DNA Polymerase II

Thomas Kornberg, Arthur Kornberg’s son, identified and described the DNA polymerase II of E. coli in 1970. In 1970, Knippers and Moses and Richardson published separate papers on DNA polymerase II.

coli mutants defective in DNA pol I were created in 1969 by De Lucia and Cairns to better understand the in vivo role of this enzyme. This novel mutant strain was more sensitive to ultraviolet light than previously thought, supporting the theory that DNA pol I was involved in repair replication. The mutant grew at the same pace as the natural type, showing that another enzyme involved in DNA replication is present. In simultaneous research done by different labs, this novel polymerase implicated in semiconservative DNA replication was isolated and characterised. DNA polymerase II was the name given to the new polymerase.

Structure

DNA Pol II is an 89.9 kD protein of 783 amino acids encoded by the polB (dinA) gene. DNA Pol II is a globular protein that works as a monomer, whereas many other polymerases form complexes. This monomer is divided into three major regions, which are informally known as the palm, fingers, and thumb. This “hand” wraps itself around a strand of DNA. In order to operate, the complex’s palm contains three catalytic residues that will interact with two divalent metal ions. DNA Pol II has a high number of copies in the cell, roughly 30-50, whereas DNA Pol III has five times less copies.

Types and Function

In E. coli, five DNA polymerases have been found. The structure, roles, rate of polymerization, and processivity of all DNA polymerases varies.

DNA Polymerase I:- The polA gene codes for DNA polymerase I. It’s a single polypeptide that helps with recombination and repair. It shows action for both 5’3′ and 3’5′ exonucleases. By using 5’3′ exonuclease activity, DNA polymerase I removes the RNA primer from the lagging strand while also filling the gap.

DNA Polymerase II:- The polB gene codes for DNA polymerase II. It consists of seven components. Its primary function is in DNA polymerase III repair and as a backup. It has activity as a 3’5′ exonuclease.

DNA Polymerase III: – In E.coli, Poly III is the most important enzyme for replication. The polC gene codes for it. DNA polymerase III has the highest polymerization and processivity rates. It also possesses 3’5′ exonuclease activity for proofreading.

E.coli DNA polymerase III is made up of 13 subunits, each of which is made up of 9 different types of subunits.

It is made up of two core domains that are made up of 𝜶, 𝟄, and 𝞱 subunits. It is connected to the 𝝲 complex, also known as the clamp-loading complex, which consists of five subunits, 𝞽2𝝲𝝳𝝳’.  𝟀 and 𝟁 Attached to the clamp-loading complex are additional subunits and. 𝞫 Subunits combine to form two clamps, each with a dimer. They improve the DNA polymerase III’s processivity.

DNA Polymerase IV: – DinB is the gene that codes for DNA Polymerase IV. When DNA replication is halted at the replication fork during the SOS reaction, it plays a key role in DNA repair. Translesion polymerases are DNA polymerase II, IV, and V.

DNA Polymerase V: – During the SOS response and DNA repair, DNA Polymerase V is also engaged in translesion synthesis. UmuC monomer and UmuD dimer make up this molecule.

Mechanism

Base pairs in the DNA sequence are susceptible to damage during replication. Replication can be slowed down by a broken DNA sequence. DNA Pol II catalyses the repair of nucleotide base pairs to correct a sequence mistake. In vitro investigations have revealed that Pol II interacts with Pol III auxiliary proteins (clamp and clamp loading complex) on occasion, allowing Pol II to access the developing nascent strand. This makes sense in terms of DNA Pol II’s role during DNA replication, because any errors produced by Pol III will be on the expanding strand rather than the conservative strand. The attachment and dissociation of the DNA strand from the catalytic subunit is controlled by DNA Pol II’s N-terminal region. Single-stranded DNA is most likely recognised by two locations in DNA Pol II’s N-terminal domain. One site is in charge of recruiting single-stranded DNA to DNA Pol II, while another is in charge of separating single-stranded DNA from DNA Pol II. When DNA Pol II connects to a substrate, it bonds nucleoside triphosphates to keep the hydrogen bound structure of DNA intact. The right dNTP is then bound, and the subdomains and amino acid residues of the enzyme complex undergo conformational modifications. The rapid rate of repair synthesis is enabled by these conformational alterations.

CONCLUSION

DNA Polymerase II is naturally prevalent in the cell, typically five times more than Polymerase III. In the instance of mispairings, Polymerase II’s greater abundance permits it to outnumber Polymerase III. This amount can be raised when the SOS response is triggered, which upregulates the polB gene, resulting in a sevenfold rise in Polymerase II. Despite the fact that Polymerase II can work on both strands, it has been discovered that it prefers the lagging strand over the leading strand.