N-terminal Truncation of Prion Protein Affects Both Formation and Conformation of Abnormal Protease-resistant Prion Protein Generatedin Vitro

Abstract

Transmissible spongiform encephalopathy diseases are characterized by conversion of the normal proteasesensitive host prion protein, PrP-sen, to an abnormal protease-resistant form, PrP-res. In the current study, deletions were introduced into the flexible tail of PrPsen (23-124) to determine if this region was required for formation of PrP-res in a cell-free assay. PrP-res formation was significantly reduced by deletion of residues 34 -94 relative to full-length hamster PrP. Deletion of another nineteen amino acids to residue 113 further reduced the amount of PrP-res formed. Furthermore, the presence of additional proteinase K cleavage sites indicated that deletion to residue 113 generated a protease-resistant product with an altered conformation. Conversion of PrP deletion mutants was also affected by post-translational modifications to PrP-sen. Conversion of unglycosylated PrP-sen appeared to alter both the amount and the conformation of protease-resistant PrPres produced from N-terminally truncated PrP-sen. The N-terminal region also affected the ability of hamster PrP to block mouse PrP-res formation in scrapie-infected mouse neuroblastoma cells. Thus, regions within the flexible N-terminal tail of PrP influenced interactions required for both generating and disrupting PrPres formation.

Essential Role of the Prion Protein N Terminus in Subcellular Trafficking and Half-life of Cellular Prion Protein

“…Prion titers and protease-resistant PrP were about 30-fold lower than in wild type mice, with no histopathology typical for scrapie (44). These results are in line with in vitro cell-free conversion studies performed with hamster PrP c , where a truncated form of PrP c lacking amino acids 32-94 influenced the quantity and the conformation of generated PrP Sc (45). In light of the results reported here, these findings could also be explained by altered intracellular trafficking of the mutated PrP because of the alterations in the N-terminal part.…”

Cyclin-dependent Kinase 5 Phosphorylation of Familial Prion Protein Mutants Exacerbates Conversion into Amyloid Structure

“…These results are consistent with other findings that show the influence of the N terminus of PrP on the conversion of the globular C-terminal region of PrP. Full-length PrP forms oligomers more readily than the N-terminally truncated PrP 90 -231 (28), deletion of amino acids 34 -94 decreases protease-resistant PrP formation in cell-free conversion assays (29), antibody 6A12 raised against amino acids 41-47 of PrP immunoprecipitates only PrP scrapie and not normal cellular PrP from bovine spongiform encephalopathy-and sheep scrapieaffected brains (30), and hydroxylation at proline 44 of PrP leads to the formation of a rigid polyproline II helix (31, 32). The N terminus of PrP is also clearly involved in familial and transmissible prion diseases.…”

Characterization of Truncated Forms of Abnormal Prion Protein in Creutzfeldt-Jakob Disease

“…The pathogenetic role of the anchorless form of PrP in prion disease is unclear. Although cell-free experiments in vitro have shown that anchorless PrP can be converted to PrP Sc (48,49), the absence of the GPI anchor reduces conversion in scrapie-infected cells (50), which led to the general belief that conversion in prion disease involves membrane-bound GPI-linked PrP. Interestingly, an anchorless version of PrP has recently been shown to sustain PrP Sc replication in vivo, although it did not generate any characteristic symptoms in GPI-negative transgenic mice (47).…”