Human serum IgA1 is substituted with up to six O-glycans as shown by matrix assisted laser desorption ionisation time-of-flight mass spectrometry.
The micro-heterogeneity of human serum IgA1 results from variable O-glycan substitutions in the 'hinge region' of the molecule and this O-glycosylation may be altered in a number of medical conditions. This micro-heterogeneity has been monitored by analysis of IgA1-derived tryptic O-glycopeptides using matrix assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI-ToF-MS) analysis. With ammonium citrate-trihydroxyacetophenone matrix, individual compositional glycoforms have been baseline resolved in more than 70 samples and these spectra revealed for the first time that, in addition to expected substitution with 3,4 and 5 GalNAcs, a sixth GalNAc substitution was also present in the hinge region of the molecule. The spectra obtained from subsequent exoglycosidase-treated samples confirmed hexa-O-substitution. Following endoprotease digestions of the exoglycosidase treated samples, possible locations for the sixth GalNAc were indicated from further MALDI-ToF-MS analysis. Hexa-substitution accounts for around 5-10% the glycoforms. This is, we believe, the first report of hexa-O-substitution with GalNAc of human serum IgA1.
Tarelli
Edward
E
Medical Biomics Centre, St. George's Hospital Medical School, Cranmer Terrace, London SW17 ORE, UK. etarelli@sghms.ac.uk
Smith
Alice C
AC
Hendry
Bruce M
BM
Challacombe
Stephen J
SJ
Pouria
Shideh
S
D000595:Amino Acid Sequence; D006020:Glycopeptides / Q000737:chemistry / Q000302:isolation & purification; D006026:Glycoside Hydrolases; D006801:Humans; D007070:Immunoglobulin A / Q000097:blood* / Q000737:chemistry* / Q000302:isolation & purification; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D010447:Peptide Hydrolases; D011134:Polysaccharides; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods; D014357:Trypsin
D006020:Glycopeptides; D007070:Immunoglobulin A; D010446:Peptide Fragments; D011134:Polysaccharides; D006026:Glycoside Hydrolases; D010447:Peptide Hydrolases; D014357:Trypsin