Molecular chaperones definition

  • A large and diverse group of proteins that assist folding/unfolding and the assembly/disassembly of other macromolecular structures. They are not permanent components of the folded structures. 
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Molecular chaperone action

  • Holders bind hydrophobic patches to prevent aggregated formation.
  • Unfolders pull apart non-native interactions using ATP.
  • Folders Rearrange disulphide bonds catalytically. 
  • The function of chaperones are to:
  •                   1. Prevent thermal inactivation and permanent aggregation.
  •                   2. To aid in protein folding/degradation of inactivated protein.

Order of chaperoning, from simple to more complex to specialised

Bacteria - Trigger factor -> DnaK -> GroEL-ES/HTPG

Archae - NAC -> Hsp70/prefoldin -> thermosome

Eukaryotic - NAC/RAC -> Hsp70/prefolding -> TriC/CCT/Hsp90

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Hsp70 - A Holder


ATP binding domain (N terminal)

Substrate binding domain (C terminal)

C terminal domain for allosteric regulation

1.       The J domain protein Hsp40 stimulates the ATPase activity of Hsp70

2.       Hsp70 is now in the high substrate binding state, and Hsp40 can bring the client protein to Hsp70. The lid closes.

3.       Bag1 (a nucleotide exchange factor) replaces the ADP with ATP, opening the lid and releasing the protein, allowing folding to occur.

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Hsp60 aka the chaperonins - An Unfolder

  • In archae it is thermosome, in eukaryotes it is TriC/CCT, in prokaryotes it is GroEL/ES


  • GroEL/ES is heptameric, but the other two are octameric. 
  • GroEL has two heptagelical rings forming a barrell, with each monomer having 3 domains:

                          Apical substrate binding domain

                          A hinge region

                          An equatorial ATP binding region

1. ATP and GroES bind to the apo ring, causing the apical domain to rotate, hiding hydrophobic residues and allowing the client to enter the cavity (the previous protein and ADP is released from the lower ring).

2. After 10-15 seconds (client has refolded in the meantime), ATPase activity kicks in, and the Upper ring ATP --> ADP. 

3. ATP/GRoES bind to the lower ring, releasing the ADP/GroES from the upper ring, and the cycle repeats.

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Prefoldin - A Holder


  • It is heterohexameric and has two alpha and four beta domains.
  • The structure looks like a jellyfish.
  • The main body of the structure is composed of beta strands which are involved in oligomerisation
  • The "tentacles" are coiled coils involved in substrate binding. 
  • The hydrophobic residues at the end of the tentacles bind non native proteins, which can be passed onto chaperonins for unfolding. 
  • In archae, prefoldin often acts as a Hsp70 substitute. 
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Hsp100 - unfolders

  • An exmaple of a Hsp100 system is ClpB in E. Coli.
  • ClpB has four domains:

                   N terminal domain

                  2x AAA+ ATPase domains  with tyrosine loops.

                 A middle domain that docks to DnaK (aka hsp70)

  • It forms a hexameric ring

1. DnaJ (aka hsp40) attaches to the client and mediates the binding of DnaK.

2. DnaK attaches to ClpB at the middle domain.

3. The N terminal domain is used by DnaK to pass the client to ClpB

4. GrpE (a nucleotide exchange factor) releases DnaK, which has stimulates the ATPase activity of ClpB

5. This drives the up and down motion of the loops, threading the protein and disaggregating it. 

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Hsp100 continued

  • In yeast, it is ClbP or Hsp104

Another example of a HSP 100 system...ClPA

1. ClpA disaggregates the polypeptide

2. The polypeptide is transferred to the ClpP peptidase, where it is cleaved into small fragments.

             The structure of ClpP is two heptameric rings. 

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  • Three domains

               N terminal domain that binds ATP

               Middle domain separated from the N terminal by a charged linker

               A C terminal dimerisation domain. 

  • Many co chaperones are involved, which deliver the client protein, regulate the system, or speed up the system.

The Basic mechanism

1. ATP binding triggers N terminal dimerisation of Hsp90

2. Hsp90 hydrolyses ATP

3. ADP is formed and the N terminal domains separate. 

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A Hsp90 system - the steroid hormone receptor

1. HOP attaches to Hsp70 and and the steroid hormone receptor, and then attaches to the MEEVD site of Hsp90 using HOPs TPR domains (the client is transferred to Hsp90, creating the early complex).

2. Immunophilin binds, causing release of HOP-Hsp70 (The intermediate complex).

3. Hsp90 binds to ATP causes N terminal dimerisation, and p23 (sba1 in yeast) binds (The mature complex).

4. Once the steroid hormone binds, ATP hydrolysis occurs and the active steroid hormone receptor is released. 

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Another Hsp90 system - Kinase activation

1. Cdc37 binds Hsp90 dependant kinases and delivers them to Hsp90.

2. ATP binds causing Hsp90 to N terminal dimerase.

3. Aha1 stimulates Hsp90 ATPase activity, causing the release of the activated kinase.

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Hsp27 - A Holder

  • It is present as a large multioligomeric complex.
  • Heat/phosphorylation causes formation of Hsp27 dimers, which are involved in actin stabilisation.
  • They can form complexes with denatured proteins through application of further heat, acting as heat shock proteins.
  • Once the stress has passed the clients can either be refolded or degraded, and sHsp dissociates:

                       To be folded by the Hsp70/Hsp100 systems

                       To be degraded by the Hsp100/protease system

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