Nuclear pore complex glycoproteins contain cytoplasmically disposed O-linked N-acetylglucosamine.
A novel form of protein-saccharide linkage consisting of single N-acetylglucosamine (GlcNAc) residues attached in O-linkages directly to the polypeptide backbone has been described (Holt, G. D., and G. W. Hart, 1986, J. Biol. Chem., 261:8049-8057). This modification was found on proteins distributed throughout the cell, although proteins bearing O-linked GlcNAc moieties were particularly abundant in the cytosolic and nuclear envelope fractions of rat liver. In the accompanying article (Snow, C. M., A. Senior, and L. Gerace, 1987, J. Cell. Biol., 104: 1143-1156), the authors describe monoclonal antibodies directed against eight proteins localized to the nuclear pore complex. These proteins occur on the cytoplasmic and nucleoplasmic (but not lumenal) sides of nuclear membranes. In this report, we demonstrate that all members of this group of pore complex proteins bear multiple O-linked GlcNAc residues. Further, we show that the O-linked GlcNAc moieties are linked via serine (and possibly threonine) side chains to these proteins. Perturbing the O-linked GlcNAc residues either by covalently attaching galactose to them or by releasing them with beta-N-acetylglucosaminidase strongly diminishes the immunoreactivity of the proteins with all of the monoclonal antibodies. However, the O-linked GlcNAc moieties are only part of the epitopes recognized, since O-GlcNAc-containing limit pronase fragments of nuclear pore complex proteins cannot be immunoprecipitated by these antibodies. These findings, taken together with those in the accompanying article, are a direct demonstration that proteins of the cytoplasm and nucleoplasm bear O-linked GlcNAc residues.
D000117:Acetylglucosamine / Q000032:analysis*; D000818:Animals; D003593:Cytoplasm / Q000032:analysis; D005944:Glucosamine / Q000031:analogs & derivatives*; D006023:Glycoproteins / Q000302:isolation & purification*; D008099:Liver / Q000032:analysis; D008565:Membrane Proteins / Q000302:isolation & purification*; D008970:Molecular Weight; D009685:Nuclear Envelope / Q000032:analysis*; D009844:Oligosaccharides; D051381:Rats
D006023:Glycoproteins; D008565:Membrane Proteins; D009844:Oligosaccharides; D005944:Glucosamine; D000117:Acetylglucosamine