Filaments of the Ure2p prion protein have a cross-beta core structure.
Formation of filaments by the Ure2 protein constitutes the molecular mechanism of the [URE3] prion in yeast. According to the "amyloid backbone" model, the N-terminal asparagine-rich domains of Ure2p polymerize to form an amyloid core fibril that is surrounded by C-terminal domains in their native conformation. Protease resistance and Congo Red binding as well as beta-sheet content detected by spectroscopy-all markers for amyloid-have supported this model, as has the close resemblance between 40 A N-domain fibrils and the fibrillar core of intact Ure2p filaments visualized by cryo-electron microscopy and scanning transmission electron microscopy. Here, we present electron diffraction and X-ray diffraction data from filaments of Ure2p, of N-domains alone, of fragments thereof, and of an N-domain-containing fusion protein that demonstrate in each case the 4.7A reflection that is typical for cross-beta structure and highly indicative of amyloid. This reflection was observed for specimens prepared by air-drying with and without sucrose embedding. To confirm that the corresponding structure is not an artifact of air-drying, the reflection was also demonstrated for specimens preserved in vitreous ice. Local area electron diffraction and X-ray diffraction from partially aligned specimens showed that the 4.7A reflection is meridional and therefore the underlying structure is cross-beta.
Baxa
Ulrich
U
Laboratory of Structural Biology, National Institute of Arthritis, Musculoskeletal, and Skin Diseases, National Institutes of Health, Bethesda, MD 20892, USA.
Cheng
Naiqian
N
Winkler
Dennis C
DC
Chiu
Thang K
TK
Davies
David R
DR
Sharma
Deepak
D
Inouye
Hideyo
H
Kirschner
Daniel A
DA
Wickner
Reed B
RB
Steven
Alasdair C
AC
D005979:Glutathione Peroxidase; D046529:Microscopy, Electron, Transmission; D011108:Polymers; D011328:Prions / Q000737:chemistry*; D011487:Protein Conformation; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry*; D014961:X-Ray Diffraction
D011108:Polymers; D011328:Prions; D029701:Saccharomyces cerevisiae Proteins; D005979:Glutathione Peroxidase; C067020:URE2 protein, S cerevisiae