Recombinant PrPScshares structural features with brain-derived PrPSc: Insights from limited proteolysis

Alejandro M. Sevillano, Natalia Fernández-Borges, Neelam Younas, Fei Wang, Saioa R. Elezgarai, Susana Bravo, Ester Vázquez-Fernández, Isaac Rosa, Hasier Eraña, David Gil, Sonia Veiga, Enric Vidal, Melissa L. Erickson-Beltran, Esteban Guitián, Christopher J. Silva, Romolo Nonno, Jiyan Ma, Joaquín Castilla, Jesús R. Requena

Research output: Contribution to journalArticle

Abstract

Very solid evidence suggests that the core of full length PrPScis a 4-rung β-solenoid, and that individual PrPScsubunits stack to form amyloid fibers. We recently used limited proteolysis to map the β-strands and connecting loops that make up the PrPScsolenoid. Using high resolution SDS-PAGE followed by epitope analysis, and mass spectrometry, we identified positions ~116/118, 133–134, 141, 152–153, 162, 169 and 179 (murine numbering) as Proteinase K (PK) cleavage sites in PrPSc. Such sites likely define loops and/or borders of β-strands, helping us to predict the threading of the β-solenoid. We have now extended this approach to recombinant PrPSc(recPrPSc). The term recPrPScrefers to bona fide recombinant prions prepared by PMCA, exhibiting infectivity with attack rates of ~100%. Limited proteolysis of mouse and bank vole recPrPScspecies yielded N-terminally truncated PK-resistant fragments similar to those seen in brain-derived PrPSc, albeit with varying relative yields. Along with these fragments, doubly N- and C-terminally truncated fragments, in particular ~89/97-152, were detected in some recPrPScpreparations; similar fragments are characteristic of atypical strains of brain-derived PrPSc. Our results suggest a shared architecture of recPrPScand brain PrPScprions. The observed differences, in particular the distinct yields of specific PK-resistant fragments, are likely due to differences in threading which result in the specific biochemical characteristics of recPrPSc. Furthermore, recombinant PrPScoffers exciting opportunities for structural studies unachievable with brain-derived PrPSc.

Original languageEnglish
Article numbere1006797
JournalPLoS Pathogens
Volume14
Issue number1
DOIs
Publication statusPublished - Jan 1 2018

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Endopeptidase K
Proteolysis
Brain
Arvicolinae
Prions
Amyloid
Epitopes
Polyacrylamide Gel Electrophoresis
Mass Spectrometry

ASJC Scopus subject areas

  • Parasitology
  • Microbiology
  • Immunology
  • Molecular Biology
  • Genetics
  • Virology

Cite this

Sevillano, A. M., Fernández-Borges, N., Younas, N., Wang, F., R. Elezgarai, S., Bravo, S., ... R. Requena, J. (2018). Recombinant PrPScshares structural features with brain-derived PrPSc: Insights from limited proteolysis. PLoS Pathogens, 14(1), [e1006797]. https://doi.org/10.1371/journal.ppat.1006797

Recombinant PrPScshares structural features with brain-derived PrPSc : Insights from limited proteolysis. / Sevillano, Alejandro M.; Fernández-Borges, Natalia; Younas, Neelam; Wang, Fei; R. Elezgarai, Saioa; Bravo, Susana; Vázquez-Fernández, Ester; Rosa, Isaac; Eraña, Hasier; Gil, David; Veiga, Sonia; Vidal, Enric; Erickson-Beltran, Melissa L.; Guitián, Esteban; Silva, Christopher J.; Nonno, Romolo; Ma, Jiyan; Castilla, Joaquín; R. Requena, Jesús.

In: PLoS Pathogens, Vol. 14, No. 1, e1006797, 01.01.2018.

Research output: Contribution to journalArticle

Sevillano, AM, Fernández-Borges, N, Younas, N, Wang, F, R. Elezgarai, S, Bravo, S, Vázquez-Fernández, E, Rosa, I, Eraña, H, Gil, D, Veiga, S, Vidal, E, Erickson-Beltran, ML, Guitián, E, Silva, CJ, Nonno, R, Ma, J, Castilla, J & R. Requena, J 2018, 'Recombinant PrPScshares structural features with brain-derived PrPSc: Insights from limited proteolysis', PLoS Pathogens, vol. 14, no. 1, e1006797. https://doi.org/10.1371/journal.ppat.1006797
Sevillano AM, Fernández-Borges N, Younas N, Wang F, R. Elezgarai S, Bravo S et al. Recombinant PrPScshares structural features with brain-derived PrPSc: Insights from limited proteolysis. PLoS Pathogens. 2018 Jan 1;14(1). e1006797. https://doi.org/10.1371/journal.ppat.1006797
Sevillano, Alejandro M. ; Fernández-Borges, Natalia ; Younas, Neelam ; Wang, Fei ; R. Elezgarai, Saioa ; Bravo, Susana ; Vázquez-Fernández, Ester ; Rosa, Isaac ; Eraña, Hasier ; Gil, David ; Veiga, Sonia ; Vidal, Enric ; Erickson-Beltran, Melissa L. ; Guitián, Esteban ; Silva, Christopher J. ; Nonno, Romolo ; Ma, Jiyan ; Castilla, Joaquín ; R. Requena, Jesús. / Recombinant PrPScshares structural features with brain-derived PrPSc : Insights from limited proteolysis. In: PLoS Pathogens. 2018 ; Vol. 14, No. 1.
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AU - Wang, Fei

AU - R. Elezgarai, Saioa

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