Mutagenic SOS repair of site-specific psoralen damage in plasmid pBR322

The mutagenic repair of psoralen damage was examined by transforming Escherichia coli with psoralen-treated pBR322. Plasmid DNA randomly reacted with psoralen was repaired only when the E. coli was uvrA⁺ and recA⁺, and only when the cells were pre-irradiated with far-ultraviolet light. The recA dependence and requirement for pre-irradiation are characteristics of SOS repair.Psoralens were placed specifically near the BamHI site, in the tetracycline-resistance gene of pBR322, using a sulfhydryl-containing psoralen derivative. Repair of this damage also required pre-irradiation of the host cells. This repair was accompanied by a 4% frequency of mutagenesis to a tetraeycline-sensitive phenotype. Sequence analysis of these mutant plasmids revealed that 75% had mutations within the targeted region, while 25% had no sequence changes within 100 bases of the BamHI site. In up to five independent isolates only one kind of mutation was observed at each site, suggesting that mutagenic SOS repair is influenced by DNA structure at the site of the psoralen. Most mutations were transitions, primarily G-C to A-T changes. Some transitions occurred at sites where psoralen crosslinks could not have formed, and these may have arisen from the repair of psoralen monoadducts.