title: Acquisition of protease resistance by prion proteins in scrapie-infected cells does not require asparagine-linked glycosylation.
abstract: The scrapie and cellular isoforms of the prion protein (PrPSc and PrPC) differ strikingly in a number of their biochemical and metabolic properties. The structural features underlying these differences are unknown, but they are thought to result from a posttranslational process. Both PrP isoforms contain complex type oligosaccharides, raising the possibility that differences in the asparagine-linked glycosylation account for the properties that distinguish PrPC and PrPSc. ScN2a and ScHaB cells in culture produce several PrP molecules with relative molecular masses of 26-35 kDa and proteinase K-resistant cores of 19-29 kDa. When the cells were treated with tunicamycin, this heterogeneity was eliminated and a single PrP species of 26 kDa was observed. Several hours after its synthesis, a fraction of this protein became insoluble in detergents and acquired a proteinase K-resistant core, thus displaying two of the biochemical hallmarks of PrPSc. Synthesis in the presence of tunicamycin restricted the proteinase K-resistant cores of PrP to a single species of 19 kDa. No proteinase K-resistant PrP was found in uninfected cells. Expression of a mutated PrP gene lacking both asparagine-linked glycosylation sites in ScN2a cells resulted in the synthesis of 19-kDa proteinase K-resistant PrP molecules. We conclude that asparagine-linked glycosylation is not essential for the synthesis of proteinase K-resistant PrP and that structural differences unrelated to asparagine-linked oligosaccharides must exist between PrPC and PrPSc. Whether unglycosylated PrPSc molecules are associated with scrapie prion infectivity remains to be established.
authors.affiliation: Department of Neurology, University of California, San Francisco 94143.
authors.forename: A
authors.initials: A
authors.lastname: Taraboulos
authors.forename: M
authors.initials: M
authors.lastname: Rogers
authors.forename: D R
authors.initials: DR
authors.lastname: Borchelt
authors.forename: M P
authors.initials: MP
authors.lastname: McKinley
authors.forename: M
authors.initials: M
authors.lastname: Scott
authors.forename: D
authors.initials: D
authors.lastname: Serban
authors.forename: S B
authors.initials: SB
authors.lastname: Prusiner
mesh_terms: D000595:Amino Acid Sequence; D001216:Asparagine; D002472:Cell Transformation, Viral; D019286:Endopeptidase K; D006031:Glycosylation; D007700:Kinetics; D008969:Molecular Sequence Data; D018620:PrPSc Proteins; D011328:Prions / Q000235:genetics*; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D012697:Serine Endopeptidases / Q000378:metabolism*; D014415:Tunicamycin / Q000494:pharmacology; D014764:Viral Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism
chemical_list: D018620:PrPSc Proteins; D011328:Prions; D014764:Viral Proteins; D014415:Tunicamycin; D001216:Asparagine; D012697:Serine Endopeptidases; D019286:Endopeptidase K