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Nucleotide-dependent substrate recognition by the AAA+ HslUV protease

Randall E Burton1, Tania A Baker, Robert T Sauer

  • 1Department of Biology, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, Massachusetts 02139, USA.

Nature Structural & Molecular Biology
|February 8, 2005
PubMed

Insights

The HslUV (ClpYQ) protease complex efficiently degrades Arc repressor, identifying a specific N-terminal sequence as a key degradation tag. This finding advances understanding of ATP-dependent protein degradation mechanisms.

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Proteolysis

Background:

  • ATP-dependent protein degradation is crucial for cellular regulation.
  • The AAA+ HslU ATPase and HslV peptidase form the HslUV (ClpYQ) protease complex.
  • Understanding substrate recognition by HslUV has been limited by a lack of suitable substrates.

Purpose of the Study:

  • To identify and characterize substrates for the HslUV (ClpYQ) protease complex.
  • To elucidate the mechanism of substrate engagement and degradation by HslUV.
  • To investigate the role of ATP in substrate binding and unfolding.

Main Methods:

  • Degradation assays using Arc repressor and its variants.
  • Fusion protein construction to test degradation tags.
  • Peptide-binding experiments with HslU.
  • Investigating the role of ATP and Mg(2+) in binding.

Main Results:

  • HslUV efficiently degrades Arc repressor, particularly under heat shock conditions.
  • Degradation is mediated by a specific N-terminal sequence of Arc repressor.
  • This N-terminal sequence acts as a direct binding site for the HslU ATPase.
  • ATP and Mg(2+) are required for strong binding of the degradation tag to HslU.
  • Fusion of the tag to a stable protein confers HslUV-mediated degradation.

Conclusions:

  • HslUV functions as a potent protein unfoldase.
  • Substrate engagement by HslU occurs after ATP binding.
  • The identified N-terminal sequence is a functional degradation tag for HslUV.

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