NR ABLY
AU Bonetto,V.; Massignan,T.; Chiesa,R.; Morbin,M.; Mazzoleni,G.; Diomede,L.; Angeretti,N.; Colombo,L.; Forloni,G.; Tagliavini,F.; Salmona,M.
TI Synthetic miniprion PrP106
QU The Journal of Biological Chemistry 2002 Aug 30; 277(35): 31327-34
IA http://www.jbc.org/cgi/content/full/277/35/31327
PT journal article
AB Elucidation of structure and biological properties of the prion protein scrapie (PrPsc) is fundamental to an understanding of the mechanism of conformational transition of cellular (PrPc) into disease-specific isoforms and the pathogenesis of prion diseases. Unfortunately, the insolubility and heterogeneity of PrPsc have limited these studies. The observation that a construct of 106 amino acids (termed PrP106 or miniprion), derived from mouse PrP and containing two deletions (Delta 23-88, Delta 141-176), becomes protease-resistant when expressed in scrapie-infected neuroblastoma cells and sustains prion replication when expressed in PrP(0/0) mice prompted us to generate a corresponding synthetic peptide (sPrP106) to be used for biochemical and cell culture studies. sPrP106 was obtained successfully with a straightforward procedure, which combines classical stepwise solid phase synthesis with a purification strategy based on transient labeling with a lipophilic chromatographic probe. sPrP106 readily adopted a beta-sheet structure, aggregated into branched filamentous structures without ultrastructural and tinctorial properties of amyloid, exhibited a proteinase K-resistant domain spanning residues 134-217, was highly toxic to primary neuronal cultures, and induced a remarkable increase in membrane microviscosity. These features are central properties of PrPsc and make sPrP106 an excellent tool for investigating the molecular basis of the conformational conversion of PrPc into PrPsc and prion disease pathogenesis.
MH Amino Acid Sequence; Chromatography, High Pressure Liquid; Circular Dichroism; Humans; Molecular Sequence Data; Peptide Fragments/chemistry; Prion Diseases/physiopathology; Prions/chemical synthesis/*chemistry/ultrastructure; Protein Conformation; Research Support, Non-U.S. Gov't; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
AD Dulbecco Telethon Institute, Department of Molecular Biochemistry and Pharmacology, Istituto di Ricerche Farmacologiche Mario Negri, via Eritrea 62, Milan 20157, Italy. bonetto@marionegri.it
SP englisch
PO USA