No abstract

We characterize a component of the E. coli bacterial nucleoid H1a, which accumulates in stationary phase. This protein, identical with the major component of a plasmid-protein complex previously isolated in our laboratory, has a pI close to 7.5. Acrylamide gel electrophoresis and sedimentation in sucrose gradient have shown that the protein H1a induces significant compaction into DNA. This compaction is equivalent to that observed in nucleosome core although it introduces only a slight change in linking number. In addition, the structural change induced in the lactose L8UV5 promoter by H1a results in the decrease in the kinetic of formation of the open complex with RNA polymerase.

Emphasis was placed in this work on the measurement of purine oxidation products generated upon nano- and picosecond UV laser biphotonic photolysis of 2‘-deoxyadenosine, 2‘-deoxyguanosine, calf thymus DNA, and a synthetic duplex oligonucleotide (37-mer) in aerated aqueous solutions. The overall formation of 8-oxo-7,8-dihydro-2‘-deoxyguanosine and 8-oxo-7,8-dihydro-2‘-deoxyadenosine was determined using a HPLC-electrochemical detection assay. Denaturing gel electrophoresis analysis in association with a formamidopyrimidine−DNA glycosylase treatment was applied 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‘-deoxyadenosine were shown to be minor decomposition products of the related nucleoside 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, was observed in DNA. It should be noted that the yield of 8-oxo-7,8-dihydro-2‘-deoxyguanosine was about 3-fold lower in heat-denatured DNA than in double-stranded DNA. These observations provide strong support to the significant involvement of base stacking and probably DNA solvatation in the chemical reactions of the purine radical cations. Other interesting information dealt with the similarity in the level of 8-oxo-7,8-dihydro-2‘-deoxyguanosine and the number of formamidopyrimidine−DNA glycosylase sensitive guanine lesions. This strongly suggests that the latter formamidopyrimidine−DNA glycosylase purine nucleoside is the major DNA photodamaged product recognized by the DNA repair glycosylase. Another striking feature is the almost 10-fold decrease in the saturation dose E s for the two-quantum ionization of the guanine base in double-stranded DNA as compared to that observed for free 2‘-deoxyguanosine. This can be explained by either an enhancement of the quantum yield of photoionization from the intermediate excited state in DNA (φ2) and/or hole migration with preferential trapping by guanine residues.