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Identification of a consensus motif in substrates bound by a Type I Hsp40

Pradeep Kota1, Daniel W Summers, Hong-Yu Ren

  • 1Department of Biochemistry and Biophysics, University of North Carolina, Chapel Hill, NC 27599, USA.

Proceedings of the National Academy of Sciences of the United States of America
|June 25, 2009
PubMed

Insights

Molecular chaperones like yeast Ydj1 recognize misfolded proteins via a specific amino acid motif. This discovery aids understanding of protein aggregation in diseases and Hsp40 chaperone function.

Area of Science:

  • Molecular biology
  • Biochemistry
  • Structural biology

Background:

  • Protein aggregation causes many conformational diseases.
  • Molecular chaperones, including Hsp40 family members (DnaJ homologs), bind misfolded proteins to prevent toxic aggregation.
  • Type I Hsp40s are effective in suppressing aggregation and refolding damaged proteins, but their recognition mechanism for nonnative polypeptides remains unclear.

Purpose of the Study:

  • To computationally identify and experimentally validate a unique motif recognized by the Type I Hsp40 chaperone yeast Ydj1.
  • To elucidate the molecular mechanism of substrate recognition by Ydj1.
  • To propose a model for Ydj1's interaction with nonnative polypeptides and its role in presenting them to Hsp70 chaperones.

Main Methods:

  • Computational motif identification.
  • Site-directed mutagenesis to verify motif recognition.
  • Yeast proteome screening to identify proteins containing the motif.
  • Biochemical assays to study substrate binding and chaperone interactions.

Main Results:

  • A unique motif with the consensus sequence GX[LMQ]{P}X{P}{CIMPVW} was identified as selectively recognized by Ydj1p.
  • Site-directed mutagenesis confirmed that Ydj1p's substrate binding is dependent on recognizing this motif.
  • A yeast proteome screen revealed that multiple proteins contain this motif, often with multiple occurrences separated by variable amino acid linkers.
  • A 2-site peptide-binding model for Ydj1p was proposed.

Conclusions:

  • Ydj1p recognizes misfolded proteins through a specific amino acid motif, GX[LMQ]{P}X{P}{CIMPVW}.
  • This motif recognition is crucial for Ydj1p's substrate binding and function in preventing protein aggregation.
  • The findings support a 2-site binding model and provide insights into Ydj1p's mechanism for presenting substrates to Hsp70 chaperones.
  • Given the functional interchangeability of yeast Ydj1p and its human ortholog Hdj2, these results have implications for understanding human conformational diseases.

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