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Identification of proteins with high affinity for refolded and native PrP C
Author(s) -
Petrakis Spyros,
Sklaviadis Theodoros
Publication year - 2006
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.200600103
Subject(s) - gene isoform , biology , microbiology and biotechnology , western blot , proteomics , blot , immunoprecipitation , signal transduction , recombinant dna , receptor , biochemistry , gene
PrP C , the cellular prion protein, is widely expressed in most tissues, including brain, muscle and the gastrointestinal tract, but its physiological role remains unclear. During propagation of transmissible spongiform encephalopathies (TSEs), prion protein is converted to the pathological isoform, PrP Sc , in a process believed to be mediated by as‐yet‐unknown host factors. The identification of proteins associated with PrP may provide information about the biology of prions and the pathogenesis of TSEs. In the present work, we report proteins identified from brain tissue based on their ability to bind to recombinant PrP (recPrP) or form multimolecular complexes with native PrP C in the presence of cross‐linkers. Immobilized his‐tagged recPrP was used as an affinity matrix to isolate PrP‐interacting proteins from brain homogenates of normal individuals. In parallel, PrP C ‐associated proteins were characterized by cross‐linking and co‐immunoprecipitation assays. The unknown molecules were identified by MS and the results of LC‐MS/MS analysis were subsequently verified by Western blot. Both techniques resulted in identification of proteins participating in the formation of cytoskeleton and signal transduction, further supporting the hypothesis that PrP is involved in the organization and function of receptors throughout the nervous system.