Conformational properties of beta-PrP.
Prion propagation involves a conformational transition of the cellular form of prion protein (PrPC) to a disease-specific isomer (PrPSc), shifting from a predominantly alpha-helical conformation to one dominated by beta-sheet structure. This conformational transition is of critical importance in understanding the molecular basis for prion disease. Here, we elucidate the conformational properties of a disulfide-reduced fragment of human PrP spanning residues 91-231 under acidic conditions, using a combination of heteronuclear NMR, analytical ultracentrifugation, and circular dichroism. We find that this form of the protein, which similarly to PrPSc, is a potent inhibitor of the 26 S proteasome, assembles into soluble oligomers that have significant beta-sheet content. The monomeric precursor to these oligomers exhibits many of the characteristics of a molten globule intermediate with some helical character in regions that form helices I and III in the PrPC conformation, whereas helix II exhibits little evidence for adopting a helical conformation, suggesting that this region is a likely source of interaction within the initial phases of the transformation to a beta-rich conformation. This precursor state is almost as compact as the folded PrPC structure and, as it assembles, only residues 126-227 are immobilized within the oligomeric structure, leaving the remainder in a mobile, random-coil state.
MRC Prion Unit, Department of Neurodegenerative Disease, Institute of Neurology, Queen Square, London WC1N 3BG; Krebs Institute for Biomolecular Research, Department of Molecular Biology and Biotechnology, University of Sheffield, Sheffield S10 2TN.
Laszlo L P
LLP
Hosszu
MRC Prion Unit, Department of Neurodegenerative Disease, Institute of Neurology, Queen Square, London WC1N 3BG.
Clare R
CR
Trevitt
MRC Prion Unit, Department of Neurodegenerative Disease, Institute of Neurology, Queen Square, London WC1N 3BG.
Samantha
S
Jones
MRC Prion Unit, Department of Neurodegenerative Disease, Institute of Neurology, Queen Square, London WC1N 3BG.
Mark
M
Batchelor
National Centre for Macromolecular Hydrodynamics, School of Biosciences, University of Nottingham, Sutton Bonington Campus, Leicestershire LE12 5RD, United Kingdom.
David J
DJ
Scott
MRC Prion Unit, Department of Neurodegenerative Disease, Institute of Neurology, Queen Square, London WC1N 3BG.
Graham S
GS
Jackson
MRC Prion Unit, Department of Neurodegenerative Disease, Institute of Neurology, Queen Square, London WC1N 3BG.
John
J
Collinge
Krebs Institute for Biomolecular Research, Department of Molecular Biology and Biotechnology, University of Sheffield, Sheffield S10 2TN.
Jonathan P
JP
Waltho
MRC Prion Unit, Department of Neurodegenerative Disease, Institute of Neurology, Queen Square, London WC1N 3BG.
Anthony R
AR
Clarke
D002942:Circular Dichroism; D004305:Dose-Response Relationship, Drug; D006801:Humans; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008954:Models, Biological; D010455:Peptides / Q000737:chemistry; D018621:PrPC Proteins / Q000737:chemistry* / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex / Q000737:chemistry; D011487:Protein Conformation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D014461:Ultracentrifugation; D014508:Urea / Q000737:chemistry
D010455:Peptides; D018621:PrPC Proteins; D014508:Urea; D046988:Proteasome Endopeptidase Complex; C061553:ATP dependent 26S protease