Question

Explain how the replication of the lagging strand occurs in E. coli. Include a description of the role of each of the key enzymes involved.

Answer 1

Replication of the lagging strand occurs in E. coli:

The strand which is discontinuosly synthesised in the form of short Okazaki Fragment is called as the lagging strand.

DNA polymerases can only make DNA in the 5' to 3' direction, and this poses a problem during replication. A DNA double helix is always anti-parallel; one strand runs in the 5' to 3' direction, while the other runs in the 3' to 5' direction.this makes the new strands which are antiparallel.

One new strand, which runs 5' to 3' towards the replication fork, is the easy one. This strand is made continuously, because the DNA polymerase is moving in the same direction as the replication fork. This continuously synthesized strand is called the leading strand.

The other new strand, which runs 5' to 3' away from the fork, is trickier. This strand is made in fragments because, as the fork moves forward, the DNA polymerase (which is moving away from the fork) must come off and reattach on the newly exposed DNA. This tricky strand, which is made in fragments, is called the lagging strand.

Okazaki Fragment is 1–2 kb in length. Lagging strand synthesis appears to be a more complicated process as it requires the cyclical repetition of several different reactions in a defined sequence, including priming of the Okazaki fragments, extension, and rapid recycling of the polymerase from a finished fragment to the next primer

Replication of the E. coli chromosome is performed by DNA polymerase III holoenzyme, an asymmetric dimeric enzyme composed of 18 subunits that simultaneously replicates the leading and lagging strand of the replication fork.

It contains two polymerase core units, one for each strand, each consisting of three tightly associated subunits, α, ɛ, and θ. Of these, α is the polymerase - dnaE gene product, ɛ - dnaQ gene product is a 3′ → 5′ exonuclease that performs an editing function, and θ is a small subunit whose function is not known.

several enzymes are also involved in replication of E.coli

The role of each of the key enzymes involved:

DNA pol I: Exonuclease activity removes RNA primer and replaces with newly synthesized DNA.

DNA pol II: Repair function.

DNA pol III: Main enzyme that adds nucleotides in the 5′-3′ direction.

Helicase: Opens the DNA helix by breaking hydrogen bonds between the nitrogenous bases.

Ligase: Seals the gaps between the Okazaki fragments to create one continuous DNA strand.

Primase: Synthesizes RNA primers needed to start replication.

Sliding clamp: Helps to hold the DNA polymerase in place when nucleotides are being added.

Topoisomerase: Helps relieve the stress on DNA when unwinding by causing breaks and then resealing the DNA.

SSB-single stranded binding protein: Binds to single-stranded DNA to avoid DNA rewinding back.