A Short DNA Sequence Confers Strong Bleomycin Binding to Hairpin DNAs

Abstract

Bleomycins A5 and B2 were used to study the structural features in hairpin DNAs conducive to strong BLM–DNA interaction. Two members of a 10-hairpin DNA library previously found to bind most tightly to these BLMs were subsequently noted to share the sequence 5′-ACGC (complementary strand sequence 5′-GCGT). Each underwent double-strand cleavage at five sites within, or near, an eight base pair region of the DNA duplex which had been randomized to create the original library. A new hairpin DNA library was selected based on affinity for immobilized Fe(III)·BLM A5. Two of the 30 newly identified DNAs also contained the sequence 5′-ACGC/5′-GCGT. These DNAs bound to the Fe(II)·BLMs more tightly than any DNA characterized previously. Surface plasmon resonance confirmed tight Fe(III)·BLM B2 binding and gave an excellent fit for a 1:1 binding model, implying the absence of significant secondary binding sites. Fe(II)·BLM A5 was used to assess sites of double-strand DNA cleavage. Both hairpin DNAs underwent double-strand cleavage at five sites within or near the original randomized eight base region. For DNA 12, four of the five double-strand cleavages involved independent single-strand cleavage reactions; DNA 13 underwent double-strand DNA cleavage by independent single-strand cleavages at all five sites. DNA 14, which bound Fe·BLM poorly, was converted to a strong binder (DNA 15) by insertion of the sequence 5′-ACGC/5′-GCGT. These findings reinforce the idea that tighter DNA binding by Fe·BLM leads to increased double-strand cleavage by a novel mechanism and identify a specific DNA motif conducive to strong BLM binding and cleavage.

Sequence-specific recognition of DNA minor groove by an NIR-fluorescence switch-on probe and its potential applications

“…Later, Wilson et al . reported phenyl-furan-benzimidazole diamidine-based cell permeable and sequence-selective minor groove-binding molecules ( 6 , 17 – 21 ). One of the common structural features of all these minor groove-binding ligands is the molecular curvature to complement the DNA minor groove concavity, referred to as ‘ isohelicity ,’ which enhances the binding affinity and selectivity of interaction ( 10 , 11 , 17 – 20 ).…”

Structural Features Facilitating Tumor Cell Targeting and Internalization by Bleomycin and Its Disaccharide

“…One of the most striking findings in the current study involved the metalloBLMs Cu(II)•BLM and Fe(III)•BLM, both of which have been studied extensively over a period of years. 39 , 40 As shown in Figure 4 b and c, Cu(II)•BLM-Cy5** was bound/internalized about twice as efficiently as the metal-free species (BLM–Cy5**) or the BLM disaccharide–Cy5** conjugate ( 1 ). The observed binding was clearly dependent on the BLM carbohydrate moiety, since Cu(II)•deglycoBLM–Cy5** showed minimal binding/uptake (Figure 4 b,c).…”

Zorbamycin has a different DNA sequence selectivity compared with bleomycin and analogues

“…In a series of reactions, this BLM-Fe(II) complex removes a hydrogen atom from the C4′ deoxyribose sugar to produce a 4′-radical intermediate which in the presence of O 2 leads to the generation of a gapped DNA site with 5′-phosphate and 3′-phosphoglycolate ends. This gapped DNA may undergo further interaction with BLM to form a double-strand DNA break [7, 8]. Alternatively, oxidation of the 4′-radical intermediate followed by addition of H 2 O generates a 4′-oxidized abasic site [9].…”

“…BLM cleavage of hairpin DNAs has been investigated and resulted in a different spectrum of BLM binding sites [7, 8, 19, 20]. The DNA sequence specificity of BLM has been determined in human cells [21–27].…”