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Purification of Hsp104, a Protein Disaggregase
07:17

Purification of Hsp104, a Protein Disaggregase

Published on: September 30, 2011

Propagation of the yeast prion-like [psi+] determinant is mediated by oligomerization of the SUP35-encoded

S V Paushkin1, V V Kushnirov, V N Smirnov

  • 1Institute of Experimental Cardiology, Cardiology Research Center, Moscow, Russia.

The EMBO Journal
|June 17, 1996
PubMed

Insights

Yeast Sup35p protein forms prion-like aggregates, causing the [psi+] nonsense suppressor trait. This aggregation is essential for [psi+] inheritance, with chaperone Hsp104p potentially regulating its propagation.

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Yeast Genetics

Background:

  • The Sup35p protein in Saccharomyces cerevisiae is homologous to mammalian polypeptide chain release factor 3 (eRF3).
  • It has been hypothesized that Sup35p can adopt a self-propagating conformation, similar to mammalian prions, leading to the [psi+] nonsense suppressor determinant.
  • This [psi+] determinant is inherited in a non-Mendelian fashion.

Purpose of the Study:

  • To confirm the prion-like nature of the [psi+] determinant in yeast.
  • To elucidate the molecular mechanisms underlying [psi+] propagation and inheritance.
  • To investigate the role of Sup35p aggregation and chaperone proteins in this process.

Main Methods:

  • Investigating Sup35p molecular interactions in [psi+] and [psi-] cells using techniques to detect protein-protein interactions.
  • Analyzing the formation of high molecular weight Sup35p aggregates in [psi+] cells via biochemical assays.
  • Assessing the impact of N-terminally altered Sup35p and Hsp104p chaperone overexpression on the [psi+] phenotype and translation termination.

Main Results:

  • Sup35p molecules interact via their N-terminal domains specifically in [psi+] cells, which is crucial for [psi+] propagation.
  • Sup35p forms high molecular weight aggregates in [psi+] cells, sequestering most of the protein and inhibiting its activity, leading to the nonsense suppressor phenotype.
  • N-terminally altered Sup35p or overexpression of Hsp104p disrupts Sup35p aggregation, causing an antisuppressor phenotype and partially solubilizing aggregates, respectively.

Conclusions:

  • The [psi+] determinant in yeast exhibits prion-like behavior, driven by self-propagating conformational changes in Sup35p.
  • Sup35p aggregation is the direct cause of the [psi+] nonsense suppressor phenotype.
  • Hsp104p chaperone may play a critical role in maintaining stable [psi+] inheritance by regulating Sup35p aggregate dynamics.

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