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Repair of 8-methoxypsoralen monoadducts and diadducts in bacteriophages and bacteria
Abstract:
The combined action of 8-methoxypsoralen (8-MOP) and light with lambda greater than 310 nm on bacteriophages and bacteria results in the formation of the following two types of photo-products in the DNA: monoadducts, in which 8-MOP is covalently bound to a pyrimidine base, and diadducts or cross links, in which the 8-MOP is covalently bound to two pyrimidines from complementary strands. The method of repeated irradiation has been proposed for analyzing the degree of lethality of the photoproducts in DNA. According to this method, the preparation is freed of free 8-MOP molecules after the first irradiation and then irradiated for a second time. In this case the monoadducts are converted into cross linkages between the strands. Approximately 3-10(-9) cross links/Dalton-min form in Escherichia coli DNA during the first irradiation. The rate of the formation of cross links drops by a factor of about 2 during the repeated irradiation. It has been shown that the 8-MOP monoadducts are repaired by the uvr system just as efficiently as are lethal photoproducts of the cyclobutane pyrimidine dimer type. Lethal cross linkages in bacteria and phages are repaired by the joint action of the uvr, recA, and lex systems. A scheme has been proposed for the repair of cross linkages in one genome by these systems. The photoreactivating enzyme is inactive on DNA subjected to the combined action of 8-MOP and light. The kinetics of the repair of monadducts in bacteria and phages with various defects in the repair systems have been studied. It has been shown that the products of genes recA and lex take part in the repair process according to an excision-resynthesis method. The use of the method of repeated irradiation with 8-MOP as an express method for detecting repair systems of the uvr type in cells has been proposed.
Insights
8-methoxypsoralen (8-MOP) and light create DNA monoadducts and cross-links in bacteria and phages. Repeated irradiation converts monoadducts to cross-links, aiding in the study of DNA repair mechanisms.
Area of Science:
- Molecular Biology
- Photochemistry
- Genetics
Background:
- 8-methoxypsoralen (8-MOP) combined with UVA light induces DNA damage.
- Two primary photoproducts are formed: monoadducts and diadducts (cross-links).
Purpose of the Study:
- To investigate the formation and lethality of 8-MOP induced DNA photoproducts.
- To elucidate the DNA repair pathways involved in removing these photoproducts.
- To propose a novel method for assessing DNA repair capacity.
Main Methods:
- Repeated irradiation technique to convert monoadducts to cross-links.
- Quantification of cross-link formation rates in Escherichia coli DNA.
- Analysis of DNA repair kinetics in wild-type and mutant bacteria/phages.
Main Results:
- Monoadducts are efficiently repaired by the uvr system, similar to cyclobutane pyrimidine dimers.
- Lethal cross-links are repaired by a coordinated action of uvr, recA, and lex systems.
- The rate of cross-link formation decreases during repeated irradiation.
- The recA and lex gene products are involved in excision-resynthesis repair of monoadducts.
Conclusions:
- 8-MOP photoproducts represent significant DNA lesions requiring complex repair pathways.
- The repeated irradiation method can serve as a rapid assay for detecting uvr-type repair systems.
- Understanding these repair mechanisms is crucial for photochemotherapy and microbial genetics.