Question

Compare and contrast the three major mechanisms of DNA repair using a VENN Diagram. Mechanisms: Proofreading by DNA Polymerase III during replication, Mismatch Repair, and Nucleotide Excision Repair

Answer 1

The transfer of genetic information from one generation to another generation is done in the form of DNA. The negligible rate of mutations in the DNA ensures the information passed on is correct because any mutation introduced at the gentic level can alter the gentic information and their expression in the organism. Mutations (alterations in the genetic material) do occur but repair mechanisms are also available which maintain the low rate of mutations. Errors during DNA replication and chemical damage being induced to the genetic material are main causes of mutations. DNA undergoes damages such as loss of bases but also undergoes the damages which ocuur due to the presence of natural or unnatural chemicals and radiations. The cell is capable of both detecting the cell damage at the level of the genome or DNA and also repairing the cellular / DNA damages.

The cell has methods of finding the parent DNA strand amongst the parent and the daughter DNA strand. The identification mechanism differs in both the prokaryotes and the eukaryotes. In prokaryotes the parent strand is found to be fully methylated but the daughter strand is not methylated i.e. the daughter DNA duplex is in the hemimethylated state and hence the daughter strand is detected by the DNA polymerase, while in eukaryotes the identification is done by identifying the single-stranded breaks (nicks) in the DNA. The nicks are found maximally in the newly synthesized daughter DNA.

The three mechanisms differ from each other and are explained as following :

a) Proofreading by DNA Polymerase III during replication : During DNA replication DNA polymerase III enzyme's subunit (eta) does proofreading as the DNA replication proceeds. This ensures correct DNA replication and it increases the sensitivity of DNA replication by a factor of 100. The 3' to 5' exonuclease activity of the DNA polymerase causes the removal of any mismatch during the replication process.

b) Mismatch repair : Mismatch repair is usually done after the DNA replication is complete. The sensitivity of DNA replication increases three to four folds by mismatch repair. This repair mechanism looks for the mismatches if any left after DNA proofreading during DNA replication. This occurs only after the second round of replication has occured and the mismatch has resulted in a mutation. Mismatch repair proteins are involved in this mechanism. any MutS scans the entire DNA for the mismatches and forms a kink at the mismatch position and undergoes a conformational change itself. MutS recruits MutL which then recruits MutH an endonuclease which cleaves and creates a nick in the DNA near the site of mismatch. Exonuclease digests the nicked strand creating a single stranded gap which is then filled by the DNA polymerase III and sealed with DNA ligase.

c) Nucleotide Excision Repair : Excision repair involves excision and not repair of the mismatched or damage DNA bases. The undamaged strand serves as the template for the incorporation of correct nucleotide bases. DNA shape gets distorted by the interaction of chemical agents such as a chemical adduct on a base. Often includes the repair of thymine dimers. In E.coli the system involves four proteins UvrA, UvrB, UvrC and UvrD. A long stretch of bases are removed which is then replaced by the use of DNA polymerase III enzyme using the parent strand as the template and is sealed by DNA ligase.

The point indicated as common is 1) which means that the involvement of DNA polymerase III and DNA ligase is common in all the three mechanisms although the role of DNA polymerase III is different in proofreading mechanism.