Author
|
Submitted to: Meeting Abstract
Publication Type: Abstract Only Publication Acceptance Date: 2/21/2015 Publication Date: 2/26/2015 Citation: Silva, C.J. 2015. Detecting prions and discriminating among prion strains by discerning the differences in absence.. Meeting Abstract. Volume 2015. Page: Lecture #23. Interpretive Summary: Prions are molecular pathogens, able to convert a normal cellular prion protein (PrPC) into a prion (PrPSc). The only demonstrated difference between PrPC and PrPSc is conformational. This means that the information necessary for this conversion is contained solely in the conformation of PrPSc. It also means that the same amino acid in the PrPC or PrPSc conformation may react differently with the same reagent in a conformation-dependent manner. The '-amino group of lysine readily reacts with a variety of activated forms of organic acids (acetic anhydride, N-hydroxysuccinimide esters, '-propiolactone, etc.). These reactions result in the conversion of the '-amino group into the amide of the corresponding organic acid. The extent of this reaction can be monitored by using monoclonal antibodies (mAbs) with lysine-containing epitopes(e.g. 3F4, 6D11, AH6 or GE8). If the lysine has been covalently modified, then the mAb will no longer bind, since its epitope has been altered by covalent modification. Monitoring the extent of this reaction requires monitoring of the loss of signal due to the covalent modification of the lysine. Lysines in PrPC conformation tend to be more reactive to the activated organic acid reagents than the same lysines in the PrPSc conformation. The extent of the reaction can be monitored by mass spectrometry. This approach was used to distinguish between the 263K and drowsy strains of hamster-adapted scrapie. In this way, prions can be detected and prion strains discriminated by discerning the differences in absence of signals. Technical Abstract: Prions are molecular pathogens, able to convert a normal cellular prion protein (PrPC) into a prion (PrPSc). The only demonstrated difference between PrPC and PrPSc is conformational. This means that the information necessary for this conversion is contained solely in the conformation of PrPSc. It also means that the same amino acid in the PrPC or PrPSc conformation may react differently with the same reagent in a conformation-dependent manner. The '-amino group of lysine readily reacts with a variety of activated forms of organic acids (acetic anhydride, N-hydroxysuccinimide esters, '-propiolactone, etc.). These reactions result in the conversion of the '-amino group into the amide of the corresponding organic acid. The extent of this reaction can be monitored by using monoclonal antibodies (mAbs) with lysine-containing epitopes(e.g. 3F4, 6D11, AH6 or GE8). If the lysine has been covalently modified, then the mAb will no longer bind, since its epitope has been altered by covalent modification. Monitoring the extent of this reaction requires monitoring of the loss of signal due to the covalent modification of the lysine. Lysines in PrPC conformation tend to be more reactive to the activated organic acid reagents than the same lysines in the PrPSc conformation. The extent of the reaction can be monitored by mass spectrometry. This approach was used to distinguish between the 263K and drowsy strains of hamster-adapted scrapie. In this way, prions can be detected and prion strains discriminated by discerning the differences in absence of signals. |
