The structural basis of mammalian prion interspecies transmissibility and strain specificity

• Antragsteller: Dr. Thorsten Lührs
• Fachliche Zuordnung: Strukturbiologie
• Förderung: Förderung von 2008 bis 2015
• Projektkennung: Deutsche Forschungsgemeinschaft (DFG) - Projektnummer 61396538

The mammalian prion protein, PrPC, is able to switch its conformation from a monomeric soluble form into an aggregated transmissible prion state, PrPSc, which leads to prion disease. Prion strains are thought to represent distinct infectious conformations of the same primary amino acid sequence. In the frame of the DFG-funded Project we have developed a novel technique for prion amplification termed Shear Induced Fragmentation/Aggregation (ShIF/A), which allows for the strain-selective in vitro amplification of mammalian prions. This includes the generation of PrPSc from brain-tissue derived PrPC, or alternatively from bacterially expressed PrPC. Thus, some 10’s of milligrams of [2H, 13C, 15N] isotopically labeled mammalian prions for NMR structural investigations can be generated. Three previously published and commonly used methods from other groups turned out to be unsuitable for sample preparation. Ongoing Ph.D. projects employ ShiF/A to characterize mouse and hamster prion strains, in vitro and in vivo. The bulk molecular structural properties of strain-specific prion aggregates are investigated by quenched hydrogen exchange NMR in solution and by cp-MAS solid-state NMR. However, in most prion diseases, amyloid deposition does not represent a pathological hallmark. Rather oligomeric forms of PrPSc (>= 300 kDa) are thought to represent the smallest prion infectious entity. Here we propose to generate such oligomeric forms of recombinant hamster Sc237 PrPSc by the means of ShIF/A in combination with preparative Asymmetric-Flow-Field-Flow fractionation. The 3D-fold of such preparations will then be investigated using 13C-methyl TROSY NMR in solution. The results from this research will make significant contributions towards understanding the molecular determinants of prion infectivity.

• DFG-Verfahren: Emmy Noether-Nachwuchsgruppen
• Internationaler Bezug: USA
• Großgeräte: Asymmetrisches Fluss Feld Fluss Fraktionierungssystem
• Gerätegruppe: 1300 Destillier- und Fraktioniergeräte, Rektifiziergeräte
• Beteiligte Personen: Dr. Byron Caughey, Ph.D.; Professor Dr. Michael Andreas Klein; Professor Dr. Bernd Reif
• Gastgeber: Professor Dr. Rudi Balling, Ph.D.