Glycopeptidome of a heavily N-glycosylated cell surface glycoprotein of Dictyostelium implicated in cell adhesion.
Genetic analysis has implicated the cell surface glycoprotein gp130 in cell interactions of the social amoeba Dictyostelium, and information about the utilization of the 18 N-glycosylation sequons present in gp130 is needed to identify critical molecular determinants of its activity. Various glycomics strategies, including mass spectrometry of native and derivatized glycans, monosaccharide analysis, exoglycosidase digestion, and antibody binding, were applied to characterize a nonanchored version secreted from Dictyostelium. s-gp130 is modified by a predominant Man(8)GlcNAc(4) species containing bisecting and intersecting GlcNAc residues and additional high-mannose N-glycans substituted with sulfate, methyl-phosphate, and/or core alpha 3-fucose. Site mapping confirmed the occupancy of 15 sequons, some variably, and glycopeptide analysis confirmed 14 sites and revealed extensive heterogeneity at most sites. Glycopeptide glycoforms ranged from Man(6) to Man(9), GlcNAc(0-2) (peripheral), Fuc(0-2) (including core alpha 3 and peripheral), (SO(4))(0-1), and (MePO(4))(0-1), which represented elements of virtually the entire known cellular N-glycome as inferred from prior metabolic labeling and mass spectrometry studies. gp130, and a family of 14 related predicted glycoproteins whose polypeptide sequences are rapidly diverging in the Dictyostelium lineage, may contribute a functionally important shroud of high-mannose N-glycans at the interface of the amoebae with each other, their predators and prey, and the soil environment.
Feasley
Christa L
CL
Department of Biochemistry & Molecular Biology and Oklahoma Center for Medical Glycobiology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104, USA.
Johnson
Jennifer M
JM
West
Christopher M
CM
Chia
Catherine P
CP
D000595:Amino Acid Sequence; D001483:Base Sequence; D002448:Cell Adhesion; D002851:Chromatography, High Pressure Liquid; D004023:Dictyostelium / Q000235:genetics*; D004591:Electrophoresis, Polyacrylamide Gel; D054794:Glycomics; D006020:Glycopeptides / Q000737:chemistry* / Q000378:metabolism; D006031:Glycosylation; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D011134:Polysaccharides; D015800:Protozoan Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D053719:Tandem Mass Spectrometry
D006020:Glycopeptides; D008562:Membrane Glycoproteins; D011134:Polysaccharides; D015800:Protozoan Proteins