Absence Of Strand Breaks Research Articles

Restriction-modification system with methyl-inhibited base excision and abasic-site cleavage activities.

The restriction-modification systems use epigenetic modification to distinguish between self and nonself DNA. A modification enzyme transfers a methyl group to a base in a specific DNA sequence while its cognate restriction enzyme introduces breaks in DNA lacking this methyl group. So far, all the restriction enzymes hydrolyze phosphodiester bonds linking the monomer units of DNA. We recently reported that a restriction enzyme (R.PabI) of the PabI superfamily with half-pipe fold has DNA glycosylase activity that excises an adenine base in the recognition sequence (5′-GTAC). We now found a second activity in this enzyme: at the resulting apurinic/apyrimidinic (AP) (abasic) site (5′-GT#C, # = AP), its AP lyase activity generates an atypical strand break. Although the lyase activity is weak and lacks sequence specificity, its covalent DNA–R.PabI reaction intermediates can be trapped by NaBH4 reduction. The base excision is not coupled with the strand breakage and yet causes restriction because the restriction enzyme action can impair transformation ability of unmethylated DNA even in the absence of strand breaks in vitro. The base excision of R.PabI is inhibited by methylation of the target adenine base. These findings expand our understanding of genetic and epigenetic processes linking those in prokaryotes and eukaryotes.

Metal-ion assisted, radiation-induced conversion of B- to Z-DNA in poly (dGdC) and two natural DNAs.

Metal-ion assisted, gamma-radiation-induced B-Z conformation changes have been observed with UV and circular dichroism spectroscopy for poly (dGdC), calf thymus and herring testis DNA. These conformational changes are similar to those induced by increasing multivalent metal ion concentrations in DNA containing alternating purine and pyrimidine base sequences. In both the metal-ion-induced and the metal-ion-assisted, radiation-induced conformation changes, the conversions were from the right-handed B-DNA to the left-handed Z-DNA conformation. It is proposed that radiation-induced DNA strand breaks markedly reduce the high activation energy barrier in the metal ion-driven B-Z conformation conversion and allow much smaller metal ion concentrations to induce this conversion than in the absence of strand breaks. The biological importance of such radiation-induced conformational changes is discussed in terms of the potential significance of the Z-DNA conformation in the control of the DNA transcription process.

Chromium (VI)-induced DNA damage in chick embryo liver and blood cells in vivo.

Uptake of chromium (VI) and subsequent induction of DNA damage was examined in liver and blood cells of 14-day chick embryos after injection of sodium dichromate onto the inner shell membrane. Maximal loss of chromium from the inner shell membrane and distribution of chromium in liver, lung and blood was observed 2 h after injection. DNA strand breaks, interstrand cross-links and DNA--protein cross-links were measured using the alkaline elution technique. In chick embryo liver, chromium (VI) induced DNA cross-links in the absence of strand breaks. Maximal DNA cross-linking was detected in the liver 8 h after injection. Little or no DNA damage remained in the liver 10-24 h after injection. In contrast, chromium (VI) induced DNA strand breaks in the absence of cross-links in chick embryo blood cells. Maximal DNA strand breakage was observed in blood cells 8 h after injection. High levels of DNA strand breaks were present in blood cells even 24 h after treatment. These intra-embryonic tissue differences in chromium (VI)-induced DNA damage may be a result of the differences in glutathione, cytochrome P-450, other pathways of chromium (VI) metabolism or chromatin organization which exist in liver and blood cells.

Absence of strand breaks in deoxyribonucleic acid treated with metronidazole.

The deoxyribonucleic acid (DNA)-degrading potential of metronidazole was evaluated in vitro by three techniques: determination of melting curve, measurement of viscosity, and centrifugation in neutral or alkaline sucrose gradients. Studies were performed on calf thymus DNA and on (3)H-labeled or unlabeled pneumococcal and T7 phage DNA after treatment with metronidazole alone or metronidazole reduced by sodium dithionite in the presence of DNA. This latter process is known to elicit covalent binding of metronidazole to DNA. Reduced or unreduced metronidazole had no effect on the melting properties, viscosity, or sedimentation velocity of the nucleic acids studied. Sodium dithionite alone, however, caused a 25% decrease in the intrinsic viscosity of pneumococcal DNA, and decreased the sedimentation velocity of pneumococcal and T7 phage DNA in both neutral and alkaline sucrose gradients. These data suggest that degradation of DNA is not important in the interaction of metronidazole with nucleic acids, an interaction assumed relevant to the cytotoxic, radiosensitizing, and mutagenic activities of this compound.