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ARS Home » Pacific West Area » Albany, California » Western Regional Research Center » Produce Safety and Microbiology Research » Research » Publications at this Location » Publication #309849
Title: Quantifying the role of lysine in prion replication by Nano-LC mass spectrometry and bioassay

Author
item Silva, Christopher - Chris
item Erickson-Beltran, Melissa
item DYNIN, IRINA - Former ARS Employee

Submitted to: Frontiers in Bioengineering and Biotechnology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/24/2020
Publication Date: 9/23/2020
Citation: Silva, C.J., Erickson-Beltran, M.L., Dynin, I.C. 2020. Quantifying the role of lysine in prion replication by Nano-LC mass spectrometry and bioassay. Frontiers in Bioengineering and Biotechnology. 8. Article 562953. https://doi.org/10.3389/fbioe.2020.562953.
DOI: https://doi.org/10.3389/fbioe.2020.562953

Interpretive Summary: Prions are misfolded proteins that cause of a set of fatal diseases in animals and humans called transmissible spongiform encephalopathies (TSEs). Prions were reacted with chemical reagents and analyzed to determine the extent of the reaction with each lysine present in the prion. The reacted prions were also analyzed to determine their infectivity. The chemical reaction of prions reduced the infectivity by 90%. Further analysis indicated that only one of the lysines present in the protein, lysine at position 220, also reacted to a 90% extent. All of the other lysines reacted to a much greater extent. This suggests that the reaction of the lysine at position 220 is responsible for the loss of infectivity. Since the other ten lysines (not the one at position 220) reacted more extensively with the chemicals, they have a minimal role in prion replication.

Technical Abstract: Prions propagate by a template driven process, inducing the normal cellular isoform (PrPC) to adopt the prion (PrPSc) conformation. In PrPC, the positions of lysines are highly conserved and strongly influence prion propagation. In this study, covalent modification was used to quantitate the role of lysines in the PrPSc template that drives prion replication. The e-amino group of lysines in the PrPSc (hamster-adapted scrapie Sc237) template was acetylated by either acetic anhydride (Ac2O) or the N-hydroxysuccinimide ester of acetic acid (Ac-NHS). The extent of lysine acetylation in PrPSc was quantitated by mass spectrometry or Western blot-based analysis. Identical samples were bioassayed to quantitate the loss of infectivity associated with lysine acetylation. The reduction of infectivity at the highest reagent concentration was approximately 90% (~10-fold). Ten of the eleven prion lysines were acetylated to a greater extent (25-400-fold) than the observed loss of infectivity. Only one lysine, at position 220 (K220), had a reactivity that is consistent with the loss of infectivity. Although lysines are highly conserved and play a crucial role in converting PrPC into the PrPSc conformation, once that conformation is adopted, the lysines present in the PrPSc template play only a limited role in prion replication. In principle, this approach could be used to clarify the role of other amino acids in the replication of prions and other prion-like protein misfolding diseases.