DNA polymerases are a group of enzymes required for DNA synthesis. Arthur Kornberg purified and characterized DNA polymerase from E.coli for the first time. It is a single-chain polypeptide now known as DNA polymerase-I. Scientists have now found five DNA polymerases in E. coli.
The main function of DNA polymerases is to synthesize DNA. They do so by adding nucleotides at 3’-OH group of the growing DNA strand.
DNA Polymerase Structure and Types
The structure of most of the DNA polymerases resembles a hand, which is holding active sites. The active site of the enzyme has two parts. At the insertion site, nucleotides are added. After adding, the newly formed base-pair migrates to the post-insertion site.
Prokaryotic DNA Polymerase
There are five DNA polymerases identified in E.coli. All the DNA polymerases differ in structure, functions and rate of polymerization and processivity.
DNA Polymerase I is coded by polA gene. It is a single polypeptide and has a role in recombination and repair. It has both 5’→3’ and 3’→5’ exonuclease activity. It removes the RNA primer from lagging strand by 5’→3’ exonuclease activity and also fills the gap.
DNA Polymerase II is coded by polB gene. It is made up of 7 subunits. It’s main role is in repair and also a backup of DNA polymerase III. It has 3’→5’ exonuclease activity.
DNA Polymerase III is the main enzyme for replication in E.coli. It is coded by polC gene. The polymerization and processivity rate is maximum in DNA polymerase III. It also has proofreading 3’→5’ exonuclease activity.
DNA polymerase III of E.coli is made up of a total of 13 subunits, which comprises 9 different types of subunits.
- It consists of two core domains made up of 𝜶, 𝟄, and 𝞱 subunits. It is attached to the 𝝲 complex or clamp-loading complex, which is made up of five subunits, 𝞽2𝝲𝝳𝝳’. Additional subunits 𝟀 and 𝟁 are attached to the clamp-loading complex. 𝞫 subunits make two clamps with a dimer each. They increase the processivity of the DNA polymerase III.
DNA Polymerase IV is coded by dinB gene. Its main role is in DNA repair during SOS response, when DNA replication is stalled at the replication fork. DNA polymerase II, IV and V are translesion polymerases.
DNA Polymerase V is also involved in translesion synthesis during SOS response and DNA repair. It is made up of UmuC monomer and UmuD dimer.
Like prokaryotic cells, eukaryotic cells also have many DNA polymerases, which perform different functions, e.g. mitochondrial DNA replication, nuclear DNA replication, etc. The nuclear DNA replication is mainly done by DNA polymerase 𝝳 and 𝜶. There are at least 15 DNA polymerases identified in human beings.
- DNA polymerase 𝝳 – It is the main enzyme for replication in eukaryotes. It also has 3’→5’ exonuclease activity for proofreading.
- DNA polymerase 𝜶 – The main function of DNA polymerase 𝜶 is to synthesize primers. The smaller subunit has a primase activity. The largest subunit has polymerization activity. It forms a primer for Okazaki fragments, which is then extended by DNA polymerase 𝝳.
- DNA polymerase 𝟄 – The main function is DNA repair. It removes primers for Okazaki fragments from the lagging strand.
- DNA polymerase 𝝲 – It is the main replicative enzyme for mitochondrial DNA.
Also Check: MCQs on DNA Replication
How does DNA Polymerase work?
The reaction is phosphoryl group transfer. The 3’-OH group of the growing strand acts as a nucleophile and attacks the incoming deoxyribonucleoside triphosphate at the 𝜶-phosphorus, leading to phosphodiester bond formation. Inorganic phosphate is released in the reaction.
(dNMP)n + dNTP → (dNMP)n+1 + PPi
All the DNA polymerases require two Mg ions at the active site. It is important to note that DNA polymerase can only add nucleotides at the 3′ end of the growing strand, that is why replication always occurs in the 5’→3’ direction. They cannot initiate the formation of new DNA.
They need a template strand, which guides the polymerisation reaction. They also need a primer for their action as they can only add nucleotides at 3’ OH group. The primer can be a short segment of RNA, DNA or both. Generally, the primer is an RNA oligonucleotide in the living system.
After adding a nucleotide, the DNA polymerase can either dissociate or move along to add more nucleotides. It depends on the processivity of DNA polymerase and it differs in different DNA polymerases.
Replication is a highly accurate process and even the change in a single nucleotide can cause mutation. To avoid this there are two mechanisms by which DNA polymerases ensure that there are no discrepancies.
- The geometry of the active sites allows only the correct nucleotide base pairs to fit. But this is not sufficient and it is seen that it can add an incorrect nucleotide after correctly adding 104 to 105 nucleotides.
- To correct this type of errors, DNA polymerase has 3’→5’ exonuclease activity. DNA polymerase checks each of the added nucleotides and removes the nucleotide if there is a mismatch. This is known as proofreading. In DNA polymerase I, there are different active sites for polymerizing and proofreading functions.
This was all about DNA Polymerase. Explore notes on Molecular Basis of Inheritance to know in detail about the replication process, only at BYJU’S.