Separation and properties of cellular and scrapie prion proteins.

Abstract

Purified preparations of scrapie prions contain a sialoglycoprotein of Mr 27,000-30,000, designated PrP 27-30, which is derived from the scrapie prion protein [Mr,33,000 Source of Scrapie Prions and Bioassay. A hamster-adapted isolate of the scrapie agent was passaged and prepared as described (1, 13).Preparation of the Subcellular Fractions. Weanling hamsters (LVG/LAK) were inoculated intracerebrally with 107 ID50 units of the scrapie agent. The brains were collected from hamsters sacrificed 60 days after infection and from age-matched uninfected animals. The brains were suspended in 0.32 M sucrose (10%, wt/vol) and homogenized with six bursts of 10 sec each by using a Polytron homogenizer set at medium speed. The homogenates were centrifuged in a Beckman 50.2 Ti rotor. Pellets were resuspended in 0.32 M sucrose solution and the volumes were adjusted to that of the supernatant. All solutions were kept on ice and all centrifugations were performed at 4°C. The fractions were adjusted to 10 mg of protein per ml with 0.32 M sucrose solutions. One-milliliter samples were centrifuged in the Beckman 50 Ti rotor at 38,000 rpm for 1 hr at 4°C. Pellets were resuspended and treated as described in Results. For digestion experiments, N-lauroylsarcosine (sarkosyl) was added from a 10% stock solution in 25 mM Tris'HCl/0.1 M NaCl, pH 7.4, to some of the samples. Control samples were diluted by the same amount of Tris buffer. Digestions were initiated by addition of an aliquot of proteinase K (2 mg/ml) in Tris buffer to give a final concentration of 0.1 mg/ml. After 30 min at room temperature, the digestions were terminated by addition of an aliquot of 0

pH-dependent Stability and Conformation of the Recombinant Human Prion Protein PrP(90–231)

“…1 The transition between the cellular form of PrP, designated PrP C , and PrP res occurs by a post-translational mechanism and appears to take place without any detectable covalent modifications to the protein molecule (7). One of the main characteristics distinguishing PrP C from PrP res is the resistance of the latter to proteolytic digestion (8,9). Furthermore, recent spectroscopic studies indicate that the two isoforms have profoundly different conformation: while the secondary structure of PrP C consists largely of ␣-helices (10), PrP res appears to be rich in ␤-sheet structure (10 -13).…”

Bovine prion protein as a modulator of protein kinase CK2

“…Prions are composed mainly of a malformed protein (PrP Sc ) generated by a post-translational misfolding of a constitutive glycoprotein, cellular PrP (PrP c ) [22][23][24], found particularly in the central nervous system. Prion infectivity is explained by the capacity of PrP Sc to convert PrP c into the pathogenic conformation through an essentially autocatalytic mechanism initiated by PrP Sc -PrP c interaction [25].…”

Risikofaktoren der sporadischen Creutzfeldt-Jakob-Krankheit

“…Während PrP C durch die Proteinase K abgebaut werden kann, ist PrP Sc zum Teil Proteinase-resistent (Oesch et al 1985). Weiterhin ist PrP C, im Gegensatz zu der pathologischen Form, in nicht-denaturierenden Lösungsmitteln löslich (Meyer et al 1986). …”