Copper-dependent cleavage of DNA by bleomycin

Guy M. Ehrenfeld, Joshua B. Shipley, Sidney Hecht, Hiroshi Sugiyama, Eric C. Long, Jacques H. Van Boom, Gijs A. Van Der Marel, Norman J. Oppenheimer, Sidney M. Hecht

Research output: Contribution to journalArticle

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Abstract

DNA strand scission by bleomycin in the presence of Cu and Fe was further characterized. It was found that DNA degradation occurred readily upon admixture of Cu(I) or Cu(II) + dithiothreitol + bleomycin, but only where the order of addition precluded initial formation of Cu(II)-bleomycin or where sufficient time was permitted for reduction of the formed Cu(II)-bleomycin to Cu(I)-bleomycin. DNA strand scission mediated by Cu + dithiothreitol + bleomycin was inhibited by the copper-selective agent bathocuproine when the experiment was carried out under conditions consistent with Cu chelation by bathocuproine on the time scale of the experiment. Remarkably, it was found that the extent of DNA degradation obtained with bleomycin in the presence of Fe and Cu was greater than that obtained with either metal ion alone. A comparison of the sequence selectivity of bleomycin in the presence of Cu and Fe using 32P-end-labeled DNA duplexes as substrates revealed significant differences in sites of DNA cleavage and in the extent of cleavage at sites shared in common. For deglycobleomycin and decarbamoylbleomycin, whose metal ligation is believed to differ from that of bleomycin itself, it was found that the relative extents of DNA cleavage in the presence of Cu were not in the same order as those obtained in the presence of Fe. The bleomycin-mediated oxygenation products derived from cis-stilbene were found to differ in type and amount in the presence of added Cu vs. added Fe. Interestingly, while product formation from cis-stilbene was decreased when excess Fe was added to a reaction mixture containing 1:1 Fe(III) and bleomycin, the extent of product formation was enhanced almost 4-fold in reactions that contained 5:1, as compared to 1:1, Cu and bleomycin. The results of these experiments are entirely consistent with the work of Sugiura [Sugiura, Y. (1979) Biochem. Biophys. Res. Commun. 90, 375-383], who first demonstrated the generation of reactive oxygen species upon admixture of O2 and Cu(I)-bleomycin.

Original languageEnglish (US)
Pages (from-to)931-942
Number of pages12
JournalBiochemistry
Volume26
Issue number3
StatePublished - 1987
Externally publishedYes

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DNA Cleavage
Bleomycin
Copper
DNA
Stilbenes
Dithiothreitol
Metals
Ligation
Reactive Oxygen Species
copper bleomycin
Degradation
Ions
Oxygenation
Experiments
Chelation
Metal ions
indium-bleomycin

ASJC Scopus subject areas

  • Biochemistry

Cite this

Ehrenfeld, G. M., Shipley, J. B., Hecht, S., Sugiyama, H., Long, E. C., Van Boom, J. H., ... Hecht, S. M. (1987). Copper-dependent cleavage of DNA by bleomycin. Biochemistry, 26(3), 931-942.

Copper-dependent cleavage of DNA by bleomycin. / Ehrenfeld, Guy M.; Shipley, Joshua B.; Hecht, Sidney; Sugiyama, Hiroshi; Long, Eric C.; Van Boom, Jacques H.; Van Der Marel, Gijs A.; Oppenheimer, Norman J.; Hecht, Sidney M.

In: Biochemistry, Vol. 26, No. 3, 1987, p. 931-942.

Research output: Contribution to journalArticle

Ehrenfeld, GM, Shipley, JB, Hecht, S, Sugiyama, H, Long, EC, Van Boom, JH, Van Der Marel, GA, Oppenheimer, NJ & Hecht, SM 1987, 'Copper-dependent cleavage of DNA by bleomycin', Biochemistry, vol. 26, no. 3, pp. 931-942.
Ehrenfeld GM, Shipley JB, Hecht S, Sugiyama H, Long EC, Van Boom JH et al. Copper-dependent cleavage of DNA by bleomycin. Biochemistry. 1987;26(3):931-942.
Ehrenfeld, Guy M. ; Shipley, Joshua B. ; Hecht, Sidney ; Sugiyama, Hiroshi ; Long, Eric C. ; Van Boom, Jacques H. ; Van Der Marel, Gijs A. ; Oppenheimer, Norman J. ; Hecht, Sidney M. / Copper-dependent cleavage of DNA by bleomycin. In: Biochemistry. 1987 ; Vol. 26, No. 3. pp. 931-942.
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abstract = "DNA strand scission by bleomycin in the presence of Cu and Fe was further characterized. It was found that DNA degradation occurred readily upon admixture of Cu(I) or Cu(II) + dithiothreitol + bleomycin, but only where the order of addition precluded initial formation of Cu(II)-bleomycin or where sufficient time was permitted for reduction of the formed Cu(II)-bleomycin to Cu(I)-bleomycin. DNA strand scission mediated by Cu + dithiothreitol + bleomycin was inhibited by the copper-selective agent bathocuproine when the experiment was carried out under conditions consistent with Cu chelation by bathocuproine on the time scale of the experiment. Remarkably, it was found that the extent of DNA degradation obtained with bleomycin in the presence of Fe and Cu was greater than that obtained with either metal ion alone. A comparison of the sequence selectivity of bleomycin in the presence of Cu and Fe using 32P-end-labeled DNA duplexes as substrates revealed significant differences in sites of DNA cleavage and in the extent of cleavage at sites shared in common. For deglycobleomycin and decarbamoylbleomycin, whose metal ligation is believed to differ from that of bleomycin itself, it was found that the relative extents of DNA cleavage in the presence of Cu were not in the same order as those obtained in the presence of Fe. The bleomycin-mediated oxygenation products derived from cis-stilbene were found to differ in type and amount in the presence of added Cu vs. added Fe. Interestingly, while product formation from cis-stilbene was decreased when excess Fe was added to a reaction mixture containing 1:1 Fe(III) and bleomycin, the extent of product formation was enhanced almost 4-fold in reactions that contained 5:1, as compared to 1:1, Cu and bleomycin. The results of these experiments are entirely consistent with the work of Sugiura [Sugiura, Y. (1979) Biochem. Biophys. Res. Commun. 90, 375-383], who first demonstrated the generation of reactive oxygen species upon admixture of O2 and Cu(I)-bleomycin.",
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AU - Hecht, Sidney

