A novel in vitro filter trap assay identifies tannic acid as an amyloid aggregation inducer for HET-s.
In this work we present an easy and low cost in vitro filter trap assay to quickly identify direct actors on amyloid prion aggregation. We chose the recombinant purified prion protein HET-s from Podospora anserina as a reference. HET-s was labelled with a fluorophore prior to aggregation assays in a 96 well micro-array system. Aggregation assays were carried out in presence of a number of chemical compounds, followed by a filter trap assay through a cellulose acetate membrane and the straight detection of retained fluorescent amyloid fibres. We tested 22 chemical compounds from which 11 have already been described to affect various prions and other amyloid proteins. Four compounds showed direct effects on the aggregation of HET-s. ZnCl seemed to prevent the formation of amyloid fibres. Puzzlingly, three members of the group of tannins (tannic acid, epigallocatechin and epigallocatechin-gallate) had accelerant properties on amyloid aggregation. Resistance of the prion forming domain (PFD) in Proteinase K proteolysis assays underlined that tannic acid favours amyloid fibre formation of HET-s.
Boyé-Harnasch
Mona
M
Institut de Biochimie et Génétique Cellulaire, 1 rue Camille St Saëns, CNRS UMR 5095, 33077 Bordeaux Cedex, France.
Cullin
Christophe
C
D000682:Amyloid / Q000096:biosynthesis / Q000187:drug effects / Q000378:metabolism*; D001709:Biotechnology / Q000295:instrumentation / Q000379:methods*; D019286:Endopeptidase K / Q000378:metabolism; D005656:Fungal Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D048468:Podospora / Q000235:genetics / Q000378:metabolism; D011328:Prions / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D013634:Tannins / Q000494:pharmacology*
D000682:Amyloid; D005656:Fungal Proteins; D011328:Prions; D011994:Recombinant Proteins; D013634:Tannins; D019286:Endopeptidase K