ENZYMATIC RECOGNITION AND BIOLOGICAL EFFECTS OF DNA DAMAGE INDUCED BY3-CARBETHOXYPSORALEN PLUS UVA

The specific recognition of DNA modifications by repair endonucleases was used to characterize damage induced by 3-carbethoxypsoralen (3-CPs ) plus UvA in M13mp8 replicative form I (RF-I) DNA. Under the conditio ns used, 3-CPs plus UVA generates DNA base modifications which are rec ognized by the UvrABC complex and the Fpg protein of E. coli. The rate of formation of UvrABC sensitive sites is 3-4-fold higher than that o f Fpg sensitive sites. In addition a small number of sites of base los s (sensitive to Nfo protein) were observed. M13mp8 RF-I DNA treated wi th 3-CPs plus UVA was tested for transfection efficiency in E. coli mu tants defective in either Fpg protein and/or UvrABC complex. The survi val of 3-CPs plus UVA damaged M13mp8 RF-I DNA was significantly reduce d when transfected into uvrA mutants compared to that in the wild-type strain. On the other hand, the survival of 3-CPs plus UVA damaged RF- I DNA was not altered in fpg-1 mutants. These results show that nucleo tide excision repair mediated by the UvrABC complex is the major repai r pathway involved in the elimination of lethal lesions induced in DNA by 3-CPs plus UVA. Our data suggest that in vitro exposure of M13mp8 RF-I DNA to 3-CPs plus UVA produces predominantly thymine photoadditio n and to a lesser extent guanine photooxidation partially due to singl et oxygen generated during photoreaction. The photoaddition products a re primarly responsible for the observed lethal effect.