NR ATMI
AU Barnicle,D.A.; Ryder,S.; Jackman,R.; Terry,L.A.
TI Differential proteolysis and PrPsc conformers
QU International Conference - Prion 2005: Between fundamentals and society's needs - 19.10.-21.10.2005, Congress Center Düsseldorf - Poster Session: Genetics, strains and emerging problems GEN-08
PT Konferenz-Poster
AB The abnormal form of the prion protein (PrPd) associated with transmissible spongiform encephalopthies has been conventionally characterised by its relative resistance to degradation by proteolytic enzymes. Recent evidence has indicated that strain diversity of prion diseases may be identified by differences in proteolysis of PrPd. The aim of this study was to determine whether characteristics of resistance to degradation may be exploited in order to identify prion strains. Using SDS-PAGE and Western blot, the kinetics of hydrolysis of PrPd derived from brain homogenates of cattle infected with BSE and sheep infected with scrapie were compared. Susceptibility of the protease 'resistant' core, as defined by binding to 6H4 mAb, to partial and complete digestion over extended periods of time (up to 48 hours) was studied. Differences in kinetics of proteolysis of the core region between scrapie and BSE were not identified, but changes in rates of digestion within the host species were found to be dependent on buffer composition. Kinetic studies to ascertain the extent of hydrolysis of N-terminus of PrP were performed using antibodies specific to epitopes between amino acids 23-102 of the PrP molecule. The N-terminus of PrPd was found to have intrinsic resistance to protease digestion challenging the premise that this region has little defined tertiary structure. In this case, the rate of digestion of the N-terminus was clearly species specific and partially dependent on buffer composition. More importantly differences in kinetics and susceptibility in this region to proteolysis were found to depend on strain or source of inoculum as shown by studies on experimental scrapie in cattle and BSE in sheep. These studies have demonstrated patterns of proteolytic digestion of PrPd that may facilitate the definition of subtle variation in PrPd conformation and enable differential identification of prion strains.
IN Die Autoren wollen die bekanntlich bei verschiedenen TSE-Erregerstämmen unterschiedliche Resistenz gegen Inkubationen mit der Proteinase K ausnutzen, um diese zu unterscheiden.
AD D.A.Barnicle, R.Jackman, VLA, Weybridge, UK; S.Ryder, Home Office, UK; L.A.Terry, VMD, UK
SP englisch
PO Deutschland