title: Structural intermediates in the putative pathway from the cellular prion protein to the pathogenic form.
abstract: The conversion of the alpha-helical, protease sensitive and noninfectious form of the prion protein (PrP(C)) into an insoluble, protease resistant, predominantly beta-sheeted and infectious form (PrP(Sc)) is the fundamental event in prion formation. In the present work, two soluble and stable intermediate structural states are newly identified for recombinant Syrian hamster PrP(90-231) (recPrP), a dimeric alpha-helical state and a tetra- or oligomeric, beta-sheet rich state. In 0.2% SDS at room temperature, recPrP is soluble and exhibits alpha-helical and random coil secondary structure as determined by circular dichroism. Reduction of the SDS concentration to 0.06% leads first to a small increase in alpha-helical content, whereas further dilution to 0.02% results in the aquisition of beta-sheet structure. The reversible transition curve is sigmoidal within a narrow range of SDS concentrations (0.04 to 0.02%). Size exclusion chromatography and chemical crosslinking revealed that the alpha-helical form is dimeric, while the beta-sheet rich form is tetra- or oligomeric. Both the alpha-helical and beta-sheet rich intermediates are soluble and stable. Thus, they should be accessible to further structural and mechanistic studies. At 0.01% SDS, the oligomeric intermediates aggregated into large, insoluble structures as observed by fluorescence correlation spectroscopy. Our results are discussed with respect to the mechanism of PrP(Sc) formation and the propagation of prions.
authors.affiliation: Institut für Physikalische Biologie, Heinrich-Heine-Universität, Düsseldorf, Germany.
authors.forename: K
authors.initials: K
authors.lastname: Jansen
authors.forename: O
authors.initials: O
authors.lastname: Schäfer
authors.forename: E
authors.initials: E
authors.lastname: Birkmann
authors.forename: K
authors.initials: K
authors.lastname: Post
authors.forename: H
authors.initials: H
authors.lastname: Serban
authors.forename: S B
authors.initials: SB
authors.lastname: Prusiner
authors.forename: D
authors.initials: D
authors.lastname: Riesner
mesh_terms: D002021:Buffers; D002850:Chromatography, Gel; D002942:Circular Dichroism; D003432:Cross-Linking Reagents / Q000737:chemistry; D011108:Polymers / Q000737:chemistry; D018620:PrPSc Proteins / Q000737:chemistry* / Q000472:pathogenicity; D011328:Prions / Q000737:chemistry*; D011487:Protein Conformation; D017510:Protein Folding; D011994:Recombinant Proteins; D012995:Solubility; D013050:Spectrometry, Fluorescence
chemical_list: D002021:Buffers; D003432:Cross-Linking Reagents; D011108:Polymers; D018620:PrPSc Proteins; D011328:Prions; D011994:Recombinant Proteins