NR ATWR
AU Chen,C.Y.; Rojanatavorn,K.; Clark,A.C.; Shih,J.C.H.
TI Characterization and enzymatic degradation of Sup35NM, a yeast prion-like protein
QU Protein Science 2005 Sep; 14(9): 2228-35
IA http://www.proteinscience.org/cgi/content/full/14/9/2228
PT journal article
AB Transmissible spongiform encephalopathies (TSEs) are believed to be caused by an unconventional infectious agent, the prion protein. The pathogenic and infectious form of prion protein, PrPsc, is able to aggregate and form amyloid fibrils, very stable and resistant to most disinfecting processes and common proteases. Under specific conditions, PrPsc in bovine spongiform encephalopathy (BSE) brain tissue was found degradable by a bacterial keratinase and some other proteases. Since this disease-causing prion is infectious and dangerous to work with, a model or surrogate protein that is safe is needed for the in vitro degradation study. Here a nonpathogenic yeast prion-like protein, Sup35NM, cloned and overexpressed in E. coli, was purified and characterized for this purpose. Aggregation and deaggregation of Sup35NM were examined by electron microscopy, gel electrophoresis, Congo red binding, fluorescence, and Western blotting. The degradation of Sup35NM aggregates by keratinase and proteinase K under various conditions was studied and compared. These results will be of value in understanding the mechanism and optimization of the degradation process.
MH Amino Acid Sequence; Blotting, Western; Congo Red/metabolism; Endopeptidase K/*metabolism; Escherichia coli/genetics; Fluorescence; Microscopy, Electron/methods; Molecular Sequence Data; Peptide Fragments/chemistry/isolation & purification/metabolism; Peptide Hydrolases/*metabolism; Prions/*chemistry/isolation & purification/*metabolism; Research Support, U.S. Gov't, Non-P.H.S.; Saccharomyces cerevisiae Proteins/*chemistry/isolation &; purification/*metabolism; Temperature; Time Factors
AD Department of Poultry Science, North Carolina State University, Raleigh, NC 27695-7608, USA. cchen8@ncsu.edu
SP englisch
PO USA