Polymerase repair

Antibiotics, such as ciprofloxacin, that inhibit normal DNA replication in bacteria can be counteracted by the polymerase repair molecules from the SOS response. However, certain polymerases in the SOS pathway are error-prone in their copying of DNA, which leads to mutations. While such mutations are often lethal to the cell, they can also improve the bacteria s survival. 

DNA/RNA polymerases repair enzymes topoisomerase I/II Intercalators berberine, indoles, isoquinolines Alkylants PAs, aristolochic acids, cycasin Protein inhibitors camptothecin... 

RNA polymerase >Repair enzymes >Topoisomerase I/II >DNA-polymerase Intercalators Alkylants Protein inhibitors... 

Polymerase II (pol II) is mostly involved in proofreading and DNA repair. Polymerase I (pol I) completes chain synthesis between Okazaki fragments on the lagging strand. Eukaryotic cells have counterparts for each of these enzymes plus some additional ones. A comparison is shown in Table 36—6. 

Figure 36-23. Base excision-repair of DNA. The enzyme uracil DNA glycosylase removes the uracil created by spontaneous deamination of cytosine in the DNA. An endonuclease cuts the backbone near the defect then, after an endonuclease removes a few bases, the defect is filled in by the action of a repair polymerase and the strand is rejoined by a ligase. (Courtesy of B Alberts.)...

Figure 36-24. Nucleotide excision-repair. This mechanism is employed to correct larger defects in DNA and generally involves more proteins than either mismatch or base excision-repair. After defect recognition (indicated by XXXX) and unwinding of the DNA encompassing the defect, an excision nuclease (exinucle-ase) cuts the DNA upstream and downstream of the defective region. This gap is then filled in by a polymerase (5/e in humans) and religated.

Cerklewski and Forbes 1976). Also, excess zinc protects zinc-containing enzymes like ALAS, ferrochelatase, and ALAD. In vivo, aqueous solution containing zinc administered to rats significantly reduced the genotoxic effects induced by lead (Kowalska-Wochna et al. 1988). It was postulated that zinc s protective action may be related to its functioning in DNA and RNA polymerases and consequent enhancement of cell repair processes. 

Genetic recombination arises by exchange of homologous segments of DNA between viral genomes, most often during the replication process. The enzymes involved in recombination are DNA polymerases, endonucleases, and ligases, which also play a role in DNA repair and synthesis processes. 

DNA polymerase I is a nonessential enzyme, since viable E. coli mutants lack it (pol A). This conclusion is complicated, however, since the enzyme catalyzes three separate chemical reactions. It polymerizes deoxyribonucleoside triphosphates, and it has two exonucleolytic activities, a 3 to 5 activity and a 5 to 3 activity. The pol A - mutants lack only the polymerization activity. Other mutants lacking both the polymerase and the 5 to 3 exonuclease activity are lethal. Thus the exonuclease function is the more important one. This fits with the role of this enzyme in removing damaged DNA segments (DNA repair) and in removing covalently attached RNA from DNA chains. We will later see that small RNAs serve as primers of DNA synthesis. 

This is a smaller, stable enzyme that has been highly purified. It is immunologically distinct from the other polymerases, indicating that it is not merely a subunit of the larger polymerases. Polymerase p is undoubtedly a repair enzyme. 

There s not just one DNA polymerase there s a whole army. DNA replication actually occurs in large complexes containing many proteins and sometimes many polymerases. In eukaryotic cells we have to replicate both mitochondrial and nuclear DNA, and there are specific DNA polymerases for each. In addition to DNA replication, you have to make new DNA when you repair. Consequently, the function may be specialized for repair or replication. There can also be specialization for making the leading or lagging strand. Some of the activities of DNA polymerases from eukaryotes and prokaryotes are shown in the table on the next page. 

Low concentrations (10 /iM or less) of mercury have little effect on cellular viability and stimulate RNA and DNA synthesis, whereas higher concentrations are cytotoxic and inhibit DNA, RNA and protein synthesis [145, 146, 246-248 ]. Mercuric chloride is able to selectively block CHO (Chinese hamster ovary) cells in S phase, which is related to the chemical reactivity and uptake into the cells [249], The cytotoxicity of mercury(II) compounds is probably related to their ability to inhibit DNA polymerase a activity and inhibit not only DNA synthesis but also DNA repair [250, 251]. 

The Polymerase Chain Reaction. In the past, a major drawback of hybridization assays was their need for relatively large amounts of sample DNA to compensate for their low sensitivity. This problem has been surmounted in recent years by the development of powerful enzymatic techniques that can exponentially replicate specific DNA sequences in the test tube. With these techniques it is now possible to analyze vanishingly small samples that initially contain fewer than 10 copies of the sequence of interest. The new methods take advantage of the chemical properties of nucleic acids and of highly specialized enzymes that can repair and replicate DNA in vitro. 

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