Enzymes and mechanism of replication

In prokaryotes, replication process requires three types of DNA polymerases (DNA polymerase I, II, and III).

DNA polymerase III is the main enzyme involved in DNA
replication.

DNA polymerase I (also known as Kornberg enzyme)and DNA polymerase II are involved in DNA repair mechanism.

Eukaryotes have five types of DNA polymerases that catalyses the polymerization of nucleotides at the 3' OH of the new strand within a short period of time.

E.coli that has 4.6 X 106 bp completes its replication process within 38 minutes. Replication takes place
faster at the same time accurately.

Any error will lead to mutation.

However replication errors are corrected by repair enzymes such as nucleases.

Deoxy nucleotide triphosphate acts as substrate and also provides energy for polymerization reaction.

Replication begins at the initiation site called the site of ‘origin of replication’ (ori).

In prokaryotes, there is only one origin of replication, whereas in eukaryotes with giant DNA molecules, there can be several origins of replication (replicons).

Since the two strands of DNA cannot be separated
throughout at a time (due to large requirement of energy) the replication occurs within a small opening of the DNA helix called as replication fork. Unwinding of the DNA strand is carried out by DNA helicase.

Thus, in one strand (template strand with polarity
3' 5') the replication is continuous and is known as the leading strand while in the other strand (coding strand with polarity 5' 3') replication is discontinuous, known as the lagging strand..

The discontinuously synthesized fragments of the lagging strand (called the Okazaki fragments) are joined by the enzyme DNA ligase.

As they move away in both directions, newly synthesized complementary nucleotides are paired with the existing nucleotides on the parent strand and covalently bonded together by DNA polymerase.

Formation of new strand requires a primer (a short stretch of RNA)for initiation.

The primer produces a 3'OH end on the sequence of ribonucleotides, to which deoxy ribonucleotides are added.

The RNA primer is ultimately removed leaving a gap in the newly synthesized DNA strand.

It is removed from 5' end one by one by the exonuclease activity of DNA polymerase.

Finally, when all the nucleotides are in position, gaps are sealed by the enzyme DNA ligase.

At the point of origin of repliction, the helicases and topoisomerases (DNA gyrase) unwind and pull apart the strands, forming a YShaped structure called the replication fork.

There are two replication forks at each origin.

The two strands of a DNA helix have an antiparallel orientation.

The enzyme DNA polymerase can only catalyse the addition of a nucleotide to the new strands in the 5' 3'
direction, as it can only add nucleotides to the 3' carbon position.