Protein Synthesis Notes

  1. Background - How does DNA "control" a cell? Where are the instructions to make a protein? How does this happen?
    1. proteins carry information in their amino acid sequence
    2. nucleic acids carry information in their nucleotide sequence
    3. transcription - copying the instructions from the master DNA Why is this necessary?
    4. translation - the instructions must then be converted from one language to the other
    5. the code for translation is a series of triplets (codons) which indicate a particular amino acid (Table 27.1N; Fig. 16.5C)
    6. Problem: only 4 bases but 20 amino acids
      1. Crick, 1961 - 41 = 4; 42 = 16; 43 = 64
      2. therefore, the code is a series of triplets which indicate a particular amino acid. Triplets are called codons.
      3. note: redundancy but no ambiguity. e.g., GAA and GAG both mean glutamic acid, but never any other amino acid
      4. remember, DNA not permitted to leave nucleus so need a messenger to take the information to cytosol where ribosomes are.
      5. messenger is messenger RNA (mRNA)
    7. note that the code contains redundancy but no ambiguity
  2. Transcription (Fig 27.4N; Fig16.7C) - the entire process is much like DNA replication except that DNA is acting as a template for the construction of mRNA
    1. initiation
      1. RNA polymerase separates the two strands and looks for the initiation sequence (TATA box)
      2. transcription begins at this site
    2. elongation
      1. RNA polymerase moves along the template (coding) strand, adding bases by base pairing rules to form mRNA
      2. works 5' to 3' at a rate of about 60 nucleotides/s
      3. if there is high demand for a protein, the cell can have several RNA polymerases transcribing the same gene simultaneously
      4. remember that in RNA, U rather than T is paired with A
    3. termination
      1. transcription continues until the termination sequence (AATAAA) is reached
      2. the mRNA is then sent to the cytosol
      3. a transcription unit is the sequence between the start and stop sequences - generally speaking, one gene
  3. Translation (Fig 27.5N; Fig 16.9C, 16.13C, 16.14C, 16.15C)
    1. Background
      1. mRNA is translated into amino acid language by tRNA while the protein is built by a ribosome
      2. each tRNA is specific for one amino acid
      3. amino acid is attached to the tRNA at the amino acid attachment site
      4. anticodon, a group of three nucleotides at the other end of the tRNA, binds by complementary base pairing to the mRNA
    2. Initiation
      1. ribosome has two tRNA binding sites called A and P
      2. ribosome binds to mRNA and begins looking for start codon (AUG)
      3. when it is found, it is displayed in the P site so tRNA molecules can attempt to recognize it by complementary base pairing with their anticodon. When this occurs, the two ribosomal subunits come together to form the functional ribosome. The tRNA with the anticodon complementary to AUG always carries methionine (met) so it is always the first amino acid in every protein tRNA carrying methionine enters the P site
      4. the ribosome now displays the next codon in the A site and waits for the tRNA with the complementary anticodon to recognize it
      5. Why can't the ribosome begin translating anywhere? AUG establishes reading frame
      6. if the cellular demand for a protein is high, several ribosomes can translate the mRNA simultaneously
    3. Elongation (~ 60 ms per peptide bond)
      1. tRNA with anticodon complementary to next codon enters the A site
      2. ribosome forms a new peptide bond by transferring the amino acid from tRNA in the P site to the amino acid on the tRNA in the A site
      3. tRNA (now empty) in the P site leaves and the ribosome moves over one codon on the mRNA
      4. the next codon is now displayed in the A site
      5. the appropriate tRNA moves in and ribosome attaches the amino acids (2) from the tRNA in the P site to the amino acid on the tRNA in the A site
      6. the polypeptide grows by the addition of one amino acid at a time in this manner
      7. if the cell has a high demand for the protein several ribosomes may translate the same mRNA simultaneously
    4. Termination
      1. this process continues until a stop codon is reached
      2. there is no tRNA which recognizes any of the stop codons
      3. the ribosome pauses briefly and then severs the bond between the tRNA in the site and the polypeptide
    5. Evolutionary link - CCG means proline in every organism studied so far
  4. Mutations
    1. substitutions
      1. 1 nucleotide is replaced by another
      2. Are these mutations always harmful?
      3. could change to a different amino acid or to a stop codon
    2. insertions or deletions
      1. addition or subtraction of one or more nucleotides
      2. usually serious because of frame shift unless three nucleotides are added or subtracted one or more nucleotides are added or deleted. Are these mutations always harmful?
    3. mutagens
      1. anything that increases the natural frequency of mutations
      2. X-rays, UV, other radiation can cause abnormal base pairing or irreversible bonding. Radiation can also break the ladder and splicing enzymes sometimes put the pieces back together in the wrong order
      3. often chemicals which have a shape similar to that of one of the nucleotides but makes different hydrogen bonds so that base pairing is not possible
      4. some chemicals change the shape of nucleotides so that base pairing is impossible or faulty
      5. exposure to mutagens is especially dangerous for pregnant women because the embryo is growing rapidly and few embryonic cells give rise to all adult cells
      6. note difference between mutation in somatic versus germ cell. Germ cells pass on mutations to all subsequent daughter cells.
  5. Gene therapy - useful to know where genes are so we can manipulate them
    1. gene insertion
      1. a normal copy of a gene is inserted into the correct position in DNA if a cell is missing it
      2. usually done by having a virus (called a vector) carry the gene into the cell
      3. e.g., nasal spray for cystic fibrosis
      4. e.g., insert gene for insulin into pancreatic cells and implant in patient. As new cells divide, pancreas wold slowly become non-diabetic
      5. one problem is that a gene are often controlled by its location on the chromosome so we could over-produce a protein
      6. one problem is that genes are often controlled by their location on the chromosome so moving them may result in genes being out of control
    2. gene transplant - defective gene can be removed and replaced by a functional one