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The interaction of two natural protoberberine alkaloids berberine and palmatine and the synthetic derivative coralyne with the RNA triplex poly(U).poly(A)(*)poly(U) was studied using various biophysical and calorimetric techniques. All the three alkaloids bind noncooperatively to the triplex. The affinity of berberine and palmatine was in the order of 10(5) M(-1), while that of coralyne was one order higher as inferred from spectroscopic studies. The alkaloids stabilized the Hoogsteen base-paired third strand of the triplex without affecting the stability of the duplex. Fluorescence quenching and viscosity studies gave convincing evidence for the partial intercalation of berberine and palmatine and a true intercalative binding of coralyne to the triplex. This was further supported from the significant polarization of the emission spectra of the complex and the energy transfer from the base triplets to the alkaloids. Circular dichroic studies suggested that the conformation of the triplex was perturbed significantly by the binding of the alkaloids, being more by coralyne compared to berberine and palmatine and also evidenced by the generation of strong induced optical activity in the bound coralyne molecules. Isothermal titration calorimetric studies revealed that the binding to the triplex was favored by a predominantly large negative enthalpy change (DeltaH degrees = -5.42 kcal/mol) with small favorable entropy contribution (TDeltaS degrees = 2.02 kcal/mol) in berberine, favored by almost equal negative enthalpy (DeltaH degrees = -3.93 kcal/mol) and entropy changes (TDeltaS degrees = 3.89 kcal/mol) in palmatine and driven by predominant entropy contributions (DeltaH degrees = -1.84 and TDeltaS degrees = 7.44 kcal/mol) in coralyne. These results advance our knowledge on the binding of small molecule isoquinoline alkaloids that are specific binders of RNA structures, particularly triplexes.

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Design and synthesis of new small molecules binding to double-stranded RNA necessitate complete understanding of the molecular aspects of the binding of many existing molecules. Toward this goal, in this work we evaluated the biophysical aspects of the interaction of a DNA intercalator (proflavine) and a minor groove binder (hoechst 33258) with two polymorphic forms of polyCG, namely, the right-handed Watson-Crick base paired A-form and the left-handed Hoogsteen base paired H(L)-form, by absorption, fluorescence, and viscometry experiments. The energetics of the interaction of these molecules with the RNA structures has also been elucidated by isothermal titration calorimetry (ITC). Results suggest that proflavine strongly intercalates in both forms of polyCG, whereas hoechst shows mainly groove-binding modes. The binding of both drugs to both forms of RNA resulted in significant conformational change to the RNA structure with the bound molecules being placed in the chiral RNA helix. ITC profiles for both proflavine and hoechst show two binding sites. Binding of proflavine to both forms of RNA is endothermic and entropy driven in the first site and exothermic and enthalpy driven in the second site, whereas hoechst binding to both forms of RNA is exothermic and enthalpy driven in the first site and endothermic and entropy driven in the second site. This study suggests that the binding affinity characteristics and energetics of interaction of these DNA binding molecules with the RNA conformations are significantly different and may serve as data for future development of effective structure-selective RNA-based drugs.

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