Sequence Specific Binding of Beta Carboline Alkaloid Harmalol with Deoxyribonucleotides: Binding Heterogeneity, Conformational, Thermodynamic and Cytotoxic Aspects

Abstract

BackgroundBase dependent binding of the cytotoxic alkaloid harmalol to four synthetic polynucleotides, poly(dA).poly(dT), poly(dA-dT).poly(dA-dT), poly(dG).poly(dC) and poly(dG-dC).poly(dG-dC) was examined by various photophysical and calorimetric studies, and molecular docking.Methodology/Principal FindingsBinding data obtained from absorbance according to neighbor exclusion model indicated that the binding constant decreased in the order poly(dG-dC).poly(dG-dC)>poly(dA-dT).poly(dA-dT)>poly(dA).poly(dT)>poly(dG).poly(dC). The same trend was shown by the competition dialysis, change in fluorescence steady state intensity, stabilization against thermal denaturation, increase in the specific viscosity and perturbations in circular dichroism spectra. Among the polynucleotides, poly(dA).poly(dT) and poly(dG).poly(dC) showed positive cooperativity where as poly(dG-dC).poly(dG-dC) and poly(dA-dT).poly(dA-dT) showed non cooperative binding. Isothermal calorimetric data on the other hand showed enthalpy driven exothermic binding with a hydrophobic contribution to the binding Gibbs energy with poly(dG-dC).poly(dG-dC), and poly(dA-dT).poly(dA-dT) where as harmalol with poly(dA).poly(dT) showed entropy driven endothermic binding and with poly(dG).poly(dC) it was reported to be entropy driven exothermic binding. The study also tested the in vitro chemotherapeutic potential of harmalol in HeLa, MDA-MB-231, A549, and HepG2 cell line by MTT assay.Conclusions/SignificanceStudies unequivocally established that harmalol binds strongly with hetero GC polymer by mechanism of intercalation where the alkaloid resists complete overlap to the DNA base pairs inside the intercalation cavity and showed maximum cytotoxicity on HepG2 with IC50 value of 14 µM. The results contribute to the understanding of binding, specificity, energetic, cytotoxicity and docking of harmalol-DNA complexation that will guide synthetic efforts of medicinal chemists for developing better therapeutic agents.

Interaction of β-Carbolines with DNA: Spectroscopic Studies, Correlation with Biological Activity and Molecular Docking

“…The binding constants obtained for β-carboline derivatives and ctDNA in the present work corroborate some literature data for this class of compounds. 40,41 The K b values ≥ 10 5 indicate high affinity between the evaluated compounds and ctDNA. 3 The Scheme S2 presents a comparative literature reports containing similar compounds to those evaluated in this work and interactions parameters with different nucleic acids.…”

Therapeutic potential of harmaline, a novel alkaloid, against cervical cancer cells in vitro: Apoptotic induction and DNA interaction study

“…CD spectra were measured on a Jasco J815 spectropolarimeter attached to a temperature controller, temperature programmer (model PFD 425 L/15) and a PC, as described previously [21,22,23,29].…”

“…(4). Generally, cooperative binding is mediated by allosteric effects [21,24,25,[29][30][31]. Scatchard plot from the interaction of harmaline with CT DNA indicates [inset of Figure 4a] intrinsic binding affinity, K i ω (product of cooperative binding affinity and the cooperative factor) in the order of 5.60 × 10 5 /M with a stoichiometry of 4 nucleotide phosphates.…”

“…Conformational changes of CT DNA-harmaline was studied by circular dichroism (CD) [21,23,[34][35][36] [ Figure 4c]. The intrinsic CD spectra of the CT DNA-alkaloid interaction at pH 6.8 shows a regular and prominent perturbation in the 278 nm positive peak while the 242 nm negative peak show only slight changes in molar ellipticity.…”

“…Previously, harmalol, another naturally occurring beta carboline alkaloid, have been reported to have strong in-vitro cytotoxic ability [19][20][21] and also have binding potential with nucleic acids [22,23]. Hence, with reference to our previous investigation, in this study we have tried to explore the binding and cytotoxic ability of harmaline on human cervical carcinoma (HeLa) cell line in detail using different biophysical, calorimetric, and biochemical assays.…”