Influence of the amino acid sequence on the MUC5AC motif peptide O-glycosylation by human gastric UDP-GalNAc:polypeptide N-acetylgalactosaminyltransferase(s).
The present work was carried out to study the role of the peptide moiety in the addition of O-linked N-acetylgalactosamineto human apomucin using human crude microsomal homogenates from gastric mucosa (as enzyme source) and a series of peptide acceptors representative of tandem repeat domains deduced from the MUC5AC mucin gene (expressed in the gastric mucosa). Being rich in threonine and serine placed in clusters, these peptides provided several potential sites for O-glycosylation. The glycosylated products were analysed by a combination of electrospray mass spectrometry and capillary electrophoresis in order to isolate the glycopeptides and to determine their sequence by Edman degradation. The O-glycosylation of our MUC5AC motif peptides gave information on the specificity and activity of the gastric microsomal UDP-N-acetylgalactosamine:polypeptide N-acetylgalactosaminyltransferase(s). The proline residues and the induced-conformations are of great importance for the recognition of MUC5AC peptides but they are not the only factors for the choice of the O-glycosylation sites. Moreover, for the di-glycosylated peptides, the flanking regions of the proline residues strongly influence the site of the second O-glycosylation.
Hennebicq
S
S
Unité INSERM N 377, Lille, France.
Tetaert
D
D
Soudan
B
B
Boersma
A
A
Briand
G
G
Richet
C
C
Gagnon
J
J
Degand
P
P
D000595:Amino Acid Sequence; D001665:Binding Sites; D005753:Gastric Mucosa / Q000201:enzymology; D006031:Glycosylation; D006801:Humans; D066298:In Vitro Techniques; D008861:Microsomes / Q000201:enzymology; D008969:Molecular Sequence Data; D055271:Mucin 5AC; D009077:Mucins / Q000737:chemistry* / Q000378:metabolism*; D017350:N-Acetylgalactosaminyltransferases / Q000378:metabolism*; D009842:Oligopeptides / Q000737:chemistry / Q000378:metabolism; D011487:Protein Conformation; D013379:Substrate Specificity; D000097763:Polypeptide N-acetylgalactosaminyltransferase
D055271:Mucin 5AC; D009077:Mucins; D017350:N-Acetylgalactosaminyltransferases; D009842:Oligopeptides; D000097763:Polypeptide N-acetylgalactosaminyltransferase; C524678:MUC5AC protein, human