Repair replication in Escherichia coli as measured by the photolysis of bromodeoxyuridine

Abstract

The ability to selectively photolyze bromouracil-(BrUra-)containing repaired regions in cellular DNA has allowed us to estimate the average size of repaired regions in ultraviolet (UV) light-irradiated Escherichia coli. Cells were labeled with thymidine-(3)H, irradiated at 254 nm, and incubated in nonradioactive bromodeoxyuridine (BrdUrd). After incubation the cells were exposed to 10(6) ergs.mm(-2) at 313 nm, lysed, and sedimented in alkaline sucrose gradients so as to measure the average molecular weight of single DNA strands. In strains that had excised approximately 45 cyclobutane pyrimidine dimers/10(8) daltons, the 313 nm treatment resulted in approximately 6 single-strand breaks/10(8) daltons. In an excisionless strain, the same treatment resulted in only 1.5 breaks/10(8) daltons. From the determination of the sensitivities of fully substituted DNAs to 313 nm light, we calculate that the repaired regions in excising strains of E. coli contain an average of 4-6 BrUra residues. Photoreactivation experiments indicate that the excision of pyrimidine dimers in the presence of BrdUrd is the primary source of repaired regions selectively photolyzed by 313 nm radiation.