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Peer-reviewedCC BYBiochemistry

A Unique Platform to Study the Dynamic Process of Prion Formation.

  • Jessica de Alcantara Ferreira1
  • Daniel J Walsh1
  • Evelyn Turnbaugh1
  • Jeremy H Mills2
  • Surachai Supattapone1,3
  1. 1Department of Biochemistry, Geisel School of Medicine at Dartmouth, Hanover, New Hampshire03755, United States.
  2. 2School of Molecular Sciences and The Biodesign Center for Molecular Design and Biomimetics, Arizona State UniversityTempe, Arizona85287, United States.
  3. 3Department of Medicine, Geisel School of Medicine at Dartmouth, Hanover, New Hampshire03755, United States.
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Abstract

The pathogenic conversion of the cellular prion protein (PrPC) into the β-sheet-rich isoform PrPSc is the pivotal pathogenic event in prion disease, yet the molecular steps that govern this structural transition remain elusive. In this study, we introduce a new approach to monitor site-specific conformational transitions that occur during infectious prion formation. The method relies on genetically encoded substitution of a fluorescent, environmentally sensitive noncanonical amino acid, l-(7-hydroxycoumarin-4-yl)ethylglycine (7-HCAA), into recombinant PrP substrate molecules, allowing real-time monitoring of structural changes in high-efficiency in vitro PrPSc conversion reactions. As proof of principle, we show that the W99 7-HCAA recPrP substrate efficiently propagates two different PrPSc conformers (infectious cofactor PrPSc and noninfectious protein-only PrPSc). Bioassays in knock-in mice expressing bank vole PrP confirm that W99 7-HCAA cofactor PrPSc produced by serial propagation is infectious, causing scrapie with an incubation period and neuropathological profile like those induced by wild-type cofactor PrPSc. Marked differences in fluorescence intensity were observed between native, misfolded, and denatured states of W99 7-HCAA PrP, confirming that 7-HCAA reports on local changes in PrP conformation. Together, these findings establish 7-HCAA as a site-specific and sensitive probe of local PrP conformation. Moreover, the results suggest a new strategy for studying conformational dynamics in amyloid-forming proteins.

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Cite

BibTeXRIS
BibTeX
@article{ferreira2026unique,
  title = {{A Unique Platform to Study the Dynamic Process of Prion Formation.}},
  author = {Jessica de Alcantara Ferreira and Daniel J Walsh and Evelyn Turnbaugh and Jeremy H Mills and Surachai Supattapone},
  journal = {Biochemistry},
  year = {2026},
  month = sep,
  doi = {10.1021/acs.biochem.6c00351},
  url = {https://europepmc.org/article/MED/42915602},
  note = {CC BY},
}
RIS
TY  - JOUR
TI  - A Unique Platform to Study the Dynamic Process of Prion Formation.
AU  - Jessica de Alcantara Ferreira
AU  - Daniel J Walsh
AU  - Evelyn Turnbaugh
AU  - Jeremy H Mills
AU  - Surachai Supattapone
PY  - 2026
DA  - 2026/09/01
JO  - Biochemistry
DO  - 10.1021/acs.biochem.6c00351
UR  - https://europepmc.org/article/MED/42915602
AB  - The pathogenic conversion of the cellular prion protein (PrPC) into the β-sheet-rich isoform PrPSc is the pivotal pathogenic event in prion disease, yet the molecular steps that govern this structural transition remain elusive. In this study, we introduce a new approach to monitor site-specific conformational transitions that occur during infectious prion formation. The method relies on genetically encoded substitution of a fluorescent, environmentally sensitive noncanonical amino acid, l-(7-hydroxycoumarin-4-yl)ethylglycine (7-HCAA), into recombinant PrP substrate molecules, allowing real-time monitoring of structural changes in high-efficiency in vitro PrPSc conversion reactions. As proof of principle, we show that the W99 7-HCAA recPrP substrate efficiently propagates two different PrPSc conformers (infectious cofactor PrPSc and noninfectious protein-only PrPSc). Bioassays in knock-in mice expressing bank vole PrP confirm that W99 7-HCAA cofactor PrPSc produced by serial propagation is infectious, causing scrapie with an incubation period and neuropathological profile like those induced by wild-type cofactor PrPSc. Marked differences in fluorescence intensity were observed between native, misfolded, and denatured states of W99 7-HCAA PrP, confirming that 7-HCAA reports on local changes in PrP conformation. Together, these findings establish 7-HCAA as a site-specific and sensitive probe of local PrP conformation. Moreover, the results suggest a new strategy for studying conformational dynamics in amyloid-forming proteins.
SN  - 0006-2960
SN  - 1520-4995
ER  -