| Summary: | 碩士 === 國立臺灣大學 === 生化科學研究所 === 103 === The prion diseases are transmissible fatal neurodegenerative disorders affecting human and other mammals. One of the crucial molecular events is the conformational conversion of the normal, membrane-anchored prion protein PrP(C) into the β-sheet-rich and misfolded isoform, PrP(Sc). Within the context of the “protein only” hypothesis, the PrP(Sc) can associate each other to form fibril-like structure. However, the molecular mechanisms underlying the conversion of PrP(C) to the PrP(Sc) form and species barrier remain unclear. Since amyloid formation is a key hallmark of prion pathogenesis, studies of prion protein segment involved in the formation of cross-β structure have provided molecular details of species barrier and insights of amyloid core. The full-length recombinant Syrian hamster prion protein (SHaPrP(23-231)) was produced by E. coli over-expression and five chemically synthetic peptides, SHaPrP(108-144), SHaPrP(108-127), SHaPrP(128-144), SHaPrP(172-194), and SHaPrP(200-227) were produced by solid-phase peptide synthesis. The amyloidogenesis of SHaPrP protein and peptides was investigated in vitro by Thioflavin T (ThT) assay mainly and circular dichroism (CD) spectroscopy. The cross-seeding capacity between SHaPrP protein and peptide fibrils was used to identify the amyloid core region. We found that short peptide SHaPrP(108-144), SHaPrP(108-127), SHaPrP(172-194), and SHaPrP(200-227) fibrils as seed have seeding effect for SHaPrP(23-231) monomer, suggesting that these regions are critical in the amyloidogenesis of SHaPrP(23-231). The full-length recombinant SHaPrP(23-231) fibrils as seed could induce the fibril formation of short peptide SHaPrP(108-144) and SHaPrP(108-127) but not for SHaPrP(128-144). These results suggest that SHaPrP(108-127) segment but not SHaPrP(128-144) segment is involved in the amyloid core of SHaPrP fibrils. This result corresponds to our previous hypothesis that the species barrier between SHaPrP and MoPrP is contributed by utilization of difference amyloid core of PrP. Moreover, this work provides evidence that amyloid core of SHaPrP fibrils is involved in hydrophobic region of residue 108-127 and helical region of residue 172-194 and 200-227.
|
|---|
