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We have examined lexA1 uvrA6 and recF143 uvrBdelta derivatives of Escherichia coli K-12 for post-replication repair and DNA synthesis after UV irradiation. Compared to corresponding lex+ rec+ strains, we found that the lexA and recF cells were defective in (1) converting short DNA segments synthesized after irradiation to DNA of normal size; (2) synthesizing high molecular weight DNA after irradiation; (3) transferring pyrimidine dimers from irradiated DNA into unirradiated daughter strands. Our results support the hypothesis that after UV irradiation the formation of large DNA molecules in excision-deficient cells of E.coli depends directly or indirectly upon joining short DNA segments into longer strands, concomitant with the transfer of DNA from irradiated tamplates into unirradiated daughter strands. This process appears to require the activity of lexA and recF genes.
to protein at 0°C. Each reaction contained '4C-labeled ColEl DNA (approximately 100 ng, 5,000 cpm) irradiated with 200 J/m2 at 254 nm. Nicking of DNA was assayed with superhelical 14C-790
We previously reported that endonuclease V of bacteriophage T4 reacts processively with pyrimidine dimers in UV-irradiated DNA, tending to react with all of the dimers on one DNA molecule before reacting with any dimers on another DNA molecule [Lloyd, R. S., Hanawalt, P. C., & Dodson, M. L. (1980) Nucleic Acids Res. 8, 5113-5127]. In this paper we show that this processivity depends upon salt concentration: it can be detected in 10 mM NaCl but not, by our methods, in 100 mM NaCl. In addition, we show that endonuclease V binds to unirradiated DNA in 10 mM NaCl but not in 100 mM NaCl. We conclude that T4 endonuclease V binds to pyrimidine dimers in a two-step process in 10 mM NaCl. It first binds electrostatically to undamaged sections of DNA, and it remains bound during the second step in which it "searches" for pyrimidine dimers. Our conclusion is analogous to the expanded target theory developed for Lac repressor [Berg, O. G., Winter, R. B., & von Hippel, P. H. (1981) Biochemistry 20, 6929-6948].
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