protein folding & degradation

Comprehensive vocabulary flashcards covering protein folding mechanisms, chaperone systems, degradation pathways, and clinical conditions like Cystic Fibrosis, Amyloidosis, and Prion diseases.

protein folding

the process by which a protein goes from an unfolded native state to form a three-dimensional structure

• occurs through progressive stabilization of intermediate states until the lowest‑energy conformation is reached

• begins during translation

• crucial for the protein's functionality

protein folding regulation

the factors that determine how a protein folds into its native conformation

• the spatial information encoded in the amino acid sequence (primary structure)

• assisted by regulatory proteins (molecular chaperones) that help other proteins fold

chaperone proteins

intracellular protein complexes that prevent protein aggregation during synthesis and permit refolding of misfolded proteins in a protected environment

• consume ATP during the folding process

• constitutively expressed → prevent denaturation and misfolding

• expression increases with heat, chemical stress, and hypoxia

• major families: Hsp60, Hsp70, Hsp90

heat shock proteins (hsp)

stress‑induced chaperone proteins that assist folding during and after translation and stabilize/refold damaged or unfolded proteins

• expression increases during heat, chemical stress, and hypoxia

• prevent protein denaturation and aggregation

• require ATP

• include Hsp60, Hsp70, Hsp90

hsp60 system

a barrel-shaped chaperone system required for the correct folding of cellular proteins that do not fold spontaneously and for aiding refolding after crossing membranes

• located in the mitochondria and cytosol (depending on isoform)

• unfolded polypeptide enters the chamber → cap closes → folding occurs

  • creates a hydrophilic environment for proper folding

  • uses ATP to change conformation and release folded protein

hsp70 system

an ATP‑dependent chaperone system that aids in the stabilization of extended chains, membrane translocation, and regulation of the heat shock response

• located in the cytosol and RER

• prevents aggregation of unfolded proteins

• requires ATP hydrolysis for binding and release

• works with co‑chaperones (e.g., Hsp40)

hsp90 system

a molecular chaperone system involved in the binding and stabilization or regulation of steroid receptors

• located in the cytosol

• essential for activation of receptors containing zinc finger motifs

• assists maturation of signaling proteins and kinases

• requires ATP

denaturation

a process where the three-dimensional structure of a protein is changed by breaking noncovalent and disulfide bonds while the primary structure remains intact

• peptide backbone is NOT broken → primary structure preserved

• denatured proteins usually cannot refold and often precipitate

• denaturation causes loss of biological activity

• dietary proteins are denatured in the stomach due to low pH

  • stomach acidity is not strong enough to cleave peptide bonds

laboratory denaturation methods

chemical and physical treatments used to disrupt protein structure by breaking stabilizing bonds

• heat, 5–10 M urea, high salt → break hydrogen bonds

• strong acids or bases → break ionic bonds

• 1 - 2% SDS (detergent) → disrupt hydrophobic interactions

• thiol compounds (β‑mercaptoethanol, 2‑mercaptoethanol) → reduce disulfide bonds

  • disulfide bonds of insulin → cleaved in the liver during degradation

protein misfolding

a change in a protein’s 3D conformation that causes loss of function and increases its tendency to aggregate

• mechanism: altered conformation → ↑ self‑association → ↑ aggregation → aggregates resist clearance

  • fail to fold correctly (e.g., cystic fibrosis, Marfan syndrome, ALS)

  • be unstable and lose normal function (many cancers)

  • fail to be correctly trafficked (familial hypercholesterolemia, α‑1 antitrypsin deficiency)

  • form insoluble toxic aggregates (Alzheimer’s, Parkinson’s, type II diabetes)

intracellular protein degradation

continuous turnover (synthesis and degradation) of cellular proteins through lysosomal and proteasomal pathways

• lysosomal proteases (cathepsins) degrade proteins at pH ~5

• proteasomes degrade ubiquitin‑tagged proteins in an ATP‑dependent process

• released amino acids are reused for new protein synthesis or energy

lysosomal degradation

acid‑dependent breakdown of extracellular material, membrane components, and cytoplasmic proteins within lysosomes

