Nuclease activity of 1,10-phenanthroline-copper. New conjugates with low molecular weight targeting ligands.

The chemical nuclease activity of 1,10-phenanthroline-copper depends on DNA sequence because the coordination complex has affinity for DNA. In order to target this efficient nucleolytic activity, it is essential to override its inherent specificity. The minimal size of ligands capable of redirecting the specificity has been investigated. A conjugate (HOP) prepared by alkylating Hoechst dye 33258 with 5-(iodoacetamido)-1,10-phenanthroline has a greater preference for A-T-rich regions than the unsubstituted 1,10-phenanthroline-copper complex, reflecting the specificity of this A-T-specific minor-groove binder. However, since quaternizing the dye with 5-(iodoacetamido)-1,10-phenanthroline increases its affinity for DNA, the specificity of cleavage by the conjugate is less than the binding selectivity of the dye. Linking 1,10-phenanthroline with the peptide of the helix-turn-helix domain of the Trp repressor specificity results in a conjugate with greater reactivity for the operator sequence than the unsubstituted complex. The intrinsic affinity of the 1,10-phenanthroline-Cu can only be partially overridden by the conformationally unstable peptide. Attachment of 1,10-phenanthroline to a deoxyoligonucleotide complementary to a single-stranded loop of RNA successfully targets the scission of the chemical nuclease. Cleavage sites are observed not only contiguous to the site of hybridization but also at nonadjacent sequence positions. The latter set of sites must be close in space to the 5' end of the hybridized deoxyoligonucleotide.