Towards a molecular Level Understanding of Prion Disease Development: Mutant forms of the Mouse Prion Protein, their Structures and Folding Properties

Prion diseases are fatal neurodegenerative disorders, which can be familial, infectious or sporadic. Prion diseases occurring in humans are: Creutzfeld-Jakob Disease (CJD), Gerstmann-Sträussler-Schenker Syndrome, Fatal Familial Insomnia (FFI) and Kuru. The genesis of those diseases is still subject of controversial discussions (1). Whereas a part of the scientific community thinks that a yet undiscovered virus is the reason for the infection, other researchers tend to believe that the infectious particle is a protein, which would mean a revolution in molecular biology. There is a lot of evidence in favour of the "protein-only" hypothesis, but the possibility of a virus involved -maybe acting together with a protein molecule - still cannot be ruled out completely. The "protein-only" hypothesis (2) for the development of prion diseases states that prion proteins occur in two different conformational isomeres: the benign form denoted by PrP C and the infectious form denoted by PrP Sc (the index Sc is derived from "scrapie", a prion disease occurring in sheep). The malfolded PrP Sc seems to be able to catalyze a process that is flipping prion proteins from the PrP C form to the abberant PrP Sc form. Different mechanisms have been proposed for this process (3). Prp Sc differs from PrP C in that it accumulates in the cell, whereas PrP Sc can form rod-shaped aggregates called plaques is of central interest for the verification of the "protein-only" hypothesis. The understanding of prion disease development is both scientifically intriguing and also of enormous economical and political importance since the possibility of a transmission of Bovine Spongiform Encephalopathy (BSE) - a prion disease occurring in cows - to humans has put scientists, politicians and consumers in Europe to a state of considerable uncertainty. The transition of PrP C to PrP Sc is ascribed to a change in coformation, because no post-translational chemical process responsible for this transition could be found (6), but large differences between the two forms in beta-sheet content could be detected (7). Certain regions of the prion protein that form alpha-helices in the PrP C form are thought to adopt a beta-sheet structure in the PrP Sc form. Thereby, hydrophobic regions could be exposed to the surface of the protein and the protein forms aggregates to avoid the unfavorable interaction with the (hydrophilic) environment (8). This process can be triggered by inoculation of healthy, PrP C containing tissue with infectious PrP Sc. There are, however, individuals who seem to develop a prion disease (and thereby PrP Sc) without being infected. They show special mutations in their prion protein which seem to make the development of a prion disease easier (ref. (9) gives some examples). An obvious explanation is that these modified prion proteins more readily fold into the PrP Sc isoform. That is where the proposed project intervenes: a variety of mutant forms of the mouse prion protein has been prepared in the group of Prof. Wüthrich at the ETH Zürich. The know-how of NMR-studies on prion proteins is also available there from previous projects such as the NMR-structure determination of the wild type mouse prion protein PrP C (10). The aim of the proposed research project is to determine the 3D-structure of the mutant forms by means of NMR. NMR spectroscopy will also be involved in studying unfolding/refolding processes of the protein molecule. In general, as much physical-chemical data as possible shall be obtained from the set of the wild type protein and its available mutants. This will help to understand the genesis of prion diseases in humans and other mammals, a point of exceeding importance as already outlined above.