pastpaperbd/ Biology/ Notes/ DNA replication and Protein synthesis
T4

3.4.2 - DNA and Protein Synthesis

This section has two parts: transcription (DNA → mRNA) and translation (mRNA → protein).

Semi-Conservative Replication

Before we get to protein synthesis, it's worth covering replication here as it naturally follows from DNA structure.

DNA replication is semi-conservative - each new DNA molecule consists of one original (template) strand and one newly synthesised strand. This was proven by the Meselson-Statt experiment using heavy (¹⁵N) and light (¹⁴N) nitrogen.

The process requires several key enzymes - Q9.1 in your paper asked you to name two:

Helicase - breaks the hydrogen bonds between base pairs, unwinding the double helix and separating the two strands at the replication fork.

DNA polymerase - adds free DNA nucleotides to the 3' end of the growing strand, using the template strand in the 3'→5' direction. It can only work in the 5'→3' direction, which is why one strand (the lagging strand) is synthesised discontinuously in Okazaki fragments.

DNA ligase - joins Okazaki fragments on the lagging strand together by forming phosphodiester bonds.

Free DNA nucleotides must be activated (as nucleoside triphosphates) before they can be incorporated. The hydrolysis of the extra phosphate groups releases energy to drive bond formation.

Transcription

Transcription is the synthesis of mRNA from a DNA template. It occurs in the nucleus.

  1. RNA polymerase binds to a promoter region upstream of the gene
  2. It unwinds and separates the DNA strands
  3. It reads the template strand (also called the antisense strand) in the 3'→5' direction
  4. Free RNA nucleotides align by complementary base pairing (A pairs with U in RNA, not T)
  5. RNA polymerase catalyses the formation of phosphodiester bonds between nucleotides, building the mRNA in the 5'→3' direction
  6. The mRNA produced is complementary to the template strand, and therefore has the same sequence as the coding strand (sense strand), except with U instead of T

In eukaryotes, the pre-mRNA initially produced contains both exons (coding sequences) and introns (non-coding sequences). The introns are removed by a process called splicing, and the exons are joined together to produce mature mRNA that leaves the nucleus through nuclear pores.

This is important - it means eukaryotic genes are split genes (mosaic structure), whereas prokaryotic genes are continuous with no introns.

Translation

Translation occurs at ribosomes in the cytoplasm (or on rough ER for secretory proteins).

The mRNA attaches to a ribosome. Transfer RNA (tRNA) molecules bring amino acids to the ribosome. Each tRNA has:

  • An anticodon - a triplet of bases that is complementary to a specific mRNA codon
  • An amino acid attachment site at the 3' end, where a specific amino acid is loaded (by an enzyme called aminoacyl-tRNA synthetase)

The ribosome has three sites - A (aminoacyl), P (peptidyl) and E (exit):

  1. The ribosome reads the mRNA codon in the A site
  2. The complementary tRNA anticodon binds by hydrogen bonds
  3. A peptide bond forms between the amino acids (condensation reaction, catalysed by peptidyl transferase - actually the rRNA itself, making ribosomes ribozymes)
  4. The ribosome moves along the mRNA (translocation), shifting the growing polypeptide to the P site and the empty tRNA to the E site
  5. This continues until a stop codon is reached - no tRNA has a complementary anticodon, so a release factor causes the polypeptide to be released

The polypeptide then folds into its secondary, tertiary and quaternary structure, often assisted by chaperone proteins. Errors during replication can lead to Mutations; the code being read is defined in Genes.

Summary

  • Semi-conservative replication: each new molecule = 1 original strand + 1 new strand. Proven by Meselson-Stahl (¹⁵N/¹⁴N experiment).
  • Helicase: breaks H-bonds between base pairs, unwinds helix at replication fork
  • DNA polymerase: adds nucleotides to 3' end in 5'→3' direction; requires template in 3'→5' direction; needs a primer. Lagging strand is synthesised discontinuously as Okazaki fragments.
  • DNA ligase: joins Okazaki fragments by forming phosphodiester bonds
  • Transcription (nucleus):
  • RNA polymerase binds promoter; unwinds DNA
  • Reads template strand 3'→5'; builds mRNA 5'→3'
  • A pairs with U; G pairs with C
  • Pre-mRNA→splicing (introns removed, exons joined)→mature mRNA exits via nuclear pores
  • Translation (ribosomes):
  • mRNA attaches to ribosome; start codon AUG
  • tRNA brings amino acids; anticodon pairs with mRNA codon
  • Ribosome has A, P, E sites; peptide bond forms between amino acids (catalysed by rRNA - ribozyme)
  • Ribosome translocates along mRNA; continues until stop codon → polypeptide released
  • Eukaryotic genes have introns; prokaryotic genes do not - important difference

AQA Exam Tips

  • Semi-conservative evidence: in the Meselson-Stahl experiment, after one round of replication in ¹⁴N, all DNA was at intermediate density (one ¹⁵N, one ¹⁴N strand). After two rounds, half was intermediate and half was light (¹⁴N/¹⁴N). This proves semi-conservative.
  • DNA polymerase direction: can only add nucleotides to the 3' end, so it reads the template 3'→5' and synthesises 5'→3'. The lagging strand is therefore synthesised discontinuously in short Okazaki fragments.
  • Transcription vs replication: both involve unwinding DNA and adding complementary nucleotides, but transcription uses RNA polymerase (not DNA polymerase), produces mRNA (not DNA), uses RNA nucleotides (with U not T), and copies only one strand of one gene (not the whole genome).
  • Why introns are removed: mature mRNA must only contain coding sequences (exons) for correct translation. Introns are non-coding - if they remained, the reading frame would be disrupted. Alternative splicing allows one gene to produce multiple proteins.
  • tRNA structure: anticodon (3 bases complementary to mRNA codon) at one end; amino acid attachment site at the other (3'-OH of adenosine). The anticodon is antiparallel to the codon.
  • Peptide bond formation: condensation reaction between the amino group of one amino acid and the carboxyl group of the next. Catalysed by rRNA in the large subunit (peptidyl transferase activity).