NR AOXM
AU Notari,S.; Capellari,S.; Giese,A.; Westner,I.; Ghetti,B.; Gambetti,P.; Kretzschmar,H.A.; Parchi,P.
TI Effects of different experimental conditions on the PrPsc core generated by protease digestion: Implications for strain typing and molecular classification of CJD
QU International Conference - Prion diseases: from basic research to intervention concepts - TSE-Forum, 08.10.-10.10.2003, Gasteig, München - Poster session - DG-75
PT Konferenz-Poster
AB
The discovery that the abnormal form of the prion protein, PrPsc, shows strain-specific physico-chemical properties, has provided a potentially powerful new method for strain typing as well as the basis for a novel molecular classification of human trasmissible spongiform encephalopathies (TSEs). Unfortunately, there is still a significant disparity in the literature regarding the existence of distinct PrPsc types. Moreover, whether a specific PrPsc structure is associated to each TSE pathological subtype is still unknown. In sporadic CJD (sCJD), the most common human TSE, 6 disease subtypes are known, but only two major types of PrPsc, type 1 and 2, showing a 2-kDa difference in molecular mass, have been consistently reproduced in most laboratories. We have further analysed the characteristics of PrPsc in 70 sCJD subjects using a gel electrophoresis system with high resolution and different buffer or protease conditions. We found that there is a significant heterogeneity in pH among sCJD brain homogenates prepared in standard buffers, which, in turn influences protease activity and electrophoretic profile of the PrPsc core. We also found that PrPsc type 1 and type 2 are heterogeneous species, which can be further distinct in molecular subtypes. PrPsc type 1 from codon 129 valine homozygotes (VV) migrated faster than the type 1 from codon 129 methionine homozygotes (MM) or heterozygotes (MV). Similarly, among type 2 samples only PrPsc from MV subjects with kuru plaques resolved as a doublet. Protease digestion at different pHs further distinguished between VV1 and MM1, since only in the second group PrPsc showed a pH-dependent shift in gel mobility. Finally, sCJD subtypes differed in PrPsc resistance to protease digestion. In particular, PrPsc from VV2 subjects showed a significantly higher protease resistance than the PrPsc from MM2-cortical cases. Our results shed lights on previous disparities in PrPsc typing, and provide a refined classification of human PrPsc types. The results also support the notion that in TSEs the pathological phenotype is largely related to PrPsc structure.
Supported by EU contract QLK3-CT-2001-02345.
AD Silvio Notari, Sabina Capellari, Piero Parchi, Dipartimento di Scienze Neurologiche, Università di Bologna, Italy; Armin Giese, Ingo Westner, Hans A. Kretzschmar, Institut für Neuropatologie, LMU, Germany; Bernardino Ghetti, Institute of Pathology, Indiana University, USA; Pierluigi Gambetti, Institute of Pathology, CWRU, USA
SP englisch
PO Deutschland