NR APJW
AU Maxson,L.; Wong,C.; Herrmann,L.M.; Caughey,B.W.; Baron,G.S.
TI A solid-phase assay for identification of modulators of prion protein interactions
QU Analytical Biochemistry 2003 Dec 1; 323(1): 54-64
PT journal article
AB The progression of the transmissible spongiform encephalopathies (TSEs) is characterized in part by accumulation of a proteinase K-resistant form of the prion protein, which has been converted from the endogenous, proteinase K-sensitive form. This conversion reaction provides a target for possible anti-TSE strategies. We have adapted a cell-free conversion reaction to a high-throughput, solid-phase format that can be used to screen possible therapeutic compounds for inhibitory activity or to illuminate inhibition and conversion mechanisms. The solid-phase assay was compatible with reactions performed under a variety of conditions. Using this assay, we report that phthalocyanine tetrasulfonate, a known modulator of conversion, inhibited conversion by interfering with binding between the protease-sensitive and the protease-resistant forms of the prion protein. A biotinylated form of the protease-sensitive prion protein was successfully converted to the protease-resistant isoform in the solid-phase assay, indicating that biotinylation provides a nonisotopic labeling strategy for large-scale screens.
MH Animals; Biotinylation; Brain; *Drug Design; Drug Evaluation, Preclinical/*methods; Endopeptidase K/pharmacology; Hamsters; Indoles/pharmacology; Prion Diseases/drug therapy; Prions/*analysis/*antagonists & inhibitors/chemistry; Protein Binding/drug effects; Protein Isoforms/analysis
AD Rocky Mountain Laboratories, NIAID, NIH, 903 S. 4th St., Hamilton, MT 59840, USA
SP englisch
PO USA