Department of Biochemistry, Institute of Medical Science, University of Tokyo, Minato-ku, Tokyo 108, Japan, and Departments of Neurology and Biochemistry and Biophysics, University of California, San Francisco, California 94143-0518 Received March 30, 1989; Revised Manuscript Received June 13, 1989 abstract: Prion proteins from humans and rodents contain two consensus sites for asparagine-linked glycosylation near their C-termini. The asparagine-linked oligosaccharides of the scrapie isoform of the hamster prion protein (PrP 27-30) were released quantitatively from the purified molecule by hydrazinolysis followed by N-acetylation and NaB3H4 reduction. The radioactive oligosaccharides were fractionated into one neutral and three acidic oligosaccharide fractions by anion-exchange column chromatography. All oligosaccharides in the acidic fractions could be converted to neutral oligosaccharides by sialidase digestion. Structural studies on these oligosaccharides including sequentialexoglycosidase digestion in combination with methylation analysis revealed that PrP 27-30 contains a mixture of bi-, tri-, and tetraantennary complex-type sugar chains withManal-* 6 (GlcNAc| 81-* 4)(Manal^-3) Manj31-,-4GlcNAC| 81-> 4-(Fucal-* 6) GlcNAc as their core. Variation is produced by the different combination of the oligosaccharides Gal/31—4GlcNAc/31—, Gal/? 1—4 (Fucal^ 3) GlcNAc01^, GlcNAqSl-^, Siaa2-* 3Gal/íl-* 4GlcNAc01—, and Siaa2-* 6Gal/31-*-4GlcN Ac/? 1 in their outer chain moieties. When both asparagine-linked consensus sites are glycosylated, the diversity of oligosaccharide structures yields over 400 different forms of the scrapie prion protein. Whether these diverse asparagine-linked oligosaccharides participate in scrapie prion infectivity or modify the function of the cellular prion protein remains to be established.

Scrapie is a degenerative neurological disease of sheep and goats, which can be transmitted to laboratory rodents (Pru-siner, 1987). Because the unusual properties of the scrapie agent distinguish it from viruses, the term prion was intro-duced (Prusiner, 1982). Three human diseases may also be caused by prions: Creutzfeldt-Jakob disease, kuru, and Gerstmann-Straussler syndrome (Gajdusek, 1977; Masters et al., 1981). Purification of scrapie prion infectivity led to isolation of a protein with relative molecular weight of 27000-30000 (Bolton et al., 1982; Prusiner et al., 1982). This protein was designated prion protein (PrP) 27-30.1 PrP cDNA clones were isolated from libraries that were constructed by using poly (A+) RNA from scrapie-infected