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.


Anfinsen CB (1973) Principles that govern the folding of protein chains. Science 181: 223–230.

Becker T, Bottinger L and Pfanner N (2012) Mitochondrial protein import: from transport pathways to an integrated network. Trends in Biochemical Sciences 37: 85–91.

Chen DH, Madan D, Weaver J, et al. (2013) Visualizing GroEL/ES in the act of encapsulating a folding protein. Cell 153: 1354–1365.

Clark PL (2004) Protein folding in the cell: reshaping the folding funnel. Trends in Biochemical Sciences 29: 527–534.

Daggett V and Fersht A (2003) The present view of the mechanism of protein folding. Nature Reviews Molecular Cell Biology 4: 497–502.

Dill KA and Chan HS (1997) From Levinthal to pathways to funnels. Nature Structural Biology 4: 10–19.

Ellis RJ (2007) Protein misassembly: macromolecular crowding and molecular chaperones. Advances in Experimental Medicine & Biology 594: 1–13.

Englander SW, Mayne L and Krishna MM (2007) Protein folding and misfolding: mechanism and principles. Quarterly Reviews of Biophysics 40: 287–326.

Fersht AR and Daggett V (2002) Protein folding and unfolding at atomic resolution. Cell 108: 573–582.

Glover JR and Lindquist S (1998) Hsp104, Hsp70, and Hsp40: a novel chaperone system that rescues previously aggregated proteins. Cell 94: 73–82.

Hipp MS, Park SH and Hartl FU (2014) Proteostasis impairment in protein‐misfolding and ‐aggregation diseases. Trends in Cell Biology 24 (9): 506–514.

Ignatova Z and Gierasch LM (2004) Monitoring protein stability and aggregation in vivo by real‐time fluorescent labeling. Proceedings of the National Academy of Sciences of the United States of America 101: 523–528.

Joerger AC and Fersht AR (2007) Structural biology of the tumor suppressor p53 and cancer‐associated mutants. Advances in Cancer Research 97: 1–23.

Kampinga HH and Craig EA (2010) The HSP70 chaperone machinery: J proteins as drivers of functional specificity. Nature Reviews Molecular Cell Biology 11: 579–592.

Karagöz GE, Duarte AMS, Akoury E, et al. (2014) Hsp90‐Tau complex reveals molecular basis for specificity in chaperone action. Cell 156: 963–974.

Kim YE, Hipp MS, Bracher A, Hayer‐Hartl M and Hartl FU (2013) Molecular chaperone functions in protein folding and proteostasis. Annual Review of Biochemistry 82: 323–355.

Knowles TP, Vendruscolo M and Dobson CM (2014) The amyloid state and its association with protein misfolding diseases. Nature Reviews Molecular Cell Biology 15: 384–396.

Levinthal C (1969) In: Debrunner P, Tsibris J and Münck E, (eds). Mössbauer Spectroscopy in Biological Systems, pp. 22–24. Urbana, IL: University of Illinois Press.

Li J, Soroka J and Buchner J (2012) The Hsp90 chaperone machinery: conformational dynamics and regulation by co‐chaperones. Biochimica et Biophysica Acta 1823: 624–635.

Mayer MP (2013) Hsp70 chaperone dynamics and molecular mechanism. Trends in Biochemical Sciences 38: 507–514.

Mayor U, Guydosh NR, Johnson CM, et al. (2003) The complete folding pathway of a protein from nanoseconds to microseconds. Nature 421: 863–867.

Merz F, Boehringer D, Schaffitzel C, et al. (2008) Molecular mechanism and structure of Trigger Factor bound to the translating ribosome. EMBO Journal 27: 1622–1632.

Morimoto RI and Cuervo AM (2014a) Proteostasis and the aging proteome in health and disease. Journals of Gerontology. Series A, Biological Sciences and Medical Sciences 69 (Suppl 1): S33–S38.

Oldfield CJ and Dunker AK (2014) Intrinsically disordered proteins and intrinsically disordered protein regions. Annual Review of Biochemistry 83: 553–584.

Preissler S and Deuerling E (2012) Ribosome‐associated chaperones as key players in proteostasis. Trends in Biochemical Sciences 37: 274–283.

Rapoport TA (2007) Protein translocation across the eukaryotic endoplasmic reticulum and bacterial plasma membranes. Nature 450: 663–669.

Rüdiger S, Germeroth L, Schneider‐Mergener J and Bukau B (1997) Substrate specificity of the DnaK chaperone determined by screening cellulose‐bound peptide libraries. EMBO Journal 16: 1501–1507.

Saibil H (2013) Chaperone machines for protein folding, unfolding and disaggregation. Nature Reviews Molecular Cell Biology 14: 630–642.

Saibil HR, Fenton WA, Clare DK and Horwich AL (2013) Structure and allostery of the chaperonin GroEL. Journal of Molecular Biology 425: 1476–1487.

Sakakibara D, Sasaki A, Ikeya T, et al. (2009) Protein structure determination in living cells by in‐cell NMR spectroscopy. Nature 458: 102–105.

Schröder H, Langer T, Hartl FU and Bukau B (1993) DnaK, DnaJ and GrpE form a cellular chaperone machinery capable of repairing heat‐induced protein damage. EMBO Journal 12: 4137–4144.

Vickery LE and Cupp‐Vickery JR (2007) Molecular chaperones HscA/Ssq1 and HscB/Jac1 and their roles in iron‐sulfur protein maturation. Critical Reviews in Biochemistry and Molecular Biology 42: 95–111.

Winkler J, Tyedmers J, Bukau B and Mogk A (2012a) Chaperone networks in protein disaggregation and prion propagation. Journal of Structural Biology 179: 152–160.

Xu Z, Horwich AL and Sigler PB (1997) The crystal structure of the asymmetric GroEL‐GroES‐(ADP)7 chaperonin complex. Nature 388: 741–750.

Further Reading

Dill K and MacCallum J (2012) The protein‐folding problem, 50 years on. Science 338: 1042–1046.

Gershenson A and Gierasch LM (2011) Protein folding in the cell: challenges and progress. Current Opinion in Structural Biology 21 (1): 32–41.

Jarosz DF, Taipale M and Lindquist S (2010) Protein homeostasis and the phenotypic manifestation of genetic diversity: principles and mechanisms. Annual Review of Genetics 44: 189–216.

Kityk R, Kopp J, Sinning I and Mayer MP (2012) Structure and dynamics of the ATP‐bound open conformation of Hsp70 chaperones. Molecular Cell 48: 863–874.

Mayer MP (2010) Gymnastics of molecular chaperones. Molecular Cell 39: 321–331.

Morimoto RI and Cuervo AM (2014b) Proteostasis and the aging proteome in health and disease. Journal of Gerontology: Biological Sciences and Medical Sciences 69 (6): S33–S38. DOI: 10.1093/gerona/glu049.

Southworth DR and Agard DA (2011) Client‐loading conformation of the Hsp90 molecular chaperone revealed in the cryo‐EM structure of the human Hsp90:Hop complex. Molecular Cell 42 (6): 771–781.

Taipale M, Jarosz DF and Lindquist S (2010) HSP90 at the hub of protein homeostasis: emerging mechanistic insights. Nature Reviews. Molecular Cell. Biology 11 (7): 515–528.

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.

Uversky VN (2014) The triple power of D(3): protein intrinsic disorder in degenerative diseases. Frontiers in Bioscience (Landmark Edition) 19: 181–258.

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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]