TY - JOUR
T1 - Differences in unwinding of supercoiled DNA induced by the two enantiomers of anti-benzo[a]pyrene diol epoxide
AU - Xu, Rong
AU - Birke, Sheryl
AU - Carberry, Susan E.
AU - Geacintov, Nicholas E.
AU - Swenberg, Charles E.
AU - Harvey, Ronald G.
N1 - Funding Information:
This work was supported by the Department of Energy (Grants DEFGO2-86ER60405, and in part by Grant CA 20851 from the US Public Health Service, Department of Health and Human Resources, awarded by the National Cancer Institute. The assistance of Dr. Y.Mnyukh with the linear dichroism measurements is gratefully acknowledged.
PY - 1992/12/11
Y1 - 1992/12/11
N2 - 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.
AB - 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.
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U2 - 10.1093/nar/20.23.6167
DO - 10.1093/nar/20.23.6167
M3 - Article
C2 - 1475180
AN - SCOPUS:0027093114
SN - 0305-1048
VL - 20
SP - 6167
EP - 6176
JO - Nucleic acids research
JF - Nucleic acids research
IS - 23
ER -