title: Contribution of specific residues of the β-solenoid fold to HET-s prion function, amyloid structure and stability.
abstract: The [Het-s] prion of the fungus Podospora anserina represents a good model system for studying the structure-function relationship in amyloid proteins because a high resolution solid-state NMR structure of the amyloid prion form of the HET-s prion forming domain (PFD) is available. The HET-s PFD adopts a specific β-solenoid fold with two rungs of β-strands delimiting a triangular hydrophobic core. A C-terminal loop folds back onto the rigid core region and forms a more dynamic semi-hydrophobic pocket extending the hydrophobic core. Herein, an alanine scanning mutagenesis of the HET-s PFD was conducted. Different structural elements identified in the prion fold such as the triangular hydrophobic core, the salt bridges, the asparagines ladders and the C-terminal loop were altered and the effect of these mutations on prion function, fibril structure and stability was assayed. Prion activity and structure were found to be very robust; only a few key mutations were able to corrupt structure and function. While some mutations strongly destabilize the fold, many substitutions in fact increase stability of the fold. This increase in structural stability did not influence prion formation propensity in vivo. However, if an Ala replacement did alter the structure of the core or did influence the shape of the denaturation curve, the corresponding variant showed a decreased prion efficacy. It is also the finding that in addition to the structural elements of the rigid core region, the aromatic residues in the C-terminal semi-hydrophobic pocket are critical for prion propagation. Mutations in the latter region either positively or negatively affected prion formation. We thus identify a region that modulates prion formation although it is not part of the rigid cross-β core, an observation that might be relevant to other amyloid models.
authors.affiliation: Institut de Biochimie et de Génétique Cellulaire, Unité Mixte de Recherche 5095, Centre National de la Recherche Scientifique Université de Bordeaux, Bordeaux, France.
authors.forename: Asen
authors.initials: A
authors.lastname: Daskalov
authors.affiliation: Laboratory of Physical Chemistry, Eidgenössische Technische Hochschule (ETH) Zürich, Zürich, Switzerland.
authors.forename: Matthias
authors.initials: M
authors.lastname: Gantner
authors.affiliation: Laboratory of Physical Chemistry, Eidgenössische Technische Hochschule (ETH) Zürich, Zürich, Switzerland.
authors.forename: Marielle Aulikki
authors.initials: MA
authors.lastname: Wälti
authors.affiliation: Laboratory of Physical Chemistry, Eidgenössische Technische Hochschule (ETH) Zürich, Zürich, Switzerland.
authors.forename: Thierry
authors.initials: T
authors.lastname: Schmidlin
authors.affiliation: Laboratory of Physical Chemistry, Eidgenössische Technische Hochschule (ETH) Zürich, Zürich, Switzerland.
authors.forename: Celestine N
authors.initials: CN
authors.lastname: Chi
authors.affiliation: Laboratory of Physical Chemistry, Eidgenössische Technische Hochschule (ETH) Zürich, Zürich, Switzerland.
authors.forename: Christian
authors.initials: C
authors.lastname: Wasmer
authors.affiliation: Laboratory of Physical Chemistry, Eidgenössische Technische Hochschule (ETH) Zürich, Zürich, Switzerland.
authors.forename: Anne
authors.initials: A
authors.lastname: Schütz
authors.affiliation: Institut de Biochimie et de Génétique Cellulaire, Unité Mixte de Recherche 5095, Centre National de la Recherche Scientifique Université de Bordeaux, Bordeaux, France.
authors.forename: Johanna
authors.initials: J
authors.lastname: Ceschin
authors.affiliation: Institut de Biochimie et de Génétique Cellulaire, Unité Mixte de Recherche 5095, Centre National de la Recherche Scientifique Université de Bordeaux, Bordeaux, France.
authors.forename: Corinne
authors.initials: C
authors.lastname: Clavé
authors.affiliation: Institut de Biochimie et de Génétique Cellulaire, Unité Mixte de Recherche 5095, Centre National de la Recherche Scientifique Université de Bordeaux, Bordeaux, France.
authors.forename: Sandra
authors.initials: S
authors.lastname: Cescau
authors.affiliation: Laboratory of Physical Chemistry, Eidgenössische Technische Hochschule (ETH) Zürich, Zürich, Switzerland.
authors.forename: Beat
authors.initials: B
authors.lastname: Meier
authors.affiliation: Laboratory of Physical Chemistry, Eidgenössische Technische Hochschule (ETH) Zürich, Zürich, Switzerland.
authors.forename: Roland
authors.initials: R
authors.lastname: Riek
authors.affiliation: Institut de Biochimie et de Génétique Cellulaire, Unité Mixte de Recherche 5095, Centre National de la Recherche Scientifique Université de Bordeaux, Bordeaux, France.
authors.forename: Sven J
authors.initials: SJ
authors.lastname: Saupe
mesh_terms: D000409:Alanine / Q000737:chemistry; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000682:Amyloid / Q000737:chemistry*; D016229:Amyloid beta-Peptides / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D017124:Conserved Sequence; D004735:Energy Transfer; D005656:Fungal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D058851:GPI-Linked Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011328:Prions / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D017510:Protein Folding; D055550:Protein Stability; D017433:Protein Structure, Secondary; D058767:Protein Unfolding; D016415:Sequence Alignment
chemical_list: D000682:Amyloid; D016229:Amyloid beta-Peptides; D005656:Fungal Proteins; D058851:GPI-Linked Proteins; C083891:HET-S protein, Podospora anserina; D050505:Mutant Proteins; C501125:PRND protein, human; D010446:Peptide Fragments; D011328:Prions; D000409:Alanine