• lysosome = trash collector of the cell

  • lysosomal enzymes active at pH < 4.8

  • nucleases, proteases, & phosphatases

• pathways:

  • phagocytosis → defense

  • endocytosis → membrane recycling

  • autophagy → degradation of cytoplasmic proteins & organelles

autophagy

aka 'self-eating' ; it is the lysosomal degradation of cytoplasmic proteins and organelles regulated by Atg genes

• nutrients are adequate → mTOR inhibits autophagy

• nutrient starvation, hypoxia, heat → ↓ mTOR → ↑ Atg → ↑ autophagy

proteasome-mediated degradation / ubiquitin–proteasome pathway

ATP‑dependent destruction of ubiquitin‑tagged misfolded, damaged, mutant, or short‑lived regulatory proteins within proteasomes

• requires ubiquitination (E1/E2/E3)

• eliminates:

  • misfolded proteins

  • mutant or damaged proteins

  • normal short‑lived regulatory proteins (mitotic cyclins, transcription factors, tumor suppressors, proto‑oncogenes)

• regulates: cell cycle, DNA repair, transcription, stress response, quality control, signal transduction, immunity

proteasomes

large, ATP‑dependent protease complexes that degrade proteins covalently linked to ubiquitin into peptides

• structure: 20S catalytic core + two 19S caps

• produces 7–9 aa peptides → released → further degraded to amino acids

• central to protein quality control and regulation

ubiquitin activating enzyme (E1)

ATP‑dependent enzyme that activates ubiquitin by forming a high‑energy thioester bond

• initiates the ubiquitination process

• transfers activated ubiquitin → ubiquitin conjugating enzyme (E2)

ubiquitin conjugating enzyme (E2)

enzyme that receives activated ubiquitin from E1 and carries it to E3 for substrate targeting

• accepts ubiquitin from E1

• holds ubiquitin in a thioester linkage

• interacts with E3 ligase

ubiquitin ligase (E3)

enzyme that catalyzes covalent attachment of ubiquitin to lysine residues on target proteins

• forms an isopeptide bond between ubiquitin’s C‑terminus and a lysine on the target protein

• recognizes the specific substrate

• transfers ubiquitin from E2 → substrate

long‑lived proteins

proteins that have a prolonged half-life in the cell & are more resistant to degradation

• includes cytoskeletal proteins, housekeeping proteins, stable protein complexes, organelle proteins

• degraded via autophagy → lysosomes

short‑lived proteins

proteins that have a rapid turnover rate and are quickly degraded

• includes transcription factors, signaling proteins, metabolic regulators

• degraded via ubiquitin–proteasome system

PEST‑sequence proteins

short‑lived proteins containing Pro‑Glu‑Ser‑Thr–rich sequences that target them for rapid proteasomal degradation (fast turnover)

• degraded via ubiquitin–proteasome pathway

proteasome inhibitors

drugs that block proteasomal degradation causing cell cycle arrest and apoptosis

• bortezomib, carfilzomib

• inhibit proteasome → accumulation of misfolded proteins

  • induce G2–M arrest

• used in multiple myeloma &mantle cell lymphoma

human papilloma virus (HPV) E6 & E7

proteins that activate host E3 ligases to destroy the p53 tumor suppressor protein and DNA repair genes

• HPV16/18 produce E6 and E7 proteins

  • E6 recruits a host E3 ligase to ubiquitinate p53 → p53 degradation → loss of tumor suppression

  • E7 binds Rb, releasing E2F → increased transcription → cell cycle activation

• results in uncontrolled proliferation and contributes to >90% of cervical carcinomas

cystic fibrosis (CF)

an autosomal recessive genetic disease caused by a defect in the CFTR gene due to a Phe508 deletion (most common) on chromosome 7

