NR ATQO
AU Rigter,A.; Langeveld,J.P.M.; Timmer-Parohi,D.; Bossers,A.
TI Probing for Sheep Prion Protein Interaction Sites
QU International Conference - Prion 2005: Between fundamentals and society's needs - 19.10.-21.10.2005, Congress Center Düsseldorf - Poster Session: Structure of PrP and molecular determinants of infectivity STRCT-04
PT Konferenz-Poster
AB
The central event in transmissible spongiform encephalopathies is the conversion of host-encoded protease sensitive cellular prion protein (PrPc) into the scrapie associated protease resistant isoform (PrPsc) of prion protein (PrP). Even though disease associated polymorphisms in sheep PrP modulate the conversion of PrPc into protease resistant isoform, the exact mechanism underlying this effect is not yet clear. It has been demonstrated that these polymorphisms do not modulate the initial binding of PrPc to PrPsc and that a subsequent step in the conversion process following initial binding should be responsible for determining differential conversion efficiencies. Because it is not clear whether the primary binding site of PrPc to PrPsc is also the site where the actual conversion is initiated (nucleation site) or whether a separate nucleation site is present in PrP, possible interaction sites need to be determined in order to elucidate which site is responsible for binding and/or initiation of conversion.
In order to determine which residues are capable of interacting with PrPc a PrP protein array containing a complete set of overlapping 15-mer peptides (PEPSCAN) was probed for interaction sites using a sheep PrPc fused to Maltose Binding Protein (MBP-PrPc). The fusion protein MBP-PrPc was incubated on the PEPSCAN after which bound MBP-PrPc was detected by indirect ELISA immunoscreening.
The PEPSCAN results revealed two distinctive high binding areas. The first area covered the ovine PrP amino acid residues 30-108, comprising the N-terminal octarepeats. The second area covered residues 128-197, encompassing the disease associated polymorphisms in sheep PrP. Since it was demonstrated that these polymorphisms do not modulate the initial binding of PrPc to PrPsc, several peptides were selected based on the PEPSCAN binding results in order to test their capability to interfere in binding of PrPc to PrPsc and/or the actual conversion of ovine PrPc.
AD Alan Rigter, Jan P.M. Langeveld, Alex Bossers, CIDC-Lelystad, the Netherlands; Drohpatie Timmer-Parohi, Pepscan Systems, the Netherlands
SP englisch
PO Deutschland