title: Osmolyte trimethylamine N-oxide converts recombinant alpha-helical prion protein to its soluble beta-structured form at high temperature.
abstract: The thermal unfolding of full-length human recombinant alpha-helical prion protein (alpha-PrP) in neutral pH is reversible, whereas, in the presence of the osmolyte N-trimethylamine oxide (TMAO), the protein acquires a beta-sheet structure at higher temperatures and the thermal unfolding of the protein is irreversible. Lysozyme, an amyloidogenic protein similar to prion protein, regains alpha-helical structure on cooling from its thermally unfolded form in buffer and in TMAO solutions. The thermal stability of alpha-PrP decreases, whereas that of lysozyme increases in TMAO solution. Light-scattering and turbidity values indicate that beta-sheet prion protein exists as soluble oligomers that increase thioflavin T fluorescence and bind to 1-anilino 8-naphthalene sulfonic acid (ANS). The oligomers are resistant to proteinase K digestion and during incubation for long periods they form linear amyloids>5 microm long. The comparable fluorescence polarization of the tryptophan groups and their accessibility to acrylamide in alpha-PrP and oligomers indicate that the unstructured N-terminal segments of the protein, which contain the tryptophan groups, do not associate among themselves during oligomerization. Partial unfolding of alpha-helical prion protein in TMAO solution leads to its structural conversion to misfolded beta-sheet form. The formation of the misfolded prion protein oligomers and their polymerization to amyloids in TMAO are unusual, since the osmolyte generally induces denatured protein to fold to a native-like state and protects proteins from thermal denaturation and aggregation.
authors.affiliation: Infectiologie Animale et Santé Publique, Institut National de la Recherche Agronomique, 37380 Nouzilly, France. nandi@tours.inra.fr
authors.forename: P K
authors.initials: PK
authors.lastname: Nandi
authors.forename: A
authors.initials: A
authors.lastname: Bera
authors.forename: P-Y
authors.initials: PY
authors.lastname: Sizaret
mesh_terms: D000817:Anilino Naphthalenesulfonates / Q000378:metabolism; D000818:Animals; D052160:Benzothiazoles; D002942:Circular Dichroism; D004591:Electrophoresis, Polyacrylamide Gel; D019286:Endopeptidase K / Q000378:metabolism; D005453:Fluorescence; D006358:Hot Temperature; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008744:Methylamines / Q000494:pharmacology*; D051379:Mice; D009113:Muramidase / Q000737:chemistry; D010446:Peptide Fragments / Q000737:chemistry; D011328:Prions / Q000737:chemistry* / Q000648:ultrastructure; D011485:Protein Binding; D017433:Protein Structure, Secondary / Q000187:drug effects*; D011994:Recombinant Proteins / Q000737:chemistry*; D012542:Scattering, Radiation; D012995:Solubility / Q000187:drug effects; D013844:Thiazoles / Q000378:metabolism; D014364:Tryptophan / Q000378:metabolism
chemical_list: D000817:Anilino Naphthalenesulfonates; D052160:Benzothiazoles; D008744:Methylamines; D010446:Peptide Fragments; D011328:Prions; D011994:Recombinant Proteins; D013844:Thiazoles; C009462:thioflavin T; C027132:1-anilino-8-naphthalenesulfonate; D014364:Tryptophan; D009113:Muramidase; D019286:Endopeptidase K; C005855:trimethyloxamine