In vitro conversion and seeded fibrillization of posttranslationally modified prion protein

Stöhr, J.; Wille, H.; Willbold, D.; Birkmann, E.; Riesner, D.; Elfrink, K.; Weinmann, N.

doi:10.1515/bc.2011.048

Journal Article

In vitro conversion and seeded fibrillization of posttranslationally modified prion protein

Stöhr, J. ; Elfrink, K. ; Weinmann, N. ; Wille, H. ; Willbold, D.FZJ* ; Birkmann, E.FZJ* ; Riesner, D.

2011
de Gruyter Berlin [u.a.]

Biological chemistry 392, 415 - 421 (2011) [10.1515/bc.2011.048]

This record in other databases:

Please use a persistent id in citations: http://hdl.handle.net/2128/18354 doi:10.1515/BC.2011.048

Abstract: The conversion of the cellular isoform of the prion protein (PrP(C)) into the pathologic isoform (PrP(Sc)) is the key event in prion diseases. To study the conversion process, an in vitro system based on varying the concentration of low amounts of sodium dodecyl sulfate (SDS) has been employed. In the present study, the conversion of full-length PrP(C) isolated from Chinese hamster ovary cells (CHO-PrP(C)) was examined. CHO-PrP(C) harbors native, posttranslational modifications, including the GPI anchor and two N-linked glyco-sylation sites. The properties of CHO-PrP(C) were compared with those of full-length and N-terminally truncated recombinant PrP. As shown earlier with recombinant PrP (recPrP90-231), transition from a soluble α-helical state as known for native PrP(C) into an aggregated, β-sheet-rich PrP(Sc)-like state could be induced by dilution of SDS. The aggregated state is partially proteinase K (PK)-resistant, exhibiting a cleavage site similar to that found with PrP(Sc). Compared to recPrP (90-231), fibril formation with CHO-PrP(C) requires lower SDS concentrations (0.0075%), and can be drastically accelerated by seeding with PrP(Sc) purified from brain homogenates of terminally sick hamsters. Our results show that recPrP 90-231 and CHO-PrPC behave qualitatively similar but quantitatively different. The in vivo situation can be simulated closer with CHO-PrP(C) because the specific PK cleave site could be shown and the seed-assisted fibrillization was much more efficient.

Keyword(s): Animals (MeSH) ; Blotting, Western (MeSH) ; CHO Cells (MeSH) ; Circular Dichroism (MeSH) ; Cricetinae (MeSH) ; Cricetulus (MeSH) ; Electrophoresis, Polyacrylamide Gel (MeSH) ; Microscopy, Electron (MeSH) ; PrPC Proteins: chemistry (MeSH) ; PrPC Proteins: drug effects (MeSH) ; PrPC Proteins: metabolism (MeSH) ; PrPSc Proteins: chemistry (MeSH) ; PrPSc Proteins: drug effects (MeSH) ; PrPSc Proteins: metabolism (MeSH) ; Protein Processing, Post-Translational (MeSH) ; Protein Structure, Secondary (MeSH) ; Sodium Dodecyl Sulfate: pharmacology (MeSH) ; PrPC Proteins ; PrPSc Proteins ; Sodium Dodecyl Sulfate ; J ; aggregation (auto) ; conversion (auto) ; fibrillization (auto) ; glycosylation (auto) ; GPI anchor (auto) ; prion protein (auto) ; seeding (auto)

Classification:

Biochemistry & Molecular Biology

Note: This work was supported by grants from the Deutsche Forschungsgemeinschaft (DFG), EUNetwork of Excellence (NeuroPrion) and the Praesidentenfond of the Helmholtzgemeinschaft (HGF, Virtual Institute of Structural Biology). Jan Stohr is supported by postdoctoral fellowship of the Deutsche Forschungsgemeinschaft (DFG). We thank S.B. Prusiner for critical reading and helpful discussion for this manuscript. The authors thank Ilka Ostermann for her assistance in the purification of CHO-PrP<SUP>C</SUP>.

Contributing Institute(s):