Formation of ssb, dsb, and uracil in monofilarly and bifilarly bromouracil-substituted DNA molecules.

Bifilarly BU-substituted ColE 1 plasmid and monofilarly BU-substituted M 13 phage DNA were irradiated with UV light of 313 nm. Using agarose gel electrophoresis and "reversed phase" HPLC technique ssb, dsb induction and uracil formation, respectively, could be detected in the irradiated DNA in dependence on the UV fluence. The analysis of the strandbreaks in bifilar ColE 1 DNA shows a significant part of directly induced dsb. Cross sections of ssb induction from 4.1 m2/J x 10(7) in 28%, 3.9 m2/J x 10(7) in 55% and 3.1 m2/J x 10(7) in 85-90% BU-substituted DNA were calculated. The cross section for dsb induction was found to be 0.04 m2/J x 10(7), estimated from the linear part of the fluence effect curve. In monofilar M 13 DNA a linear fluence effect curve for dsb induction was obtained. Excluding other than the direct production of dsb by using an in vitro approach for M 13 DNA, the results strongly support the hypothesis that dsb can be induced by one photochemical absorption event. The cross section for ssb was 3.8 m2/J x 10(7) and for dsb 0.05 m2/J x 10(7) in 41.5% monofilarly BU-substituted M 13 DNA. The comparison of ssb, dsb, and uracil production in bifilar and monofilar DNA with similar BU substitution showed no significant difference between the two DNA systems (ColE 1, M 13), indicating that the location of BU molecules in one or in both DNA strands will not lead to a different number of lesions after UV313 exposure.