title: The native-like conformation of Ure2p in fibrils assembled under physiologically relevant conditions switches to an amyloid-like conformation upon heat-treatment of the fibrils.
abstract: The [URE3] phenotype in the yeast Saccharomyces cerevisiae is inherited by a prion mechanism involving self-propagating Ure2p aggregates. It is believed that assembly of intact Ure2p into fibrillar polymers that bind Congo Red and show yellow-green birefringence upon staining and are resistant to proteolysis is the consequence of a major change in the conformation of the protein. We recently dissected the assembly process of Ure2p and showed the protein to retain its native alpha-helical structure upon assembly into protein fibrils that are similar to amyloids in that they are straight, bind Congo red and show green-yellow birefringence and have an increased resistance to proteolysis (). Here we further show using specific ligand binding, FTIR spectroscopy and X-ray fiber diffraction that Ure2p fibrils assembled under physiologically relevant conditions are devoid of a cross-beta core. The X-ray fiber diffraction pattern of these fibrils reveals their well-defined axial supramolecular order. By analyzing the effect of heat-treatment on Ure2p fibrils we bring evidences for a large conformational change that occurs within the fibrils with the loss of the ligand binding capacity, decrease of the alpha helicity, the formation of a cross-beta core and the disappearance of the axial supramolecular order. The extent of the conformational change suggests that it is not limited to the N-terminal part of Ure2p polypeptide chain. We show that the heat-treated fibrils that possess a cross-beta core are unable to propagate their structural characteristic while native-like fibrils are. Finally, the potential evolution of native-like fibrils into amyloid fibrils is discussed.
authors.affiliation: Laboratoire d'Enzymologie et Biochimie Structurales, CNRS, Bâtiment 34 Avenue de la Terrasse, Gif-sur-Yvette Cedex FR-91198, France.
authors.forename: Luc
authors.initials: L
authors.lastname: Bousset
authors.forename: Fatma
authors.initials: F
authors.lastname: Briki
authors.forename: Jean
authors.initials: J
authors.lastname: Doucet
authors.forename: Ronald
authors.initials: R
authors.lastname: Melki
mesh_terms: D000682:Amyloid / Q000737:chemistry*; D003224:Congo Red / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D005978:Glutathione / Q000737:chemistry; D005979:Glutathione Peroxidase; D006358:Hot Temperature; D009281:Naphthalenes / Q000737:chemistry; D010641:Phenotype; D011328:Prions / Q000737:chemistry*; D011485:Protein Binding; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry*; D017550:Spectroscopy, Fourier Transform Infrared; D013997:Time Factors; D014961:X-Ray Diffraction
chemical_list: D000682:Amyloid; D009281:Naphthalenes; D011328:Prions; D029701:Saccharomyces cerevisiae Proteins; D003224:Congo Red; D005979:Glutathione Peroxidase; C067020:URE2 protein, S cerevisiae; D005978:Glutathione