AU - Sugiyama, Hiroshi

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AU - Van Boom, Jacques H.

AU - Van Der Marel, Gijs A.

AU - Oppenheimer, Norman J.

AU - Hecht, Sidney M.

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N2 - DNA strand scission by bleomycin in the presence of Cu and Fe was further characterized. It was found that DNA degradation occurred readily upon admixture of Cu(I) or Cu(II) + dithiothreitol + bleomycin, but only where the order of addition precluded initial formation of Cu(II)-bleomycin or where sufficient time was permitted for reduction of the formed Cu(II)-bleomycin to Cu(I)-bleomycin. DNA strand scission mediated by Cu + dithiothreitol + bleomycin was inhibited by the copper-selective agent bathocuproine when the experiment was carried out under conditions consistent with Cu chelation by bathocuproine on the time scale of the experiment. Remarkably, it was found that the extent of DNA degradation obtained with bleomycin in the presence of Fe and Cu was greater than that obtained with either metal ion alone. A comparison of the sequence selectivity of bleomycin in the presence of Cu and Fe using 32P-end-labeled DNA duplexes as substrates revealed significant differences in sites of DNA cleavage and in the extent of cleavage at sites shared in common. For deglycobleomycin and decarbamoylbleomycin, whose metal ligation is believed to differ from that of bleomycin itself, it was found that the relative extents of DNA cleavage in the presence of Cu were not in the same order as those obtained in the presence of Fe. The bleomycin-mediated oxygenation products derived from cis-stilbene were found to differ in type and amount in the presence of added Cu vs. added Fe. Interestingly, while product formation from cis-stilbene was decreased when excess Fe was added to a reaction mixture containing 1:1 Fe(III) and bleomycin, the extent of product formation was enhanced almost 4-fold in reactions that contained 5:1, as compared to 1:1, Cu and bleomycin. The results of these experiments are entirely consistent with the work of Sugiura [Sugiura, Y. (1979) Biochem. Biophys. Res. Commun. 90, 375-383], who first demonstrated the generation of reactive oxygen species upon admixture of O2 and Cu(I)-bleomycin.

AB - DNA strand scission by bleomycin in the presence of Cu and Fe was further characterized. It was found that DNA degradation occurred readily upon admixture of Cu(I) or Cu(II) + dithiothreitol + bleomycin, but only where the order of addition precluded initial formation of Cu(II)-bleomycin or where sufficient time was permitted for reduction of the formed Cu(II)-bleomycin to Cu(I)-bleomycin. DNA strand scission mediated by Cu + dithiothreitol + bleomycin was inhibited by the copper-selective agent bathocuproine when the experiment was carried out under conditions consistent with Cu chelation by bathocuproine on the time scale of the experiment. Remarkably, it was found that the extent of DNA degradation obtained with bleomycin in the presence of Fe and Cu was greater than that obtained with either metal ion alone. A comparison of the sequence selectivity of bleomycin in the presence of Cu and Fe using 32P-end-labeled DNA duplexes as substrates revealed significant differences in sites of DNA cleavage and in the extent of cleavage at sites shared in common. For deglycobleomycin and decarbamoylbleomycin, whose metal ligation is believed to differ from that of bleomycin itself, it was found that the relative extents of DNA cleavage in the presence of Cu were not in the same order as those obtained in the presence of Fe. The bleomycin-mediated oxygenation products derived from cis-stilbene were found to differ in type and amount in the presence of added Cu vs. added Fe. Interestingly, while product formation from cis-stilbene was decreased when excess Fe was added to a reaction mixture containing 1:1 Fe(III) and bleomycin, the extent of product formation was enhanced almost 4-fold in reactions that contained 5:1, as compared to 1:1, Cu and bleomycin. The results of these experiments are entirely consistent with the work of Sugiura [Sugiura, Y. (1979) Biochem. Biophys. Res. Commun. 90, 375-383], who first demonstrated the generation of reactive oxygen species upon admixture of O2 and Cu(I)-bleomycin.

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