Visualizing the ATPase cycle in a protein disaggregating machine: structural basis for substrate binding by ClpB

Mol Cell. 2007 Jan 26;25(2):261-71. doi: 10.1016/j.molcel.2007.01.002.


ClpB is a ring-shaped molecular chaperone that has the remarkable ability to disaggregate stress-damaged proteins. Here we present the electron cryomicroscopy reconstruction of an ATP-activated ClpB trap mutant, along with reconstructions of ClpB in the AMPPNP, ADP, and in the nucleotide-free state. We show that motif 2 of the ClpB M domain is positioned between the D1-large domains of neighboring subunits and could facilitate a concerted, ATP-driven conformational change in the AAA-1 ring. We further demonstrate biochemically that ATP is essential for high-affinity substrate binding to ClpB and cannot be substituted with AMPPNP. Our structures show that in the ATP-activated state, the D1 loops are stabilized at the central pore, providing the structural basis for high-affinity substrate binding. Taken together, our results support a mechanism by which ClpB captures substrates on the upper surface of the AAA-1 ring before threading them through the ClpB hexamer in an ATP hydrolysis-driven step.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Adenine Nucleotides / metabolism
  • Adenosine Triphosphatases / chemistry*
  • Adenosine Triphosphatases / genetics
  • Adenosine Triphosphatases / metabolism*
  • Adenosine Triphosphatases / ultrastructure
  • Bacterial Proteins / chemistry*
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism*
  • Bacterial Proteins / ultrastructure
  • Cryoelectron Microscopy
  • Endopeptidase Clp
  • Enzyme Activation
  • Escherichia coli Proteins / chemistry
  • Escherichia coli Proteins / genetics
  • Escherichia coli Proteins / metabolism
  • Heat-Shock Proteins / chemistry*
  • Heat-Shock Proteins / genetics
  • Heat-Shock Proteins / metabolism*
  • Heat-Shock Proteins / ultrastructure
  • Image Processing, Computer-Assisted
  • Models, Molecular
  • Protein Structure, Quaternary
  • Protein Structure, Tertiary
  • Substrate Specificity
  • Thermus thermophilus / genetics
  • Thermus thermophilus / metabolism*


  • Adenine Nucleotides
  • Bacterial Proteins
  • Escherichia coli Proteins
  • Heat-Shock Proteins
  • Endopeptidase Clp
  • Adenosine Triphosphatases
  • ClpB protein, E coli