Protein Folding

Molecular Chaperones and Heat Shock Proteins

Summary

Molecular chaperones are essential proteins that assist in the folding of other proteins, prevent aggregation, and maintain proteostasis. Heat shock proteins (Hsps) are a major chaperone class upregulated during cellular stress. They are classified by molecular weight (Hsp40, Hsp60, Hsp70, Hsp90, Hsp100) and employ diverse ATP-dependent and ATP-independent mechanisms to maintain the cellular proteome.

Key Points

  • 1Heat shock proteins are classified by size: sHsps, Hsp40, Hsp60, Hsp70, Hsp90, Hsp100
  • 2Hsp70 uses ATP-driven cycles to bind/release substrates through lid opening/closing
  • 3Chaperonins provide isolated folding chambers preventing aggregation
  • 4HSF1 master regulator senses proteostasis stress by competition for chaperones
  • 5Chaperone dysfunction underlies neurodegeneration, cancer, and aging

Molecular chaperones are the guardians of the proteome—proteins that assist in the folding, assembly, and quality control of other proteins without becoming part of the final structure.

Historical Discovery

The term "heat shock proteins" originated from the observation that exposing Drosophila to elevated temperatures induced a characteristic set of proteins. We now know these proteins are:

  • Present constitutively in all cells

    • - Dramatically upregulated during stress (heat, oxidative damage, heavy metals)

  • Essential for normal protein folding even under optimal conditions
  • Highly conserved from bacteria to humans

    • Classification by Molecular Weight

    Small Heat Shock Proteins (sHsps, 12-43 kDa)

    - ATP-independent holdases

  • Form large oligomeric assemblies (12-40 subunits)
  • Bind partially unfolded proteins to prevent aggregation
  • Examples: α-crystallin, Hsp27
  • Act as a first line of defense during acute stress

    • Hsp40 (DnaJ Family)

    - Co-chaperones that work with Hsp70

  • Contain characteristic J-domain for Hsp70 interaction
  • Stimulate Hsp70 ATPase activity 5-10 fold
  • Deliver substrates to Hsp70
  • Provide substrate specificity to the Hsp70 system

    • Hsp60 (Chaperonins)

  • Form large barrel-shaped complexes

    • - Two classes:

    - Group I (GroEL/GroES in bacteria, Hsp60/Hsp10 in mitochondria)

    - Group II (TRiC/CCT in eukaryotic cytosol)

  • Provide isolated folding chambers
  • Essential for folding ~10-15% of cellular proteins

    • Hsp70 (DnaK Family)

    - Central hub of the chaperone network

  • Consist of:

    • - N-terminal ATPase domain (NBD)

    - C-terminal substrate-binding domain (SBD)

    - Lid that traps substrates

  • Recognize hydrophobic segments exposed in unfolded proteins
  • ATP binding opens the lid; ATP hydrolysis closes it

    • Hsp90

    - Specialized for signaling proteins (kinases, transcription factors, hormone receptors)

  • Works late in folding—near-native substrates
  • Requires multiple co-chaperones (Hop, p23, Cdc37, Aha1)
  • Major target for cancer therapy (many oncoproteins are Hsp90 clients)

    • Hsp100/Clp Family

    - Disaggregases that can unfold aggregated proteins

  • Use ATP to thread substrates through a central pore
  • Can rescue proteins from aggregates for refolding or degradation
  • Hsp104 in yeast; not present in metazoans (Hsp70/Hsp40 disaggregase system instead)

    • The Hsp70 Cycle

    The Hsp70 system exemplifies chaperone action:

    1. Substrate Recognition

  • J-domain proteins (Hsp40) deliver substrates
  • Hsp70 binds exposed hydrophobic sequences

    • 2. ATP Hydrolysis

  • J-domain stimulates ATPase activity
  • Lid closes, trapping substrate in high-affinity state
  • Nucleotide exchange factors (NEFs: Bag1, Hsp110) release ADP

    • 3. ATP Binding and Release

  • ATP binding opens the lid
  • Substrate is released for:

    • - Productive folding

    - Another chaperone cycle

    - Transfer to downstream chaperones (Hsp90, chaperonins)

    The Chaperonin Mechanism

    GroEL/GroES (bacterial chaperonin) provides the best-understood folding chamber:

    Structure

  • GroEL: Two heptameric rings stacked back-to-back
  • GroES: Heptameric "lid" that caps one ring
  • Together create an enclosed ~85Å cavity

    • Folding Cycle

  • Substrate binds to hydrophobic apical domains of open ring
  • ATP and GroES bind, triggering conformational change
  • Cavity becomes hydrophilic—releases substrate into chamber
  • Protein folds in isolation for ~10 seconds
  • ATP hydrolysis in opposite ring triggers release
  • Substrate released—folded or returned for another cycle

    • Anfinsen Cage Model

    The chamber provides an environment where protein can fold without:

  • Intermolecular aggregation
  • Aberrant interactions
  • Influence of crowded cytoplasm

    • Heat Shock Response

    Stress triggers a coordinated upregulation of chaperones:

    Regulation

    - HSF1 (Heat Shock Factor 1) is the master regulator

  • Under normal conditions: HSF1 is sequestered by Hsp70/Hsp90
  • Under stress: Chaperones release HSF1 to handle misfolded proteins
  • Free HSF1 trimerizes and activates heat shock genes

    • Heat Shock Elements (HSEs)

  • DNA sequences (nGAAn repeats) in promoters of heat shock genes
  • HSF1 trimers bind HSEs and activate transcription
  • Results in rapid induction of chaperone expression

    • Disease Connections

    Chaperone dysfunction is linked to numerous pathologies:

    - Neurodegenerative diseases: Overwhelmed chaperone capacity leads to aggregation

    - Cancer: Tumor cells are "addicted" to Hsp90 for mutant oncoprotein stabilization

    - Aging: Proteostasis network declines, contributing to age-related dysfunction

    - Cataracts: α-crystallin mutations cause lens protein aggregation

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