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U.S. Department of Energy
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The impact of energy-related pollutants on chromosome structure: Progress report, May 1, 1987-April 30, 1988

Technical Report ·
OSTI ID:5672360

We have examined at the DNA sequence level the sequence specificity of binding of bis-(1,10)-phenanthroline Cu(I) complex ((Phen)/sub 2/Cu/sup +/), benzo(a)pyrene-7,8-diol-9,10-epoxide, acetylacetoxyaminofluorene, the monoazide derivative of ethidium bromide, and the monoazide derivative of actinomycin D. The techniques utilized are analogous to those of DNA sequencing. Binding sites were mapped in sea urchin and mouse histone genes cloned into pUC plasmids. The sequence specificity of (Phen)/sub 2/Cu/sup +/ induced DNA cleavage was determined by mapping of cleavage sites on >2000 base pairs. We have found that (Phen)/sub 2/Cu/sup +/ has a predominant preference for TAT triplets, and a moderate preference for TGT triplets; and TAAT, TAGPy, and CAGT quartets. Oligopurine and oligopyrimidine tracts are poorly reactive. We have also shown that (Phen)/sub 2/Cu/sup +/ binds in the minor groove at preferred sequences. These reaction specificities are unusual for a DNA ligand and can be explained mainly in terms of a requirement for binding at the minor groove side of a pyrimidine-3', 5'-purine step and the stereolelectronic effects of 2-amino groups of guanine, which inhibit minor groove binding. Studies with synthetic minihelices have confirmed that only the local sequence is required for specificity, and that the 2-amino group of guanine does inhibit binding (for these experients inosine was substituted or guanine). We have examined, at sequence resolution, the formation of alkali-labile sites by each of the other reagents on 500 bp or more of cloned histone genes. Moderate to high specificity was observed in all cases but actinomycin D monoazide. 10 refs., 6 figs.

Research Organization:
Florida State Univ., Tallahassee (USA). Inst. of Molecular Biophysics
DOE Contract Number:
FG05-87ER60588
OSTI ID:
5672360
Report Number(s):
DOE/ER/60588-11; ON: DE88004921
Country of Publication:
United States
Language:
English