Characterization of bleomycin cleavage sites in strongly bound hairpin DNAs

Rachel A. Giroux, Sidney Hecht

Research output: Contribution to journalArticlepeer-review

27 Scopus citations


The first complete, systematic study of DNA degradation by bleomycin under conditions analogous to those likely in a therapeutic setting has been carried out. Hairpin DNAs selected for their ability to bind strongly to BLM A 5 were used to determine the relationship between high-affinity DNA binding sites and the cleavage efficiency and selectivity of BLM A5 and deglycoBLM A5 on these DNAs. Of the 10 hairpin DNAs examined, 8 contained at least one 5′-GC-3′ or 5′-GT-3′ cleavage site, which have traditionally been associated with strong cleavage by Fe•BLM. In the hairpin DNAs, these included the strongest cleavage sites for BLM A5 and were generally among those for deglycoBLM A 5. However, numerous additional cleavages were noted, many at sequences not usually associated with (deglyco)BLM-mediated cleavage. The remaining DNAs lacked the preferred (5′-GC-3′ or 5′-GT-3′) BLM cleavage sequences; however, strong cleavage was nonetheless observed at a number of unusual cleavage sites. The most prominent cleavage sequences were 5′-AT-3′, 5′-AA-3′, 5′-GA-3′, and 5′-TT-3′ treatment with Fe(II)•BLM A5 or Fe(II)•deglycoBLM A5 resulted in strong cleavage at these sequences. Additionally, in contrast with BLM A5, which produced cleavage within the randomized and flanking invariant regions, deglycoBLM A5 showed a preference for cleavage in the randomized region of the DNAs. Previous reports have established that deglycoBLM exhibits decreased DNA cleavage efficiency relative to BLM. This was also generally observed when comparing cleavage efficiencies for the strongly bound hairpin DNAs. However, some cleavage bands produced by Fe•deglycoBLM A5 were stronger in intensity than those produced by BLM A5 at concentrations optimal for both compounds. To investigate the chemistry of DNA degradation, selected hairpin DNAs were treated with n-butylamine following cleavage with Fe(II)•BLM A5 or Fe(II)•deglycoBLM A 5 to explore the alkali labile pathway of DNA degradation by BLM. While all 10 DNAs showed evidence of alkali labile products, five DNA hairpins afforded some products formed solely via the alkali labile pathway.

Original languageEnglish (US)
Pages (from-to)16987-16996
Number of pages10
JournalJournal of the American Chemical Society
Issue number47
StatePublished - Dec 1 2010

ASJC Scopus subject areas

  • Catalysis
  • Chemistry(all)
  • Biochemistry
  • Colloid and Surface Chemistry


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