DNA replication is the process by which a single strand of DNA is duplicated to produce two identical strands of DNA. It occurs during the S phase of Mitosis and ensures that any Daughter cells produced during mitosis will be genetically identical.
DNA replication is semi conservative, as proposed by Watson and Crick in 1953. This means that both of the two strands of the parental DNA molecule are used as templates upon which complementary strands of DNA will be built, resulting in the formation of DNA with one strand of parental DNA and one strand of new DNA.
Replication initiation begins at specific sites within the DNA called origin of replication. Circular prokaryotic DNA has one origin of replication (oriC) while linear eukaryotic DNA has multiple origins. The origin of replication has protein binding sites to which an initiator protein can bind. This protein recruits another enzyme called helicase and a special group of proteins called single stranded DNA binding proteins. The initiator protein also uses ATP to unwind the DNA slightly.
The helicase binds to the unwound DNA and continues to unwind it further and moves the replication fork along. As the helicase unwinds, the separated strands are prevented from rewinding by the single-stranded binding proteins. Next, the enzyme primase synthesizes a short sequence of RNA primer complementary to the template. At the 3’ end of the leading strand which runs 3’ to 5’. This primer is used as the starting point for DNA polymerase III to begin forming the complementary DNA strand in the 5’ to 3’ direction.
The lagging strand runs in the 5’ to 3’ direction so the complementary strand has to run in the 3’ to 5’ direction. However, since DNA polymerase III can only add new nucleotides to the 3’ hydroxyl group of the previous nucleotide it can only synthesise strands in the 5’ to 3’ direction so synthesis on of a complement for the lagging strand is discontinuous. For this reason, multiple primers need to be formed on the lagging strand so DNA polymerase can synthesize in the 5’ to 3’ direction from every primer. These fragments of DNA are called Okazaki fragments.
The polymerase III enzyme also has a 3' to 5' exonuclease proofreading activity by which it makes sure that the new nucleotide base being added to synthesize the complementary strands are accurately paired with the correct complementary bases on the template strand.
Following polymerase III, another enzyme called polymerase I with 5’ to 3’ exonuclease activity, begins to remove the primers by replacing the ribonucleotides with deoxyribonucleotides. On the complement of the lagging strand, the fragments of DNA sequence are linked together by DNA ligase.
- ↑ Hardin, J., Becker, W., Kleinsmith, L. and Bertoni, G. (2012). Becker's world of the cell. Boston: Pearson, p.563