HIGH-INTENSITY UV LASER PHOTOLYSIS OF DNA AND PURINE 2'-DEOXYRIBONUCLEOSIDES - FORMATION OF 8-OXOPURINE DAMAGE AND OLIGONUCLEOTIDE STRAND CLEAVAGE AS REVEALED BY HPLC AND GEL-ELECTROPHORESIS STUDIES

Emphasis was placed in this work on the measurement of purine oxidatio n products generated upon nano- and picosecond UV laser biphotonic pho tolysis of 2'-deoxyadenosine, 2'-deoxyguanosine, calf thymus DNA, and a synthetic duplex oligonucleotide (37-mer) in aerated aqueous solutio ns. The overall formation of 8-oxo-7,8-dihydro-2'-deoxyguanosine and 8 -oxo-7,8-dihydro-2'-deoxyadenosine was determined using a HPLC-electro chemical detection assay. Denaturing gel electrophoresis analysis in a ssociation with a formamidopyrimidine-DNA glycosylase treatment was ap plied to reveal the sites recognized by this DNA repair enzyme. Both 8 -oxo-7,8-dihydro-2'-deoxyguanosine and 8-oxo-7,8-dihydro-2'-deoxyadeno sine were shown to be minor decomposition products of the related nucl eoside purine radical cations in agreement with earlier observations. Interestingly, a dramatic increase in the yield of both photoproducts, this applying particularly to 8-oxo-7,8-dihydro-2'-deoxyguanosine, wa s observed in DNA. It should be noted that the yield of 8-oxo-7,8-dihy dro-2'-deoxyguanosine was about 3-fold lower in heat-denatured DNA tha n in double-stranded DNA. These observations provide strong support to the significant involvement of base stacking and probably DNA solvata tion in the chemical reactions of the purine radical cations. Other in teresting information dealt with the similarity in the level of 8-oxo- 7,8-dihydro-2'-deoxyguanosine and the number of formamidopyrimidine-DN A glycosylase sensitive guanine lesions. This strongly suggests that t he latter formamidopyrimidine-DNA glycosylase purine nucleoside is the major DNA photodamaged product recognized by the DNA repair glycosyla se. Another striking feature is the almost 10-fold decrease in the sat uration dose E-s for the two-quantum ionization of the guanine base in double-stranded DNA as compared to that observed for free 2'-deoxygua nosine, This can be explained by either an enhancement of the quantum yield of photoionization from the intermediate excited state in DNA (p hi 2) and/or hole migration with preferential trapping by guanine resi dues.