLEC8 protein, human; D001216:Asparagine; D009439:Neuraminidase; D005644:alpha-L-Fucosidase; D019158:N-Acetylneuraminic Acid