Dinuclear ruthenium(II) complexes as potential probes for RNA bulge sites

¹ H NMR spectroscopy and molecular modelling have been used to investigate the binding of the ΔΔ- and ΛΛ-enantiomers of the dinuclear ruthenium(II) complex [{Ru(Me₂bpy)₂}₂(μ-bpm)]⁴⁺ {Me₂bpy = 4,4′-dimethyl-2,2′-bipyridine; bpm = 2,2′-bipyrimidine} to an RNA tridecanucleotide duplex containing a single-base bulge {r(CCGAGAAUUCCGG)₂]}, and the corresponding control dodecanucleotide {r(CCGGAAUUCCGG)₂}. Both enantiomers bound the control RNA sequence weakly. From upfield shifts of the metal complex H3 and H3′ protons throughout the titration of the control dodecanucleotide with ΔΔ-[{Ru(Me₂bpy)₂}₂(μ-bpm)]⁴⁺, a binding constant of 1 × 10³ M⁻¹ was determined. In NOESY spectra of the control sequence with added ΔΔ-[{Ru(Me₂bpy)₂}₂(μ-bpm)]⁴⁺, NOEs were only observed to protons from the terminal base-pair residues. No significant changes in chemical shift were observed for either the metal complex or RNA protons upon addition of the ΛΛ-enantiomer to the control dodecanucleotide. The ΔΔ-[{Ru(Me₂bpy)₂}₂(μ-bpm)]⁴⁺ complex bound the bulge-containing RNA with a significantly greater affinity (6 × 10⁴ M⁻¹) than the non-bulge control RNA duplex. Competition binding experiments indicated that the ΛΛ-isomer bound the tridecanucleotide with similar affinity to the ΔΔ-enantiomer. Addition of ΔΔ-[{Ru(Me₂bpy)₂}₂(μ-bpm)]⁴⁺ to the bulge-containing tridecanucleotide induced selective changes in chemical shift for the base H8 and sugar H1′ resonances from the adenine bulge residue, and resonances from nucleotide residues adjacent to the bulge site. Intermolecular NOEs observed in NOESY spectra of the tridecanucleotide with added ΔΔ-[{Ru(Me₂bpy)₂}₂(μ-bpm)]⁴⁺ confirmed the selective binding of the ruthenium complex at the bulge site. Preliminary binding models, consistent with the NMR data, showed that the ruthenium complex could effectively associate in the RNA minor groove at the bulge site.