Biology Lecture 8: mRNA Translation and Protein Synthesis

Translation of mRNA-Protein Synthesis

• Translation occurs in three distinct phases: initiation, elongation, and termination
• Key components required for translation initiation:
  • Three types of RNA: mRNA, rRNA, and tRNA
  • GTP (guanosine triphosphate)
  • A set of soluble protein factors (initiation factors)

mRNA Structure

• mRNA consists of:
  • Untranslated regions (UTR)
  • 5'-UTR (upstream of start codon)
  • 3'-UTR (downstream of stop codon)
  • Protein coding sequence (start codon to stop codon)
  • Polyadenylation signal in 3'-UTR

Genetic Code

• 64 possible codons
• 61 codons encode specific amino acids
• 1 codon encodes start codon (methionine, AUG)
• 3 codons are stop codons (UAA, UAG, UGA)
• Similar genetic code in mitochondria with few variants

Ribosomes

• Ribosomes are the sites of protein synthesis
• Composed of rRNA and proteins
• Large and small subunits
• Sedimentation coefficient or S value determines ribosome size
• E. coli has 20,000 ribosomes, while mammalian cells have 10 million ribosomes

rRNA Synthesis and Processing

• In eukaryotes, rRNA synthesis occurs in multiple genes organized in clusters
• Transcribed by RNA polymerase I
• Post-transcriptional cleavages occur

tRNA Structure

• tRNAs are short, single-stranded RNA molecules (80 bases long)
• Contain stem loop structures, modified bases, and unusual bases
• 3'-end is the amino acid attachment site
• Anticodon site base pairs with the codon at mRNA

tRNA-Anticodon and Codon Pairing

• Correct match between tRNA anticodon and mRNA codon is necessary
• Amino acid attached to tRNA is specified by the anticodon and identifier sequences
• Anticodon is 3 nucleotides in tRNA that pair with a mRNA codon
• Codon is 3 nucleotides in mRNA that code for a single amino acid

Wobble Base Pairing

• Wobble base pairing allows some tRNAs to recognize more than one codon
• Redundancy/degeneracy of the genetic codons acts as a buffer against deleterious mutations

Amino Acid Attachment to tRNA

• Amino acid attachment to tRNA occurs through aminoacyl-tRNA synthetase
• The process requires energy in the form of ATP
• The amino acid is covalently attached to the ribose of the terminal adenosine at CCA

Translation Initiation

• Formation of initiation complex involves:
  • Ribosome binding site
  • P site (holds the tRNA that carries the growing polypeptide chain)
  • A site (holds the tRNA that carries the next amino acid to be added to the chain)
  • E site (tRNAs leave the ribosome)
• Initiation site is specific: AUG (methionine) in both prokaryotes and eukaryotes
• Large and small ribosomal subunits
• In prokaryotes, initiation sites have a Shine-Dalgarno sequence that precedes the AUG initiation codon
• In eukaryotes, mRNAs are bound to the 40S ribosomal subunit by their 5' caps, and the ribosome then scans along the mRNA until it encounters an AUG initiation codon

Translation Elongation

• Three steps:
  1. Next aminoacyl-tRNA binding
  2. Peptide bond formation
  3. Translocation
• eEF1α (eukaryotic elongation factor) brings the next aminoacyl-tRNA to the A site
• Peptide bond is formed by the transfer of methionine to the aminoacyl tRNA at the A site
• The ribosome then moves three nucleotides along the mRNA, translocating the peptidyl tRNA to the P site and the uncharged tRNA to the E site

Translation Termination

• A termination codon (e.g., UAA) at the A site is recognized by release factors
• The result is the release of the completed polypeptide chain, followed by the dissociation of tRNA and mRNA from the ribosome

Polyribosome

• Polyribosome is a cluster of ribosomes bound to a single mRNA molecule
• Allows for simultaneous translation of the same mRNA to make many copies of the polypeptide quickly

Post-Translational Modifications

• 1. Chaperon: protein responsible for proper folding of protein
• 2. Addition of lipid
• 3. Addition of carbohydrate
• 4. Proteolytic processing

Regulation of Translation

• 1. Global Regulation: translation can be controlled globally by regulating the activity of initiation factors
• 2. Binding of repressor proteins: regulated by proteins that bind to specific sequences in the 3' untranslated regions of some mRNAs
• 3. Binding of non-coding RNA (miRNA): miRNAs bind to 3' UTR sequences to inhibit translation and stimulate mRNA degradation