Alternative prion structural changes revealed by high pressure.
At high temperature, recombinant hamster prion protein (SHaPrP(90-231)) undergoes aggregation and changes from a predominantly alpha-helical to beta-sheet conformation. We then applied high pressure (200 MPa) to the beta-sheet-rich conformation. The aggregation was reversed, and the original tertiary and secondary structures were recovered at ambient pressure, after pressure release. The application of a pressure of 200 MPa thus allowed studying the heat-induced equilibrium refolding in the absence of protein aggregation. Prion protein unfolding as a function of high pressure was also investigated. Simple two-state, reversible unfolding transitions were observed, as monitored by spectral changes in the UV and fluorescence of the hydrophobic probe 8-anilino-1-naphthalene sulfonate. However, these heat- and pressure-induced conformers differed in their unfolding free energy. At pressures over 400 MPa, strong thioflavin-T binding was observed, suggesting a further structural change to a metastable oligomeric structure.
Torrent
Joan
J
INSERM U128, 1919 Route de Mende, F-34293 Montpellier cedex 5, France.
Alvarez-Martinez
Maria Teresa
MT
Heitz
Frédéric
F
Liautard
Jean-Pierre
JP
Balny
Claude
C
Lange
Reinhard
R
D000818:Animals; D011232:Chemical Precipitation; D006224:Cricetinae; D006358:Hot Temperature; D007700:Kinetics; D008027:Light; D008647:Mesocricetus; D010446:Peptide Fragments / Q000737:chemistry* / Q000378:metabolism; D011312:Pressure; D011328:Prions / Q000737:chemistry* / Q000378:metabolism; D011487:Protein Conformation; D017510:Protein Folding; D020033:Protein Isoforms / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012542:Scattering, Radiation; D013056:Spectrophotometry, Ultraviolet; D017550:Spectroscopy, Fourier Transform Infrared; D013329:Structure-Activity Relationship
D010446:Peptide Fragments; D011328:Prions; D020033:Protein Isoforms; D011994:Recombinant Proteins; C428437:prion protein (23-231)