NR AFCO
AU Haraguchi,T.; Fisher,S.; Olofsson,S.; Endo,T.; Groth,D.; Tarentino,A.; Borchelt,D.R.; Teplow,D.; Hood,L.; Burlingame,A.L.; Lycke,E.; Kobata,A.; Prusiner,S.B.
TI Asparagine-linked glycosylation of the scrapie and cellular prion proteins
QU Archives of Biochemistry and Biophysics 1989 Oct; 274(1): 1-13
PT journal article
AB Post-translational modification of the scrapie prion protein (PrP) is thought to account for the unusual features of this protein. Molecular cloning of a PrP cDNA identified two potential Asn-linked glycosylation sites. Both the scrapie (PrPsc) and cellular (PrPc) isoforms were susceptible to digestion by peptide N-glycosidase F (PNGase F) but resistant to endoglycosidase H as measured by migration in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. PNGase F digestion of PrPc yielded two proteins of Mr26K and 28K; however, the 26-k species was only a minor component. In contrast, PNGase F digestion of PrPsc yielded equimolar amounts of two proteins of Mr26K and 28K. The significance of this altered stoichiometry between the 26- and 28-kDa deglycosylated forms of PrP during scrapie infection remains to be established. Both isoforms as well as PrP 27-30, which is produced by limited proteolysis of PrPsc, exhibited a reduced number of charge isomers after PNGase F digestion. The molecular weight of PrP 27-30 was reduced from 27K-30K by PNGase F digestion to 20K-22K while anhydrous hydrogen fluoride or trifluoromethanesulfonic acid treatment reduced the molecular weight to 19K-21K and 20K-22K, respectively. Denatured PrP 27-30 was radioiodinated and then assessed for its binding to lectin columns. PrP 27-30 was bound to wheat germ agglutinin (WGA) or lentil lectins and eluted with N-acetylglucosamine or alpha-methyl-mannoside, respectively. Digestion of PrP 27-30 with sialidase prevented its binding to WGA but enhanced its binding to Ricinus communis lectin. These findings argue that PrP 27-30 probably possesses Asn-linked, complex oligosaccharides with terminal sialic acids, penultimate galactoses, and fucose residues attached to the innermost N-acetyl-glucosamine. Whether differences in Asn-linked oligosaccharide structure between PrPc and PrPsc exist and are responsible for the distinct properties displayed by these two isoforms remain to be established.
MH Animal; *Asparagine; Brain/metabolism/microbiology; Cloning, Molecular; DNA, Viral/genetics; Electrophoresis, Gel, Two-Dimensional; Electrophoresis, Polyacrylamide Gel; Glycoside Hydrolases; Glycosylation; Hamsters; Microsomes/metabolism/microbiology; Prions/genetics/*metabolism; *Protein Processing, Post-Translational; Support, Non-U.S. Gov't; Support, U.S. Gov't, P.H.S.; Viral Proteins/*genetics/isolation & purification
AD David R. Borchelt, Darlene Groth, Tokuko Haraguchi, Department of Neurology, University of California, San Francisco, California 94143, U.S.A.; Susan Fisher, Departments of Biochemistry and Biophysics, University of California, San Francisco, California 94143, U.S.A.; Alma Burlingame, Department of Stomatology, University of California, San Francisco, California 94143, U.S.A.; Stanley B. Prusiner, Department of Pharmaceutical Chemistry, University of California, San Francisco, California 94143, U.S.A.; Leroy Hood, Department of Anatomy, University of California, San Francisco, California 94143, U.S.A.; Erik Lycke, Sigvard Olofsson, Department of Virology, University of Goteborg, 41346, Goteborg, Sweden; Tamao Endo, Akira Kobata, Department of Biochemistry, Institute of Medical Science, University of Tokyo, Tokyo 108, Japan; Anthony Tarentino, New York State Department of Health, Albany, New York 12201, U.S.A.; David Teplow, Division of Biology, California Institute of Technology, Pasadena, California 91125, U.S.A.
SP englisch
PO USA