Translation Part 2

• tRNA = Transfer RNA

  • A single stranded non-codingRNA that base pairs within itself, forming “t”-like structure
  • Job: Carry amino acids to the ribosome
• Two components of tRNA to know

  1. The sequence 5′-3′ at the 3’ end, this is where the amino acid gets attached
  2. The anticodon loop, a set of 3 RNA nucleotides which “reads” the codons in mRNA through complementary base pairing
• • A codon on the mRNA will attract, then bind to, a tRNA with complementary anticodon sequence via normal base pairing.
  • Like all nucleotide sequences, the anticodon of tRNA and codon of mRNA base pair in an antiparallel fashion.
• As we’ll learn, the ribosome moves along the mRNA 5’ to 3'
• This means the tRNA’s anticodon is oriented 3’ to 5’
• Aminoacyl tRNA synthetase enzymes match each tRNA with its appropriate amino acid
• Each aminoacyl synthetase enzyme has a specific amino acid it can bind to

  • This means there are at least 20 different types of aminoacyl synthetase, one type for each of the 20 amino acids
• Aminoacyl tRNA synthetase

  • Other parts of this protein enzyme “recognize” and bind to the unique shape of the tRNA with the anticodon corresponding to that enzyme’s specific amino acid
• When both amino acid and tRNA are bound, aminoacyl synthetase catalyzes formation of a bond between the amino acid and the tRNA

  • The tRNA is now “charged”
• The ribosome

  • Present in all living cells
  • Similar but not identical structure in prokaryotes and eukaryotes
  • Made up of 3-4 different (non-coding) RNA molecules; 52-79 protein molecules
  • It is the RNA components that catalyze peptide bond formation between amino acids
• How ribosomes interact with mRNA transcript (1)

  • The mRNA is bound by the large and small ribosomal subunits and moves through the center of the ribosome.
• How ribosomes interact with mRNA transcript (2)

  • The ribosome reads mRNA in 5 to′ 3 in multiples of three (in other words, codons)
  • The large subunit has 3 binding sites for tRNAs: E, P and A sites
• The A site accepts new, incoming aminoacyl (“charged”) tRNA
• The P site is where peptide bond formation between amino acids occurs.
• The E site is where the used, uncharged tRNA exits the ribosome.
• Initiation is where the initiation factorscome into play
• How initiation starts (eukaryotes)

  • The 5’ cap of mRNA serves as a binding site for an initiation complex that includes:

  1. Initation factors
  2. tRNA Met (has anticodon complementary to start codon)
  3. Small ribosomal subunit
• Initiation

  • Each gene has a different length of nucleotides before the actual coding region that acts as binding sites for various regulatory factors
• Initiation complex will start moving downstream (from 5’ cap to 3’ end) along mRNA to scan for START codon
• Initiation

  • Once complex reaches AUG, the large ribosomal subunit is recruited and binds to initiation complex
  • Ribosome positions tRNAMet at P site, establishing proper reading frame for subsequent codons
• What is a reading frame?

  • refers to how the string of nucleotides on mRNA is parsed into 3-letter “words” (codons)
• Elongation factors help to coordinate and catalyze reactions during the elongation step
• Elongation

  • The codon after the start codon in frame at the A site is now prepared to bind the complementary tRNA anticodon
• Elongation

  • When a charged tRNA with complementary anticodon bumps into complex, it binds through complementary base pairing
• Elongation

  • After the new tRNA binds, Met getstransferred to the amino acid on the tRNA in the A site and a peptide bond forms
• Elongation (when ribosome is moving)

  • This pushes tRNAMet-Val over to P site, and the first – now “uncharged” – tRNA over to the E site, where it gets ejected.
  • A new tRNA complementary to the codon in-frame in A site pairs with mRNA
• Elongation

  • The growing polypeptide chain transfers to the amino acid on the new tRNA in the A site
  • Another peptide bond is formed between polypeptide and new amino acid in A site Ribosome again moves down (5’ to 3’) one codon and repeats the process
• • peptide chain is built in the N- terminus to C-terminus direction
  • (amino group of new amino acid will always bind to carboxyl group of last amino acid in the polypeptide chain)
• Termination

  • Release factors
• Termination

  1. The process continues until ribosome encounters one of the stop codons (UAA, UAG, UGA)
  2. When the stop codon is encountered, a protein release factor binds to the A site of the ribosome, causing the bond connecting polypeptide to tRNA to break
  3. The breaking of the bond creates the carboxyl terminus of the polypeptide and completes the polypeptide primary structure