N-Glycosylation of the MUC1 mucin in epithelial cells and secretions.
The MUC1 mucin is an important tumor-associated antigen that shows extensive glycosylation in vivo. The O-glycosylation of this molecule, which has been well characterized in many cell types and tissues, is important in conferring the unusual biochemical and biophysical properties on a mucin. N-Glycosylation is crucial to the folding, sorting, membrane trafficking, and secretion of many proteins. Here, we evaluated the N-glycosylation of MUC1 derived from two sources: endogenous MUC1 isolated from human milk and a recombinant epitope-tagged MUC1F overexpressed in Caco2 colon carcinoma cells. N-Glycans on purified MUC1F/MUC1 were analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS), gas chromatography-mass spectrometry (GC-MS), and CAD-ESI-MS/MS. The spectra indicate that MUC1F N-glycans have compositions consistent with high-mannose structures (Hex(5-9)HexNAc(2)) and complex/hybrid-type glycans (NeuAc(0-3)Fuc(0-3)Hex(3-8)HexNAc(3-7)). Many of the N-glycan structures are identical on MUC1F and native MUC1; however, a marked difference is seen between the N-glycans on membrane-bound and secreted forms of the native molecule.
Parry
Simon
S
Division of Molecular Biosciences, Imperial College London, UK.
Hanisch
Franz Georg
FG
Leir
Shih-Hsing
SH
Sutton-Smith
Mark
M
Morris
Howard R
HR
Dell
Anne
A
Harris
Ann
A
D018938:Caco-2 Cells; D002240:Carbohydrate Sequence; D004847:Epithelial Cells / Q000378:metabolism*; D008401:Gas Chromatography-Mass Spectrometry; D006031:Glycosylation; D006801:Humans; D008358:Mannose / Q000737:chemistry; D008895:Milk, Human / Q000737:chemistry; D008969:Molecular Sequence Data; D018396:Mucin-1 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D009237:N-Acetyllactosamine Synthase / Q000737:chemistry; D011134:Polysaccharides / Q000032:analysis*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D021241:Spectrometry, Mass, Electrospray Ionization; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D001616:beta-Galactosidase / Q000737:chemistry
D018396:Mucin-1; D011134:Polysaccharides; D011994:Recombinant Proteins; D009237:N-Acetyllactosamine Synthase; D001616:beta-Galactosidase; D008358:Mannose