Protein Folding and Chaperones


Proteins fold via specific pathways to achieve their native structure. Protein structures are, however, inherently unstable, hence folding and unfolding are in equilibrium. Protein instability is a major concern inside the cell. Specialised proteins called molecular chaperones are, therefore, required to assist proteins in folding and to prevent aggregation of folding intermediates. Many different classes of chaperones are conserved throughout all kingdoms of life, many of which are known as heat shock proteins. Chaperones typically recognise hydrophobic patches, but the exact functions and mechanisms of action of the various chaperone classes are very different. The main chaperone classes Hsp70, Hsp90, Hsp100 and chaperonins all depend on ATPase cycles, whose activities are fine‐tuned by co‐chaperones. The molecular understanding of the mechanism of both chaperones and protein folding are key problems in present‐day life sciences and molecular medicine.

Key Concepts

  • Proteins fold via pathways.
  • Protein structures are labile.
  • Protein folding in vivo is assisted by molecular chaperones.
  • Assisted protein folding requires ATP energy.
  • Molecular chaperones are evolutionary conserved.
  • Chaperone activity is controlled by co‐chaperones and co‐factors.
  • Chaperone families differ in structure and function.

Keywords: protein folding; protein stability; molecular chaperones; folding pathways; Hsp70; Hsp90; heat shock response; protein misfolding; intrinsically disordered proteins

Figure 1. Energy landscape describing protein folding and aggregation. The unfolded polypeptide chain moves towards conformations with lower free energy. In this process, the number of available conformations, hence entropy, decreases. The landscape consists of two funnels: one leading to the native state of the protein, the other to an aggregate. Alternatively, the polypeptide may become trapped in a local energy minimum, such as a partially folded state or an oligomer.
Figure 2. Chaperones in the bacterial and eukaryotic cytosol. The ATP‐dependent chaperones of the E. coli and eukaryotic cytosol and the nascent chain binding trigger factor are shown in surface representation. The molecules are shown to scale. GroEL (in complex with GroES 7 mer), Hsp60 family, oligomeric state 14mer, pdb file 1aon; ClpB, Hsp100, hexamer, 1qvr; Hsp90, dimer (Karagöz et al., ); DnaK, Hsp70, monomer, 2kho; trigger factor, no eukaryotic homologues, monomer, 1w26. Nota bene: all chaperones undergo conformational changes upon binding nucleotide, co‐factors and/or substrate. Pictures were made using Pymol.
Figure 3. Protein folding in the cytosol, assisted by ATP‐driven machines. (a) Protein folding in the cytosol of E. coli. The nascent chain emerges from the ribosome, where it meets its first chaperone, trigger factor (TF). TF is ATP‐independent but profits from the ATP‐driven growth of the polypeptide by the ribosome. Proteins may then either fold on their own, or assisted by the Hsp70 system DnaK and its co‐factors (not shown), or assisted by the chaperonin, GroEL (shown in complex with its co‐factor GroES). Nascent polypeptides may travel between chaperonin and DnaK in case they first bind to a chaperone system that fails in folding them. Proteins may unfold again, in particular under stress conditions, which may lead to aggregation. Even large aggregates can be reversed by concerted action of the DnaK system and the Hsp100 chaperone ClpB. The general action of the bacterial Hsp90 homologue HtpG remaines largely elusive. (b) Protein folding in the human cytosol. The processes are similar to those in E. coli, with the following exceptions: (1) The eukaryotic ribosome does not have a trigger factor homologue but functional analogues nascent chain associated complex (NAC; pdb 1tr8) and RAC (pdb 4gni for the Ssz1 ATPase domain in orange and 4gmq for Zuo1 in red). (2) Hsp90 is essential for folding of a subset of substrate, often following Hsp70 action. (3) There is no Hsp100 chaperoning activity. Chaperone pictures are based on the pictures in Figure . Human Hsp70 is depicted as its E. coli counterpart and NAC is represented by the archaeal homologue. TRiC/CCT (pdb 4v94) is the eukaryotic Hsp60 family member. Co‐chaperones are omitted, except for the GroEL‐GroES complex.
Figure 4. The Hsp70 ATPase cycle, as the substrate sees it. Hsp70 consist of an ATPase domain (blue) and a substrate binding domain that has a substrate holding segment (red) and a lid segment (green). ATP binding triggers opening of the substrate binding domain, ATP hydrolysis encloses the substrate. Release of ADP and rebinding of ATP opens Hsp70, leading to release of the substrate that subsequently may fold into the native state. It is not known whether the substrate has a different structure after Hsp70 release than before Hsp70 interaction. Hsp70 chaperone activity is tuned by stimulating ATP hydrolysis by J proteins and by triggering nucleotide exchange. Protein folding/unfolding processes that are not assisted by Hsp70 are indicated by grey arrows, all other processes are indicated by black arrows. The pictures are based on pbd file 2kho of DnaK for the closed ADP conformation and on pdb file 2qxl of yeast Hsp70 sse1 for the open ATP conformation.
Figure 5. Recognition of hydrophobic residues by Hsp70 and Hsp90 during the folding pathway. Directly after translation, the unfolded polypeptide exposes hydrophobic residues that are recognised by Hsp70. In later stages of folding, these form the hydrophobic core of the protein, leaving Hsp70 unable to bind, but exposing scattered hydrophobic residues that allow binding of Hsp90. Finally, the protein adopts its correct fold and does not present surfaces for chaperone binding anymore.


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Further Reading

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Winkler J, Tyedmers J, Bukau B and Mogk A (2012b) Hsp70 targets Hsp100 chaperones to substrates for protein disaggregation and prion fragmentation. Journal of Cell Biology 198: 387–404.

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Sinnige, Tessa, Karagöz, G Elif, and Rüdiger, Stefan GD(Apr 2015) Protein Folding and Chaperones. In: eLS. John Wiley & Sons Ltd, Chichester. [doi: 10.1002/9780470015902.a0005721.pub3]