The HET-s prion protein of the filamentous fungus Podospora anserina aggregates in vitro into amyloid-like fibrils. |
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Authors: | Suzana Dos Reis Bénédicte Coulary-Salin Vincent Forge Ioan Lascu Jo?l Bégueret Sven J Saupe |
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Affiliation: | les Laboratoires Génétique Moléculaire des Champignons, Institut de Biochimie et de Génétique Cellulaires, UMR 5095 CNRS/Université de Bordeaux 2, 33077 Bordeaux cedex, France. |
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Abstract: | The HET-s protein of Podospora anserina is a fungal prion. This protein behaves as an infectious cytoplasmic element that is transmitted horizontally from one strain to another. Under the prion form, the HET-s protein forms aggregates in vivo. The specificity of this prion model compared with the yeast prions resides in the fact that under the prion form HET-s causes a growth inhibition and cell death reaction when co-expressed with the HET-S protein from which it differs by 13 residues. Herein we describe the purification and initial characterization of recombinant HET-s protein expressed in Escherichia coli. The HET-s protein self-associates over time into high molecular weight aggregates. These aggregates greatly accelerate precipitation of the soluble form. HET-s aggregates appear as amyloid-like fibrils using electron microscopy. They bind Congo Red and show birefringence under polarized light. In the aggregated form, a HET-s fragment of approximately 7 kDa is resistant to proteinase K digestion. CD and FTIR analyses indicate that upon transition to the aggregated state, the HET-s protein undergoes a structural rearrangement characterized by an increase in antiparallel beta-sheet structure content. These results suggest that the [Het-s] prion element propagates in vivo as an infectious amyloid. |
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