Differences in unwinding of supercoiled DNA induced by the two enantiomers of anti-benzo[a]pyrene diol epoxide

Rong Xu, Sheryl Birke, Susan E. Carberry, Nicholas E. Geacintov, Charles E. Swenberg, Ronald G. Harvey

Research output: Contribution to journalArticlepeer-review


The unwinding of supercoiled φX174 RFI DNA induced by the tumorigenic (+) and non-tumorigenic (-) enantiomers of trans-7,8-dihydroxy-anti-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE) has been investigated by agarose slab-gel and ethidium titration tube gel electrophoresis. The differences in adduct conformations were verified by flow linear dichroism techniques. Both enantiomers cause a reversible unwinding by the formation of noncovalent intercatative complexes. The effects of covalently bound BPDE residues on the electrophoretic mobilities of the RF I DNA form in agarose gels were investigated in detail in the range of binding ratios rb {equivalent to} 0.0-0.06 (covalently bound BPDE residues/nucleotide). In this range of rb values, there is a striking difference in the mobilities of (+)-BPDE- and (-)-BPDE-adducted φX174 DNA in agarose slab-gels, the covalently bound (+)-BPDE residues causing a significantly greater retardation than (-)-BPDE residues. Increasing the level of covalent adducts beyond rb {equivalent to} 0.06 in the case of the (+)-BPDE enantiomer, leads to further unwinding and a minimum in the mobliities (corresponding to comigration of the nicked form and the covalently closed relaxed modified form) at rb 0.10 ± 0.01; at still higher rb values, rewinding of the modified DNA in the opposite sense is observed. From the minimum in the mobility, a mean unwinding angle (per BPDE residue) of θ = 12±1.5° is determined, which is in good agreement the value of θ = 11 ± 1.8° obtained by the tube gel titration method. Using this latter method, values of θ = 6.8±1.7° for (-)-BPDE-{equivalent to}X174 adducts are observed. It is concluded that agarose slab gel techniques are not suitable for determining unwinding angles for (-)-BPDE-modified {equivalent to}X174 DNA because the alterations in the tertiary structures for rb < 0.06 are too small to cause sufficiently large changes in the electrophoretic mobilities. The major trans (+)-BPDE-N2-guanosine covalent adduct is situated at external binding sites and the mechanisms of unwinding are therefore different from those relevant to noncovalent intercalative BPDE-DNA complexes or to classical intercalating drug molecules; a flexible hinge joint and a widening of the minor groove at the site of the lesion may account for the observed unwinding effects. The more heterogeneous (-)-BPDE-nucleoside adducts (involving cis and trans N and adenosine adducts) are less effective in causing unwinding of supercoiled DNA for reasons which remain to be elucidated.

Original languageEnglish (US)
Pages (from-to)6167-6176
Number of pages10
JournalNucleic acids research
Issue number23
StatePublished - Dec 11 1992

ASJC Scopus subject areas

  • Genetics


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