GER lecture 3

• Genome stability

  Keeping all gene bases unchanged
• Mutations and genetic variation

  Essential for evolution but can also often lead to undesirable traits
• Genome stability

  • Genome needs to be stable to ensure survival
  • There also needs to be a little bit of instability for evolution
• DNA damage

  Can be caused by intrinsic and extrinsic factors
• Extrinsic DNA damaging agents


  • External DNA damaging agents
• Intrinsic DNA damaging agents


  • Damaging agents inside the cell and are not influenced by external factors
• DNA damage/lesion ≠ mutation !!!
• DNA damage turns into a mutation

  When it is not fixed before DNA replication
• Spontaneous hydrolytic damage

  Depurination and deamination
• Uracil

  Cytosine but without the amine group
• Deamination of cytosine

  Leads to uracil
• Deamination of 5-methylcytosine

  Leads to thymine
• Uracil

  Relatively easy to identify and repair – because it's not supposed to be in DNA at all
• Thymine

  Harder for DNA repair proteins to detect than uracil – because it is a DNA base
• Methylated DNA bases are common because they are used to regulate eukaryotic genes
• CpG sequences

  C on the 5' end and G on the 3' end
• Mutations often happen in CpG sequences
• Alkylating agents

  Reactive compounds that can transfer methyl or ethyl groups to DNA
• To minimise oxidative damage, the nucleus keeps oxygen away from the DNA
• Mitochondrial DNA is highly exposed to oxygen and ROS due to the electron transport chain, therefore the mutation rate is much higher in mitochondria than in nuclear DNA
• Benzopyrene

  Metabolised and modifies guanine
• Benzopyrene

  Blocks DNA replication and transcription (cytotoxic)
• There is evidence of links between benzopyrene and cancer
• UV light induced damage

  Chemically/covalently joins 2 DNA bases together (cyclobutane pyrimidine dimer)
• Cyclobutane pyrimidine dimer

  Blocks replication and transcription (cytotoxic and mutagenic)
• Cisplatin

  Covalently links 2 adjacent guanine bases – intrastrand crosslink
• Cisplatin

  Blocks replication and transcription (cytotoxic) → used to kill cancer cells
• Cisplatin

  Kills more cancer cells than normal cells because cancer cells undergo more replication
• Cisplatin also has a lot of side effects since it does kill normal cells as well
• Direct reversal repair pathway

  Most simple way of reversing damage is to run the damaging reaction backwards
• DNA photolyases

  Enzymes that repair UV-damaged DNA by using visible light energy to break the covalent bonds in cyclobutane pyrimidine dimer (CPDs)
• DNA photolyases are widely distributed in prokaryotes and eukaryotes – BUT they are not present in humans (or any other placental mammal)
• DNA alkyltransferases

  Enzymes that transfer the alkyl group from the damaged guanine to a cysteine residue within the alkyltransferase
• DNA alkyltransferases act in 'suicidal reactions' because they don't turnover like a normal enzyme, after they've been alkylated, they are degraded
• Spontaneous oxidative damage (e.g. 8-oxo-guanine)

  One of the most common DNA lesions resulting from reactive oxygen species (ROS).

  Base pairs ambiguously with cytosine or adenine (C → A mutation).

  Intrinsic DNA damage factor.

  To minimise this, the nucleus keeps oxygen away from the DNA.

  However, mitochondrial DNA is highly exposed to oxygen and ROS due to the electron transport chain.

  Therefore, the mutation rate is much higher in mitochondria than in nuclear DNA.
• Chemical induced damage – carcinogens.

  • E.g. Benzopyrene is metabolised and modifies guanine 
  • Block DNA replication and transcription (cytotoxic)
  • Evidence of links to cancer
  • Extrinsic DNA damage factor
• Cisplatin as a chemical treatment for cancer (i.e. chemotherapy)
• Direct reversal repair pathway

  Most simple way of reversing damage is to run the damaging reaction backwards.

  DNA returns to its original state – like it was before it was damaged.
• DNA photolyases repair UV-induced photoproducts.

  • DNA photolyases are flavoproteins (enzymes) that repair UV-damaged DNA by using visible light energy to break the covalent bonds in cyclobutane pyrimidine dimer (CPDs)
  • Flavoproteins are light-harvesting cofactors that transfer energy to FADH– 
  • FADH– can then act as a redox-active cofactor to break the cyclobutane ring
  • This process is known as photoreactivation 
  • DNA photolyases are widely distributed in prokaryotes and eukaryotes – BUT they are not present in humans
  • So, we are a lot more prone to skin cancer