NR AMCK
AU Vorberg,I.; Chan,K.; Priola,S.A.
TI Deletion of beta-strand and alpha-helix secondary structure in normal prion protein inhibits formation of its protease-resistant isoform
QU Journal of Virology 2001 Nov; 75(21): 10024-32
PT journal article
AB A fundamental event in the pathogenesis of transmissible spongiform encephalopathies (TSE) is the conversion of a normal, proteinase K-sensitive, host-encoded protein, PrP-sen, into its protease-resistant isoform, PrPres. During the formation of PrPres, PrP-sen undergoes conformational changes that involve an increase of beta-sheet secondary structure. While previous studies in which PrP-sen deletion mutants were expressed in transgenic mice or scrapie-infected cell cultures have identified regions in PrP-sen that are important in the formation of PrPres, the exact role of PrP-sen secondary structures in the conformational transition of PrP-sen to PrPres has not yet been defined. We constructed PrP-sen mutants with deletions of the first beta-strand, the second beta-strand, or the first alpha-helix and tested whether these mutants could be converted to PrPres in both scrapie-infected neuroblastoma cells (Sc(+)-MNB cells) and a cell-free conversion assay. Removal of the second beta-strand or the first alpha-helix significantly altered both processing and the cellular localization of PrP-sen, while deletion of the first beta-strand had no effect on these events. However, all of the mutants significantly inhibited the formation of PrPres in Sc(+)-MNB cells and had a greatly reduced ability to form protease-resistant PrP in a cell-free assay system. Thus, our results demonstrate that deletion of the beta-strands and the first alpha-helix of PrP-sen can fundamentally affect PrPres formation and/or PrP-sen processing.
MH Animal; Endopeptidase K/*pharmacology; Mice; Mutagenesis, Site-Directed; Prions/*chemistry/metabolism; *Protein Structure, Secondary; Tumor Cells, Cultured
AD Laboratory of Persistent Viral Diseases, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Hamilton, Montana 59840, USA
SP englisch
PO USA