Conformational transitions, dissociation, and unfolding of scrapie amyloid (prion) protein

J Biol Chem. 1993 Sep 25;268(27):20276-84.

Abstract

The infectious form of the scrapie amyloid (prion) precursor, PrPSc, is a host-derived protein and a component of the infectious agent causing scrapie. PrPSc and the carboxyl-terminal proteinase K resistant core, PrP27-30, have the potential to form amyloid as a result of a post-translational event or conformational abnormality. We have studied the conformational transitions of both proteins reconstituted into liposomes, associated in solid state in thin films, and dissociated by guanidine HCl. The secondary structure of PrPSc in liposomes deduced from analysis of circular dichroism spectra contained approximately 34% beta-sheets, approximately 20% alpha-helix, and approximately 46% beta-turns and random coil. Cleavage of the amino-terminal region of PrPSc resulted in all-beta PrP27-30, with an estimated approximately 43% beta-sheet, no alpha-helix, and approximately 57% beta-turns and random coil. The PrPSC associated in thin films with a tertiary structure perturbation corresponding to unfolding, while the secondary structure was preserved. The PrP27-30 assembled into the solid state with a similar perturbation of tertiary structure but with a large increase in the beta-sheet content, probably due to an intermolecular alignment of the external beta-sheets, or to a secondary structure transition, or both. The various conformational states had little or no impact on infectivity. Equilibrium dissociation and unfolding demonstrated a greater resistance of PrP27-30 to denaturation. The dissociated monomers unfolded through intermediate(s), suggesting the presence of protein domains with distinct secondary structure stabilities. The results provide experimental evidence for the beta-sheet type assembly of scrapie amyloid PrP27-30 in the solid state and demonstrate the importance of amino-terminal cleavage in the stability and alignment of the amyloid-forming monomers.

Publication types

  • Comparative Study

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Blotting, Western
  • Circular Dichroism
  • Cricetinae
  • Electrophoresis, Polyacrylamide Gel
  • Female
  • Mesocricetus
  • Molecular Sequence Data
  • PrP 27-30 Protein
  • PrPSc Proteins
  • Prions / chemistry*
  • Prions / metabolism*
  • Prions / pathogenicity
  • Protein Conformation*
  • Protein Structure, Secondary*
  • Scrapie / microbiology
  • Spectrometry, Fluorescence
  • Spectrophotometry, Ultraviolet
  • Virulence

Substances

  • PrPSc Proteins
  • Prions
  • PrP 27-30 Protein