Protein Synthesis, Folding, and Degradation - Quick Notes

Ribosome sites: P site (peptidyl), A site (aminoacyl); tRNA entry at A-site; peptide bond forms between amino acids; ribosome advances along mRNA (translocation).
mRNA and tRNA interplay: codon-anticodon recognition drives incorporation of amino acids (e.g., Met initiator).
Key sequence: Met on initiator tRNA; subsequent amino acids added as ribosome slides.
Core process: Entry of aminoacyl-tRNA to A-site → peptide bond formation → translocation to P-site → A-site accepts next tRNA.

Non-ribosomal protein factors: Initiation factors (IF), Elongation factors (EF), Release factors (RFs).
In eukaryotes, IFs are regulated by cellular signals; global control of protein synthesis and cellular stress responses.

Most proteins fold spontaneously to their stable forms; some need help.
Chaperones assist folding during/after translation.
Major chaperone systems:
- Hsp70 system: small, works during translation; binds exposed hydrophobic regions; uses ATP to drive correct folding.
- Chaperonin system (GroEL/GroES, Hsp60-like): large barrel-like complex; acts post-translation to aid refolding.
Misfolding is detected by exposed hydrophobic residues on unfolded/misfolded proteins.

Hsp70 binds exposed hydrophobic regions as proteins emerge from the ribosome; ATP hydrolysis drives conformational changes and release for proper folding.
Hsp70 also participates in mitochondrial protein import via cooperation with mitochondrial Hsp70.

Similarities: both recognize exposed hydrophobic regions, assist refolding via ATP hydrolysis.
Differences:
- Hsp70: smaller complex, active during translation (co-translationally).
- Chaperonin: larger barrel, acts mainly after translation.
- Both help avoid aggregation and assist achieving native structure.

Prion protein (PrP): normal cellular form is non-toxic; rare conformational change can produce toxic PrP that induces others to misfold.
PrP can act as infectious agent without nucleic acids; misfolded PrP aggregates and propagates.
Misfolding can lead to a spectrum of diseases via protein aggregation.

Neurodegenerative diseases: Alzheimer's (β-amyloid), Parkinson's (α-synuclein), Huntington's (huntingtin), ALS (SOD1), prion diseases (PrP).
Localized/systemic amyloidoses: various proteins (e.g., amylin, transthyretin).

Some mutant proteins may be functional but misfolded recognition (degraded); e.g., CFTR is often functional biochemically but not expressed on membrane due to misfolding recognition by chaperones.

Regulation occurs via small molecules, covalent modifications (e.g., phosphorylation), and protein–protein interactions.

Three important proteins: E1 (ubiquitin-activating), E2 (ubiquitin-conjugating), E3 (ubiquitin ligase).
Step 1: Activation
- E1 binds free ubiquitin and activates it using ATP:
- ext{E1} + ext{Ub} + ext{ATP}
  ightarrow ext{E1–Ub} + ext{AMP} + ext{PP}_i
Step 2: Transfer to E2
- E1 transfers Ub to E2; E1 is released. Complex becomes E2–Ub (often with E3 present as ubiquitin ligase).
Step 3: Substrate tagging
- Degradation signal (e.g., exposed hydrophobic region) is recognized by E3.
- Ubiquitin is transferred from E2 to a lysine on the substrate.
- Repeated cycles build a polyubiquitin chain on the substrate: $ext{Ub}_n ext{ chain}$ .
Regulation
- There are hundreds of E3 ubiquitin ligases with different substrate specificities; not all tagging targets misfolded proteins—many regulate cellular processes.

Multi-ubiquitinated substrates are recognized by the proteasome, a large ATP-dependent protease complex.
Proteasome structure: a barrel-like protease core with proteolytic sites aligned for degradation.
Degradation is selective and regulated, ensuring proper turnover of many cellular proteins.

Protein degradation can regulate cell cycle proteins (cyclins, securin, ORC components) via specific E3s.
The proteasome-mediated pathway accounts for a substantial fraction of newly synthesized protein turnover in cells.