Premium
Identification of substituted sites on MUC5AC mucin motif peptides after enzymatic O‐glycosylation combining β‐elimination and fixed‐charge derivatization
Author(s) -
Czeszak X.,
Ricart G.,
Tetaert D.,
Michalski J. C.,
Lemoine J.
Publication year - 2001
Publication title -
rapid communications in mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.528
H-Index - 136
eISSN - 1097-0231
pISSN - 0951-4198
DOI - 10.1002/rcm.532
Subject(s) - chemistry , glycosylation , ethanethiol , derivatization , tandem mass spectrometry , peptide , edman degradation , threonine , biochemistry , mass spectrometry , mucin , enzyme , chromatography , peptide sequence , organic chemistry , serine , gene
A strategy for determination of O‐glycosylation site(s) in glycopeptides has been developed using model compounds obtained by enzymatic glycosylation (by human GaNTase‐T2 isoform) on peptides derived from the human MUC5AC mucin tandem repeat motif. The β‐elimination‐addition reaction (using dimethylamine and concomitantly ethanethiol) on the formerly glycosylated sites through a Michael‐type condensation produced efficient deglycosylation with appropriate chemical modification. After N‐terminal derivatization by a phosphonium group, peptide sequencing was then carried out by nanospray tandem mass spectrometry experiments. The highly predictable fragmentation pathways of these fixed‐charge phosphonium derivatives enable straightforward recognition of glycosylation site(s) based on the mass increment of +44 Da for originally glycosylated threonine compared to the mass of fragments containing nonglycosylated residues. Copyright © 2001 John Wiley & Sons, Ltd.