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.