Mass spectroscopic analysis of Sup35NM prion polymerization.
Sup35NM, the prion determining domain of the protein responsible for the yeast prion phenomenon [Psi], has become a powerful model for studying key processes in amyloid-related human diseases. One of these processes is a conformational conversion of soluble precursor protein into insoluble fibrillar structures. In this study, we created a set of Sup35NM mutants and used proteolytic digestion coupled with mass spectroscopy to monitor local structure of the protein during polymerization. Experimental data were compared to a network model and showed that during the conformational conversion residue Arg-28 became highly protected from cleavage, residue Arg-98 remained partially solvent exposed, and residues between 28 and 98 showed an intermediate degree of protection. In addition, we found that a distinct subset of proteolytic polypeptides spanning 28-98 residues segment spontaneously formed stable dimers. This finding suggests that the [29-98] region is the key interacting region of Sup35NM responsible for amyloid conversion.
Goncharov
Vladimir A
VA
Whitehead Institute for Biomedical Research, Cambridge, Massachusetts 02142, USA. goncharov@wi.mit.edu
D000682:Amyloid / Q000032:analysis / Q000737:chemistry* / Q000648:ultrastructure*; D001665:Binding Sites; D003198:Computer Simulation; D019281:Dimerization; D013058:Mass Spectrometry / Q000379:methods*; D008956:Models, Chemical; D008958:Models, Molecular; D046912:Multiprotein Complexes / Q000032:analysis / Q000737:chemistry / Q000648:ultrastructure; D010454:Peptide Termination Factors; D011108:Polymers / Q000737:chemistry; D011328:Prions / Q000032:analysis / Q000737:chemistry* / Q000648:ultrastructure*; D011485:Protein Binding; D011487:Protein Conformation; D029701:Saccharomyces cerevisiae Proteins / Q000032:analysis / Q000737:chemistry* / Q000648:ultrastructure*
D000682:Amyloid; D046912:Multiprotein Complexes; D010454:Peptide Termination Factors; D011108:Polymers; D011328:Prions; C068068:SUP35 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins