Biophysical Characterization of the Strong Stabilization of the RNA Triplex poly(U)•poly(A)*poly(U) by 9-O-(ω-amino) Alkyl Ether Berberine Analogs

Background: Binding of two 9-O-(v-amino) alkyl ether berberine analogs BC1 and BC2 to the RNA triplex poly(U)Npoly(A)*poly(U) was studied by various biophysical techniques. Methodology/Principal Findings: Berberine analogs bind to the RNA triplex non-cooperatively. The affinity of binding was remarkably high by about 5 and 15 times, respectively, for BC1 and BC2 compared to berberine. The site size for the binding was around 4.3 for all. Based on ferrocyanide quenching, fluorescence polarization, quantum yield values and viscosity results a strong intercalative binding of BC1 and BC2 to the RNA triplex has been demonstrated. BC1 and BC2 stabilized the Hoogsteen base paired third strand by about 18.1 and 20.5uC compared to a 17.5uC stabilization by berberine. The binding was entropy driven compared to the enthalpy driven binding of berbeine, most likely due to additional contacts within the grooves of the triplex and disruption of the water structure by the alkyl side chain. Conclusions/Significance: Remarkably higher binding affinity and stabilization effect of the RNA triplex by the amino alkyl berberine analogs was achieved compared to berberine. The length of the alkyl side chain influence in the triplex stabilization phenomena

binding to natural organic matter: competition, heterogeneity, stoichiometry and thermodynamic consistency

The general principles of cation binding to humic materials are discussed. Important aspects that need to be included in general purpose speciation models are: the extreme binding heterogeneity of natural humic materials, the variable stoichiometry of binding (monodentate, bidentate and tridentate), the competition between specifically-bound ions, especially protons and metal ions, and electrostatic effects which give rise to ionic strength effects and the non-specific binding of counterions. The NICCA–Donnan model is a semi-empirical model that addresses these issues. It is similar to the previously published NICA–Donnan model except that it introduces an additional degree of scaling that ensures thermodynamic consistency and allows for variable stoichiometry of binding. It implicitly accounts for the large degree of chemical heterogeneity of humic particles. The NICCA (consistent NICA) model also recognizes that the affinity distributions are ion specific and are not fully correlated. The model requires no assumptions to be made about the geometry of the humic particles, but the Donnan submodel does allow for shrinking and swelling. Important model parameters such as the site density and median binding constants are not dependent on pH, metal ion concentration, ionic strength, etc. Data are analysed for H+, Ca2+, Cd2+, Cu2+, Pb2+ and Al3+ binding to a single purified peat humic acid. The NICCA–Donnan model captures the non-linearity of the observed isotherms even at very low free metal ion concentrations. After fitting the model to datasets containing only the proton and one metal ion, the model was able to predict Cd2+–Ca2+, Cu2+–Ca2+ and Pb2+–Al3+ competition reasonably well. It also gave satisfactory predictions of the H+/Mz+ molar exchange ratios. These ratios varied strongly with metal ion: Ca2+ (0.2–0.5); Cd2+ (0.5–1.0); Pb2+ (1.1–1.2); Cu2+ (1.2–1.7) and Al3+ (2.1–2.7), and also to a varying degree with pH and free metal ion concentration