DNA Replication

DNA Replication

1. DNA replication is a semi-conservative process that is carried out by a complex system of enzymes

2. Helicase

   • Helicase unwinds and separates the double-stranded DNA by breaking the hydrogen bonds between base pairs
   • This occurs at specific regions (origins of replication), creating a replication fork of two strands running in antiparallel directions

3. DNA Gyrase

   • DNA gyrase reduces the torsional strain created by the unwinding of DNA by helicase
   • It does this by relaxing positive supercoils (via negative supercoiling) that would otherwise form during the unwinding of DNA

4. Single Stranded Binding (SSB) Proteins

   • SSB proteins bind to the DNA strands after they have been separated and prevent the strands from re-annealing
   • These proteins also help to prevent the single stranded DNA from being digested by nucleases
   • SSB proteins will be dislodged from the strand when a new complementary strand is synthesised by DNA polymerase II

5. DNA Primase

   • DNA primase generates a short RNA primer (~10–15 nucleotides) on each of the template strands
   • The RNA primer provides an initiation point for DNA polymerase III, which can extend a nucleotide chain but not start one

6. DNA Polymerase III

   • Free nucleotides align opposite their complementary base partners (A = T ; G = C)
   • DNA pol III attaches to the 3’-end of the primer and covalently joins the free nucleotides together in a 5’ → 3’ direction
   • As DNA strands are antiparallel, DNA pol III moves in opposite directions on the two strands
     • On the leading strand, DNA pol III is moving towards the replication fork and can synthesise continuously
     • On the lagging strand, DNA pol III is moving away from the replication fork and synthesises in pieces (Okazaki fragments)

7. DNA Polymerase I

   • As the lagging strand is synthesised in a series of short fragments, it has multiple RNA primers along its length
   • DNA pol I removes the RNA primers from the lagging strand and replaces them with DNA nucleotides

8. DNA Ligase

   • DNA ligase joins the Okazaki fragments together to form a continuous strand
   • It does this by covalently joining the sugar-phosphate backbones together with a phosphodiester bond