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Prion and non-prion amyloids of the HET-s prion forming domain

Raimon Sabaté1, Ulrich Baxa, Laura Benkemoun

  • 1Laboratoire de Génétique Moléculaire des Champignons, Institut de Biochimie et de Génétique Cellulaires,UMR 5095 CNRS/Université de Bordeaux 2, 1 rue Camille St Saëns, 33077 Bordeaux cedex, France.

Journal of Molecular Biology
|May 29, 2007
PubMed

Insights

Researchers explored the diversity of HET-s prion protein amyloids formed in vitro. Different pH conditions yielded distinct amyloid structures with varying infectivity, highlighting the role of electrostatic interactions in prion formation.

Area of Science:

  • Biochemistry
  • Structural Biology
  • Mycology

Background:

  • HET-s is a fungal prion protein from Podospora anserina.
  • The HET-s prion-forming domain (HET-s(218-289)) serves as a model for studying amyloid assembly and prion propagation.
  • A structural model exists for the infectious amyloid fold of HET-s(218-289).

Purpose of the Study:

  • To investigate the diversity of HET-s(218-289) amyloids formed in vitro.
  • To characterize the structural and functional differences between amyloids formed under different pH conditions.
  • To elucidate the role of electrostatic interactions in the formation and stability of infectious prion folds.

Main Methods:

  • In vitro amyloid formation assays at different pH values (pH 7 and pH 2).
  • Morphological analysis of amyloid fibrils.
  • Prion infectivity assays.
  • Thioflavin T (ThT) binding assays.
  • Denaturant resistance studies.
  • Kinetic analysis of amyloid assembly.
  • Secondary structure determination (e.g., using spectroscopy).
  • Intrinsic fluorescence measurements.

Main Results:

  • Two distinct types of HET-s(218-289) amyloids were formed at pH 7 and pH 2.
  • Amyloids formed at pH 2 exhibited significantly reduced prion infectivity compared to those formed at pH 7.
  • Differences were observed in ThT binding, denaturant resistance, assembly kinetics, secondary structure, and intrinsic fluorescence between pH 7 and pH 2 amyloids.
  • pH 7 amyloids consisted of bundled or disordered 5 nm fibrils, while pH 2 amyloids formed tightly twisted protofibrils.
  • Electrostatic interactions were identified as critical for the formation and stability of the infectious prion fold.

Conclusions:

  • The in vitro assembly of HET-s(218-289) can produce structurally diverse amyloids with distinct functional properties.
  • Prion infectivity is sensitive to the conditions of amyloid formation, particularly pH.
  • Altered properties of pH 2 amyloids may result from changes in subunit fold or fibrillar stacking due to perturbed electrostatic interactions.

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