title: Identification of a human gastric mucin precursor: N-linked glycosylation and oligomerization.
abstract: Gastric mucin plays an important role in the protection of the stomach wall from chemical, microbiological and mechanical damage. We have previously isolated human gastric mucus glycoproteins and raised a polyclonal antiserum against these macromolecules. This antiserum specifically reacted with gastric mucins in immunoblotting experiments and stained mucous granules at the apical side of gastric surface epithelial cells. A similar staining pattern was obtained after incubation with an antiserum against rat gastric mucin. Next we used the antiserum in pulse-chase experiments of human stomach tissue explants. After short labelling periods with [35S]methionine and [35S]cysteine, the antiserum reacted with a polypeptide with an apparent molecular mass of approx. 500 kDa as determined by SDS/PAGE, which was converted after 90 min into a heterogeneous high-molecular-mass glycoprotein. This high-molecular-mass form, but not the 500 kDa polypeptide, was detectable in the culture medium after 2 h. This strongly suggests that the 500 kDa polypeptide is the precursor of the purified gastric mucin. Analysis of pulse-chase experiments by non-reducing SDS/PAGE revealed that the precursors form disulphide-linked oligomers early in biosynthesis, before the addition of O-linked sugars. After preincubation with the N-glycosylation inhibitor, tunicamycin, the apparent molecular mass of the precursor decreased marginally but consistently, indicating that N-linked glycan chains are present on the mucin precursor.
authors.affiliation: Laboratory of Cell Biology, University of Utrecht, The Netherlands.
authors.forename: L W
authors.initials: LW
authors.lastname: Klomp
authors.forename: L
authors.initials: L
authors.lastname: van Rens
authors.forename: G J
authors.initials: GJ
authors.lastname: Strous
mesh_terms: D005260:Female; D005748:Gastric Fundus / Q000737:chemistry / Q000378:metabolism; D005752:Gastric Mucins / Q000096:biosynthesis*; D005753:Gastric Mucosa / Q000378:metabolism*; D006031:Glycosylation; D006801:Humans; D007120:Immunochemistry; D008297:Male; D011233:Precipitin Tests; D011498:Protein Precursors / Q000032:analysis / Q000378:metabolism*; D013270:Stomach / Q000737:chemistry
chemical_list: D005752:Gastric Mucins; D011498:Protein Precursors