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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 #297157
Title: Distinguishing between PrPC and PrPSc using small molecule reagents

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

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 10/1/2013
Publication Date: 8/15/2013
Citation: Silva, C.J., Erickson-Beltran, M.L., Dynin, I., Carter, J.M. 2013. Distinguishing between PrPC and PrPSc using small molecule reagents. Meeting Abstract. Prion 2013 Banff, Alberta, Canada May 26-29, 2013. jdc..

Interpretive Summary:

Technical Abstract: Background/Introduction. The structural difference between PrPSc and PrPC is entirely conformational: they are isoforms. Both isoforms possess identical covalent structures and identical post-translational modifications. This means that the same amino acid can react differently with the same chemical reagent, depending upon which of the isoforms is reacted. The site of covalent modification can be identified by mass spectrometry or by Western blot, if the epitope of the primary antibody contains an amino acid that can be covalently modified by a selected reagent. Materials and Methods. A set of small molecules reagents was synthesized. These reagents preferentially react with the (epsilon)-amino group of lysine. They were reacted with PrPSc, PrPC, and recombinant PrP. The reaction mixtures were analyzed by mass spectrometry and by Western blot to determine the relative reactivity of the various lysines. Four of the antibodies we used recognize an epitope that is encrypted in the PrPSc isoform, but exposed in the PrPC isoform. Results and Conclusions. Because these reagents block the recognition of PrPC, Western blot analysis of these reactions permits the detection of prion infected brain extracts without the need for proteinase K digestion. This is important, because although proteinase K-resistant PrP is diagnostic for disease, much of infectious PrP is proteinase K-sensitive. In addition these reagents can be used, with an appropriate antibody, to determine which amino acids of PrPSc are exposed on the surface and which are encrypted, thus providing useful structural information. This approach was used to distinguish among strains of hamster-adapted scrapie without the use of proteinase K. The mass spectrometry-based analysis was used to quantitate these differences and analyze the relative reactivity of the various lysines present in hamster PrP.