Protein Synthesis and Post-Translational Modifications

Genetic Code & Ribosomes - Code Crackers Crew

• Genetic Code: Triplet, non-overlapping, commaless, degenerate, nearly universal.
  • 64 codons: 61 sense (code for amino acids), 3 nonsense (stop).
  • Start codon: AUG (Methionine; rarely GUG).
  • Stop codons: UAA, UAG, UGA. 📌 Mnemonic: "U Are Away, U Are Gone, U Go Away".
• Ribosomes: rRNA + protein; site of translation.
  • Prokaryotic: 70S (50S + 30S subunits).
  • Eukaryotic: 80S (60S + 40S subunits).
  • A, P, E sites for tRNA binding.
• mRNA: Carries genetic information from DNA as codons.
• tRNA: Adaptor molecule; anticodon pairs with mRNA codon, carries specific amino acid.

⭐ The peptidyl transferase activity, forming peptide bonds, is a ribozyme function of the large ribosomal subunit's rRNA (23S in prokaryotes, 28S in eukaryotes).

Translation Steps - Protein Assembly Line

• Initiation:
  • • Small subunit (30S/40S) + mRNA + initiator tRNA (fMet/Met) + IFs → complex.
  • • Prok: Shine-Dalgarno. Euk: 5' cap, Kozak seq.
  • • Large subunit (50S/60S) joins; GTP used.
• Elongation: (📌 APE sites: Aminoacyl, Peptidyl, Exit)
  • • Codon Recognition: aa-tRNA to A-site (EF-Tu/eEF1A, GTP).
  • • Peptide Bond: Peptidyl transferase (rRNA ribozyme) links AAs. Peptide to A-tRNA.
  • • Translocation: Ribosome moves 1 codon (EF-G/eEF2, GTP). P→E, A→P.
• Termination:
  • • Stop codon (UAA, UAG, UGA) in A-site.
  • • Release Factors (RFs/eRFs) bind.
  • • Polypeptide released; complex dissociates (GTP).

⭐ Peptidyl transferase is a ribozyme (catalytic rRNA: 23S in prokaryotes, 28S in eukaryotes).

Translation Control & Inhibitors - Quality & Sabotage

• Control:
  • Initiation: Key regulatory step. eIF2-P (stress, ↓ global), eIF4E (cap-binding, ↑ cancer). IRES (cap-independent translation).
  • mRNA stability (e.g., AU-rich elements affecting decay).
• Prokaryotic Inhibitors (70S): 📌 "Buy AT 30, CELL at 50"
  • 30S: Aminoglycosides (misreading mRNA), Tetracyclines (block A-site tRNA entry).
  • 50S: Macrolides (translocation), Chloramphenicol (peptidyltransferase), Clindamycin (peptide bond), Linezolid (prevents 70S initiation complex).
• Eukaryotic Inhibitors (80S):
  • Diphtheria toxin: ADP-ribosylates eEF-2, halting elongation.
  • Ricin (plant toxin): Cleaves rRNA in 60S subunit.
• Quality/Sabotage:
  • NMD: Degrades mRNA with PTCs (Premature Termination Codons), preventing truncated proteins.
  • Viral shutoff: Poliovirus protease cleaves eIF4G.

⭐ Diphtheria toxin inactivates eukaryotic elongation factor 2 (eEF-2) via ADP-ribosylation, stopping protein synthesis.

Post-Translational Modifications - Protein Finishing Touches

• Post-synthesis changes; vital for protein folding, stability, localization, activity.
• Common Modifications:
  • Phosphorylation: Ser/Thr/Tyr. Kinases add, phosphatases remove. Regulates signaling, enzyme activity.
  • Glycosylation: N-linked (Asn, ER), O-linked (Ser/Thr, Golgi). Folding, targeting, recognition.

    ⭐ Mannose-6-phosphate (M6P) targets enzymes to lysosomes; M6P pathway defect causes I-cell disease.

  • Ubiquitination: Lys. Tags proteins for proteasomal degradation.
  • Hydroxylation: Pro, Lys in collagen. Needs Vit C (📌 Collagen C needs Vit C). Deficiency: Scurvy.
  • Carboxylation: Glu in clotting factors. Needs Vit K (📌 K for Koagulation).
  • Acetylation/Methylation: Lys, Arg in histones. Regulates gene expression.
  • Proteolytic Cleavage: Zymogen activation (trypsinogen), signal peptide removal.

High‑Yield Points - ⚡ Biggest Takeaways

• Wobble hypothesis: Inosine in tRNA anticodon allows binding multiple codons, explaining code degeneracy.
• Antibiotics: Streptomycin (30S) causes mRNA misreading; Chloramphenicol (50S) inhibits peptidyl transferase.
• Puromycin: Aminoacyl-tRNA analog, causes premature termination in prokaryotes and eukaryotes.
• Protein targeting: Signal Recognition Particle (SRP) directs proteins to ER for N-linked glycosylation; O-linked glycosylation occurs in Golgi.
• Modifications: Ubiquitination targets proteins for proteasomal degradation; SUMOylation alters function/localization.
• Protein folding: Assisted by chaperones (e.g., Hsp70); misfolding is linked to diseases like Alzheimer's.
• Kozak sequence (eukaryotes) or Shine-Dalgarno sequence (prokaryotes) initiates translation at the start codon.