Quantitating PrP Polymorphisms Present in Prions from Heterozygous Scrapie-Infected Sheep.
Anal Chem
; 89(1): 854-861, 2017 01 03.
Article
em En
| MEDLINE
| ID: mdl-27936597
ABSTRACT
Scrapie is a prion (PrPSc) disease of sheep. The incubation period of sheep scrapie is strongly influenced by polymorphisms at positions 136, 154, and 171 of a sheep's normal cellular prion protein (PrPC). Chymotrypsin was used to digest sheep recombinant PrP to identify a set of characteristic peptides [M132LGSXMSRPL141 (X = A or V), Y153XENMY158 (X,= H or R), and Y166RPVDXY172 (X = H, K, Q, or R)] that could be used to detect and quantitate polymorphisms at positions 136, 154, and 171 of sheep PrPC or PrPSc. These peptides were used to develop a multiple reaction monitoring method (MRM) to detect the amounts of a particular polymorphism in a sample of PrPSc isolated from sheep heterozygous for their PrPC proteins. The limit of detection for these peptides was less than 50 attomole. Spinal cord tissue from heterozygous (ARQ/VRQ or ARH/ARQ) scrapie-infected Rasa Aragonesa sheep was analyzed using this MRM method. Both sets of heterozygotes show the presence of both polymorphisms in PrPSc. This was true for samples containing both proteinase K (PK)-sensitive and PK-resistant PrPSc and samples containing only the PK-resistant PrPSc. These results show that heterozygous animals contain PrPSc that is composed of significant amounts of both PrP polymorphisms.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Polimorfismo Genético
/
Scrapie
/
Príons
Tipo de estudo:
Prognostic_studies
Limite:
Animals
Idioma:
En
Ano de publicação:
2017
Tipo de documento:
Article