Exploring the comparative binding aspects of benzophenanthridine plant alkaloid chelerythrine with RNA triple and double helices: a spectroscopic and calorimetric approach

Abstract

A comparative study on the interaction of a benzophenanthridine alkaloid chelerythrine (CHL) with RNA triplex poly(U).poly(A)*poly(U) (hereafter U.A*U, .(dot) and *(asterisk) represent Watson-Crick and Hoogsteen base pairing respectively) and its parent duplex poly(A).poly(U) (A.U) was carried out by using a combination of various spectroscopic, viscometric and calorimetric techniques. The interaction was characterized by hypochromic and bathochromic effects in the absorption spectrum, the increase of thermal melting temperature, enhancement in solution viscosity, and perturbation in the circular dichroic spectrum. The binding constant calculated by using spectrophotometric data was in the order of 10(5) for both forms of RNA, but it was greater for triplex RNA (30.2 × 10(5) M(-1)) than duplex RNA (3.6 × 10(5) M(-1)). Isothermal titration calorimetric data are in good agreement with the spectrophotometric data. The data indicated stronger binding of CHL to the triplex structure of RNA compared to the native duplex structure. Thermal melting studies indicated greater stabilization of the Hoogsteen base paired third strand of the RNA triplex compared to its Watson-Crick strands. The mode of binding of CHL to both U.A*U and A.U was intercalation as revealed from fluorescence quenching, viscosity measurements and sensitization of the fluorescence experiment. Thermodynamic data obtained from isothermal calorimetric measurements revealed that association was favoured by both a negative enthalpy change and a positive entropy change. Taken together, our results suggest that chelerythrine binds and stabilizes the RNA triplex more strongly than its respective parent duplex. The results presented here may be useful for formulating effective antigene strategies involving benzophenanthridine alkaloids and the RNA triplex.

A Spectroscopic Approach towards the Comparative Binding Studies of the Antioxidizing Flavonol Myricetin with Various Single‐Stranded RNA

“…When MTN binds to a polyribonucleotide sequence, the intensity of absorption at the absorption maximum decreases as deciphered from MTN's UV spectra. This is due to the fact that the empty π*‐orbital of MTN (LUMO) is now partially filled with electrons on interaction with the π*‐orbital of the nitrogenous base pairs of RNA homo‐polymer [31] . The spectral features display distinct hypochromic effects, in presence of poly(rA) (∼30 %), poly(rU) (∼28 %) and poly(rC) (∼17.5 %) respectively at the absorption maxima without any significant shift of the absorption peak.…”

“…This is due to the fact that the empty π*-orbital of MTN (LUMO) is now partially filled with electrons on interaction with the π*orbital of the nitrogenous base pairs of RNA homo-polymer. [31] The spectral features display distinct hypochromic effects, in presence of poly(rA) (∼ 30 %), poly(rU) (∼ 28 %) and poly(rC) (∼ 17.5 %) respectively at the absorption maxima without any significant shift of the absorption peak. As observed, the maximum decrease in absorption intensity occurred in the case of poly(rA) followed by poly(rU) and lastly poly(rC).…”

Multispectroscopic and Theoretical Exploration of the Comparative Binding Aspects of Bioflavonoid Fisetin with Triple- and Double-Helical Forms of RNA

“…A similar behavior of other molecules has been reported with DNA as well as RNA triple helices. 15,55,56 Thermal Melting Study. Nucleic acids are characterized by the ability of a single molecular strand to particularly couple with a second strand or a third strand using the intrinsic pairing potential of the nucleotide bases to form a double-stranded or triple-stranded helical structure.…”

“…15 In practice, thermal stability of the triplex structure increases on binding with intercalating compounds, resulting in an enhancement in the melting temperature of the parent triplex and duplex. 56,58,59 Controlling factors 60 behind the triplex stabilization by intercalators are (1) good stacking interactions between the base triplets and the intercalator due to the planar aromatic ring structure;…”

“…A lower stoichiometry was found in the case of triplex-bound FTN than that in duplex-bound FTN, which indicates closer binding sites of the FTN−triplex complex as well as stronger association of FTN with the triple-helical form of RNA. 56 Lifetime Measurements. To exploit the excited-state dynamics of fluorophore FTN within the RNA environment, the time-resolved fluorescence decay of FTN has been recorded in the absence and presence of increasing concentrations of U.A*U and A.U.…”