• Phe508 deletion → misfolded CFTR retained in RER → not transported to membrane

  • ↓ Cl⁻ and H₂O secretion → ↑ intracellular Cl⁻ → ↑ Na⁺ reabsorption → ↑ H₂O reabsorption

  • results in abnormally thick mucus in lungs & GI tract

  • sweat glands: ↓ Cl⁻ reabsorption → ↑ Cl⁻ in sweat

• causes more negative transepithelial potential difference

cystic fibrosis transmembrane conductance regulator (CFTR)

a protein that functions as an ATP-gated chloride channel in epithelial cells

• secretes Cl⁻ in the lungs & GI tract

• reabsorbs Cl⁻ in sweat glands

cystic fibrosis – diagnosis

elevated chloride concentration in pilocarpine‑induced sweat test

• CFTR dysfunction → ↓ Cl⁻ reabsorption in sweat glands

• results in ↑ Cl⁻ in sweat

cystic fibrosis – complications

multisystem consequences of CFTR dysfunction and thick mucus secretions

• recurrent respiratory infections

• pancreatic insufficiency → malabsorption, steatorrhea

• fat‑soluble vitamin deficiencies (A, D, E, K)

• liver disease

• infertility in men (absence of vas deferens)

• subfertility in women (thick cervical mucus)

cystic fibrosis – treatment

combination therapy that corrects misfolding and improves chloride transport in CFTR used specifically for Phe508 deletion, the most common mutation

• lumacaftor → corrects misfolded CFTR and improves trafficking to membrane

• ivacaftor → opens CFTR Cl⁻ channels to enhance chloride transport

Prion

An infective protein that does not contain DNA or RNA and causes normal proteins to misfold into an infectious form.

PrPc

The normal, alpha-helical form of the Prion protein.

PrPsc

The misfolded, infectious form of the Prion protein characterized by abundant $\beta$-pleated sheets.

Spongiform change

The irreversible brain damage resulting from the non-degradable accumulation of PrPsc, leading to ataxia and dementia.

Creutzfeldt-Jacob disease

A human prion disease characterized by rapidly progressive dementia and startle myoclonus.

Scrapie

A prion disease found in sheep.

Mad cow disease

The common name for Bovine spongiform encephalopathy, which can be transmitted from cows to humans.

Amyloidosis

A condition resulting from extracellular deposits of fibrillar proteins (amyloid) that cause tissue damage and functional compromise.

Congo red

A dye used to identify amyloid deposits; it produces red-green dichroism (birefringence) under polarized microscopy.

Amyloid fibril structure

Nonbranching fibrils, $7.5$ to $10\,nm$ in diameter, formed of $\beta$-sheet polypeptide chains wound together.

AL (amyloid light chain) protein

Amyloid protein produced by plasma cells and made up of complete immunoglobulin light chains.

AA (amyloid-associated) fibril

A non-immunoglobulin amyloid protein derived from the serum precursor SAA protein.

SAA (serum amyloid-associated) protein

A precursor protein synthesized in the liver that can form AA fibrils.

A$\beta$ amyloid

The type of amyloid found in the cerebral lesions of Alzheimer disease.

Birefringence

The red-green dichroism produced when Congo red dye binds to amyloid fibrils under polarized light.

Ataxia

A clinical symptom of prion disease pathogenesis involving lack of muscle coordination.

Startle myoclonus

A symptom associated with Creutzfeldt-Jacob disease and the rapid progression of dementia.

Multiple myeloma

A malignancy treated with proteasome inhibitors like Bortezomib.

G2-M phase

The stage of the cell cycle where proteasome inhibitors induce arrest in cancer cells.

Alpha 1 antitrypsin deficiency

A genetic disease associated with the failure of protein to be correctly trafficked due to misfolding.

Familial hypercholesterolemia

A condition characterized by the failure of a protein to be correctly trafficked.

Beta-mercaptoethanol

A thiol-containing compound used in laboratories to reduce disulfide bonds.

Glomerulus (Amyloid deposition)

A site in the kidney where amyloid deposits can be visualized using Congo Red stain and polarized microscopy.