Notes on DNA Supercoiling and Topoisomerases

DNA Structure in Bacterial Cells

• E. coli DNA Length: DNA strand released from a lysed E. coli cell is 1,500 times longer than the cell itself.
• Compaction: Inside the cell, DNA is compacted into a structure known as a nucleoid.
  • Nucleoid Structure: DNA is arranged in tightly wound loops.
  • Anchoring Proteins: Loop boundaries defined by histone-like anchoring proteins.
  • Supercoiling: DNA in loops is supercoiled (helix coils upon itself for compactness).

Supercoiling

• Relaxation of DNA: If one strand of DNA is cut, it loses supercoils due to tension dissipation.
• Constrained Loops: Other domains remain supercoiled, anchored by proteins preventing rotation.
• Concept of Supercoiling:
  • A relaxed circular DNA naturally forms a helix with ~10 base pairs per turn.
  • Cutting a strand and unwinding creates an underwound state that forms negative supercoils to relieve stress.
  • Positive supercoils relieve stress from overwound DNA.

Topoisomerases

• Function: Enzymes that modify DNA supercoiling by changing DNA topology.
• Types of Topoisomerases:
  • Type 1: Cleave one strand of a double helix, generally used to relieve or unwind supercoils.
  • Type 2: Cleave both strands, use energy to introduce supercoils.

Type 1 Topoisomerases

• Mechanism:
  • Binds to DNA, opens strands, loosens double helix.
  • Loss of a turn in the helix converts supercoils (e.g., from five to four).
  • Cleaves one strand, passes intact strand through the break, and reseals.
  • Example: Reduces number of supercoils in a DNA molecule.

Type 2 Topoisomerases

• DNA Gyrase: Adds negative supercoils to DNA.
  • Structure: Tetramer with two GyrA and two GyrB proteins.
  • Mechanism:
    • GyrB grabs DNA section, GyrA introduces double strand break.
    • ATP hydrolysis by GyrA provides energy to pass intact DNA through break.
    • Re-seals DNA, introducing one new negative supercoil.

Balancing Supercoiling

• Importance: Proper DNA supercoiling balance is crucial for cell function.
  • Negative Supercoiling in Bacteria:
    • Nucleoids and nuclear DNA of eukaryotes are kept negatively supercoiled.
    • Easier strand separation aids transcription by RNA polymerase.
  • Positive Supercoiling in Archaea:
    • Species in acidic, high-temperature environments maintain DNA in positive supercoils.
    • Positively supercoiled DNA resists denaturing due to extra tightness.

Conclusion

• The supercoiling state of DNA is managed by topoisomerases, ensuring proper cellular function in various environments and conditions.