In vivo glycosylation of mucin tandem repeats.
The biochemical and biophysical properties of mucins are largely determined by extensive O-glycosylation of serine- and threonine-rich tandem repeat (TR) domains. In a number of human diseases aberrant O-glycosylation is associated with variations in the properties of the cell surface-associated and secreted mucins. To evaluate in vivo the O-glycosylation of mucin TR domains, we generated recombinant chimeric mucins with TR sequences from MUC2, MUC4, MUC5AC, or MUC5B, which were substituted for the native TRs of epitope-tagged MUC1 protein (MUC1F). These hybrid mucins were extensively O-glycosylated and showed the expected association with the cell surface and release into culture media. The presence of different TR domains within the chimeric mucins appears to have limited influence on their posttranslational processing. Alterations in glycosylation were detailed by fast atom bombardment mass spectrometry and reactivity with antibodies against particular blood-group and tumor-associated carbohydrate antigens. Future applications of these chimeras will include investigations of mucin posttranslational modification in the context of disease.
Silverman
H S
HS
Paediatric Molecular Genetics, Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Oxford, OX3 9DS, UK.
Parry
S
S
Sutton-Smith
M
M
Burdick
M D
MD
McDermott
K
K
Reid
C J
CJ
Batra
S K
SK
Morris
H R
HR
Hollingsworth
M A
MA
Dell
A
A
Harris
A
A
D000595:Amino Acid Sequence; D015295:Antigens, Tumor-Associated, Carbohydrate / Q000096:biosynthesis; D001789:Blood Group Antigens; D002240:Carbohydrate Sequence; D006031:Glycosylation; D006801:Humans; D008969:Molecular Sequence Data; D018396:Mucin-1 / Q000378:metabolism; D009077:Mucins / Q000235:genetics / Q000378:metabolism*; D009844:Oligosaccharides / Q000737:chemistry; D011499:Protein Processing, Post-Translational; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D020449:Repetitive Sequences, Amino Acid; D016339:Spectrometry, Mass, Fast Atom Bombardment
D015295:Antigens, Tumor-Associated, Carbohydrate; D001789:Blood Group Antigens; D018396:Mucin-1; D009077:Mucins; D009844:Oligosaccharides; D011993:Recombinant Fusion Proteins