mRNA Translation and tRNA

• In the nucleus, an mRNA copy of a gene is created via transcription.
• The mRNA then exits the nucleus through a nuclear pore.
• In the cytoplasm, a ribosome binds to the mRNA to initiate translation.
• During translation, amino acids are linked together at the ribosome, forming a polypeptide chain based on the mRNA sequence.
• The language of nucleotides is translated into the language of amino acids during translation.
• The nucleotide sequence of an mRNA dictates the amino acid sequence of a protein through the genetic code.
• Each set of three nucleotides (a codon) corresponds to a specific amino acid.

Transfer RNA (tRNA)

• The genetic code is read by tRNA molecules.
• tRNAs are about 80 nucleotides long.
• It is composed of a single-stranded RNA molecule that folds back on itself to form a cloverleaf-like structure.
• tRNAs contain modified versions of uracil such as pseudouridine (Ψ) and dihydrouridine (D).
• It attaches to an amino acid at one end and binds to a codon at the other end.
• The anticodon is the triplet in the tRNA that binds to a codon in mRNA.
• Aminoacyt-tRNA synthetase attaches an amino acid to a tRNA, there is a specific one for each amino acid.

Importance of Wobble Base Pairing

• Flexibility exists in pairing the third base of the codon with the first base of the anticodon.
• At the wobble position, hydrogen bonds can form between G:U, I:U, I:A, and I:C.
• As a result, roughly 30 tRNA types are necessary to accommodate the 61 sense codons.

Ribosome Structure

• Ribosomes serve as workbenches for translation.
• Ribosomes are composed of a large and a small subunit.
• Each subunit is composed of multiple proteins and several RNA molecules.
• Eukaryotic ribosomes (80S) are larger than prokaryotic ribosomes.
• Eukaryotic ribosomal RNAs include 28S, 5S, 5.8S, and 18S.
• Prokaryotic ribosomal RNAs are 23S, 5S, and 16S.
• Each ribosome has one mRNA binding site and three tRNA binding sites; the A (aminoacyl-tRNA) site, the P (peptidyl-tRNA) site, and the E (exit) site.
• The A site is where the charged tRNA anticodon binds to the mRNA codon.
• At the P site the tRNA carrying the growing peptide chain resides here.
• The E site is where the uncharged tRNA resides before exiting the ribosome.

Translation Initiation

• Translation initiation in prokaryotes requires sequence recognition, with 16S rRNA recognizing the Shine-Dalgarno sequence.
• The Shine-Dalgarno sequence is upstream of the start codon (AUG) in the small subunit of the ribosome.
• Translation initiation in eukaryotes involves recognizing the 5' cap of the mRNA.

Translation Elongation and Peptide Bond Formation

• The 23S rRNA is responsible for peptide bond formation in prokaryotes.
• The 28S rRNA is responsible for peptide bond formation in eukaryotes.

Translation Termination

• Stop codons (UAA, UAG, and UGA) do not correspond to any amino acids, but bind release factors instead of tRNA.
• The release factor binding allows hydrolysis of the bond between the polypeptide chain and the tRNA at the P site.

Polysomes

• A polysome (or polyribosome) is an mRNA assemblage, ribosomes, and their accumulating polypeptides.
• Polysome formation increases the rate of protein synthesis.

Post-translational Modifications

• Required for a new protein to become fully functional proteins.
• Examples include:
  • Folding
  • Cofactor binding
  • Noncovalent interactions
  • Covalent modifications such as phosphorylation.
  • Noncovalent binding to other protein subunits.
  • Adding phosphate groups after the shape of the protein.
  • Adding sugars after the targeting and recognition
  • Cleaving the polypeptide that allows the fragments to fold into different shapes.

Protein Destination

• Proteins have diverse destinations in the cell.
• Such as the Cytosol, Lysosome, Endoplasmic Reticulum, Golgi Apparatus, Protein, and Mitochondrion.
• Protein destination is encoded in its sequence.
• Signal sequences guide a protein to its final location.
• Altering signal sequences changes protein destination in the cell.
• The following are some typical signal sequences in the cell:
  • Import into ER
  • Retention in lumen of ER
  • Import into mitochondria
  • Import into nucleus
  • Export from nucleus
  • Import into peroxisomes
• The information required to guide a protein to its final destination is encoded in its sequence.
• Adding or removing signal sequences changes protein destination in the cell.

Proteins Entering Organelles

• An import receptor protein recognizes the mitochondrial protein.
• The import receptor is associated with a protein translocator.
• A second translocator recognizes the signal sequence found in the inner membrane.
• The two translocators transport the protein into the matrix.
• The nuclear localization signal of prospective nuclear proteins are recognized by nuclear import receptors (a transport receptor).

Endoplasmic Reticulum

• An ER signal sequence guides a ribosome to the ER membrane.
• Proteins with an ER signal sequence enter via a protein translocator.
• An ER signal recognition particle (SRP) and its receptor (SRP receptor) direct a ribosome to the ER membrane.
• Proteins lacking a stop-transfer sequence enter the ER lumen.
• Vesicles transport these proteins to their final destination.
• Stop-transfer sequences allow transmembrane proteins to be incorporated in the plasma membrane
• Transmembrane proteins can be single-pass or multiple-pass.