Translation Overview

• Ribosomes translate mRNA sequence into proteins
• mRNA is read 5' to 3'
• Polypeptide is synthesized from amino end toward carboxyl end
• Rate about 15-20 peptides per sec

Amino acids and polypeptides

• The general form of an amino acid is H2N -C(R)- COOH
• The general structure is the same for all, but they differ in their R group.
• Polypeptides are chains of amino acids linked with peptide bonds One end is the amino or N-teminus and the other is the carboxyl or C-terminus
• See Figs. 6.1 and 6.3 for the genereal form of amino acids and polypeptides

The 20 amino acids can be divided into four groups:

• Acidic:
• Basic:
• Neutral Hydrophilic
• Neutral Hydrophobic

I recommend that you learn which amino acid is in which group, and learn to recognize the amino acids by their one-letter abbreviations, though that is not required for this course.

The genetic code

• The code is a triplet code, divided into 3-letter words that each specify an amino acid.
• The first evidence for triplet code came from deletion experiments- a one base deletion leads to nonsense, because there is a shift in the reading frame of the triplets.
• • the new boy saw the big cat eat the hot dog
  • the neb oys awt heb igc ate att heh otd og
• Function could be restored by an insertion nearby
• • the neb oys aaw the big cat eat the hot dog
• or by two more deletions to get it back into the correct "reading frame"
• • the neb osa wte big cat eat the hot dog
• As long as there were 3 insertions or 3 deletions, most of the code would be correct.

Deciphering the Code

• Your book tells how people used synthetic mRNAs and in vitro translation to determine decipher the codons.
• E.g. UUUUUUUU -> phenylalanine only
• UCUCUCUCUCU -> mix of leucine and serine
• Why is it a mixture?

General features of the genetic code:

• The code is redundant
• • Several different codons encode the same amino acid
• The code is comma free
• • It does have start and stop signals, however.
  • Start: AUG
  • Stop: UAG, UAA, UGA
• The code has wobble
• • The third base is less important than the first two
  • The third base of the anticodon can often pair with several bases
  • • G with C or U
    • U with A or G
    • I with C or A or U

Molecular basis of Translation

Charging the tRNA

• Aminoacyl tRNA synthetase adds the amino acid to a tRNA
• One synthetase for each tRNA
• See Fig. 6.10

Initiation

• Initiation factors and ribosome small subunit bind to mRNA
• Prokaryotes: AUG and Shine-Delgarno seq.
• Eukaryotes: initiators bind to 5' cap then scans for start codon
• Special methionine binds to start codon
• Large subunit binds

See and study Fig. 6.12

Elongation

• Ribosome has three sites (A, P, E)
• Three-step cycle
• 1. recruitment of aminoacyl-tRNAs to A site
• 2. formation of peptide bonds
• 3. Translocation to P site
• Exiting tRNAs can be charged again
• Polypeptide chain remains attached to the most recent tRNA

See and study Fig. 6.15 There are a lot of details in the book about the machinery involved in elongation, but the basic concept is pretty simple.

Elongation is similar in prokaryotes and eukaryotes, but the elongation factors have different names.

Termination

• Stop codons don't have a matching tRNA, so the ribosome pauses.
• Special release factors bind to A site and release the polypeptide.

Take another look at the translocation animation at www.dnai.org

Cost of protein biosynthesis

• Synthesis of aminoacyl tRNAs: 2 ATPs
• Proofreading (deacylation): 1 ATP/error
• Formation of 1 peptide bond: 4 GTPs

Protein Sorting

Many polypeptides have signal sequences at N-end which are necessary to direct the new protein to the correct part of the cell.

Chapter 6 problems

6, 8, 12, 16, 18, 19, 22, 24, 25