Exploring the binding mode of donepezil with calf thymus DNA using spectroscopic and molecular docking methods

Donepezil (DNP) is one of approved drugs to treat Alzheimer's disease (AD). However, the potential effect of DNP on DNA is still unclear. Therefore, the interaction of DNP with calf thymus DNA (DNA) was studied in vitro using spectroscopic and molecular docking methods. Steady‐state and transient fluorescence experiments showed that there was a clear binding interaction between DNP and DNA, resulting from DNP fluorescence being quenched using DNA. DNP and DNA have one binding site between them, and the binding constant (K_b) was 0.78 × 10⁴ L·mol^?1 at 298 K. In this binding process, hydrophobic force was the main interaction force, because enthalpy change (ΔH) and entropy change (ΔS) of DNP–DNA were 67.92 kJ·mol^?1 and 302.96 J·mol^?1·K^?1, respectively. DNP bound to DNA in a groove‐binding mode, which was verified using a competition displacement study and other typical spectroscopic methods. Fourier transform infrared (FTIR) spectrum results showed that DNP interacted with guanine (G) and cytosine (C) bases of DNA. The molecular docking results further supported the results of spectroscopic experiments, and suggested that both Pi‐Sigma force and Pi‐Alkyl force were the major hydrophobic force functioning between DNP and DNA.     

Spectroscopic and molecular docking studies on the interaction of antiviral drug nevirapine with calf thymus DNA

The interaction of calf thymus DNA with nevirapine at physiological pH was studied by using absorption, circular dichroism, viscosity, differential pulse voltammetry, fluorescence techniques, salt effect studies and computational methods. The drug binds to ct-DNA in a groove binding mode, as shown by slight variation in the viscosity of ct-DNA. Furthermore, competitive fluorimetric studies with Hoechst 33258 indicate that nevirapine binds to DNA via groove binding. Moreover, the structure of nevirapine was optimized by DFT calculations and was used for the molecular docking calculations. The molecular docking results suggested that nevirapine prefers to bind on the minor groove of ct-DNA.     

Multi-spectroscopic and molecular docking studies on the interaction of neotame with calf thymus DNA

Abstract

In this paper, we have studied the in vitro binding of neotame (NTM), an artificial sweetener, with native calf thymus DNA using different methods including spectrophotometric, spectrofluorometric, competition experiment, circular dichroism (CD), and viscosimetric techniques. From the spectrophotometric studies, the binding constant (K_b) of NTM-DNA was calculated to be 2?×?10³ M^?1. The quenching of the intrinsic fluorescence of NTM in the presence of DNA at different temperatures was also used to calculate binding constants (K_b) as well as corresponding number of binding sites (n). Moreover, the obtained results indicated that the quenching mechanism involves static quenching. By comparing the competitive fluorimetric studies with Hoechst 33258, as a known groove probe, and methylene blue, as a known intercalation probe, and iodide quenching experiments it was revealed that NTM strongly binds in the grooves of the DNA helix, which was further confirmed by CD and viscosimetric studies. In addition, a molecular docking method was employed to further investigate the binding interactions between NTM and DNA, and confirm the obtained results.     

Understanding nucleosomal histone and DNA interactions: a biophysical study

Histones are associated with DNA to form nucleosome essential for chromatin structure and major nuclear processes like gene regulation and expression. Histones consist of H1, H2A, H2B and H3, H4 type proteins. In the present study, combined histones from calf thymus were complexed with ct DNA and their binding affinities were measured fluorimetrically. All the five histones were resolved on SDS page and their binding with DNA was visualized. The values of biding affinities varied with pH and salt concentration. Highest affinity (4.0?×?10⁵ M^?1) was recorded at pH 6.5 in 50 mM phosphate buffer and 1.5?×?10⁴ M^?1 in 2 M NaCl at pH 7.0. The CD spectra support the highest binding affinity with maximum conformational changes at pH 7.0. The time-resolved fluorescence data recorded two life times for histone tyrosine residues at 300 nm emission in phosphate buffer pH 6.5. These life times did not show much change upon binding with DNA in buffer as well as in 2 M NaCl. The isothermal calorimetric studies yielded thermodynamic parameters ΔG, ΔH and ΔS as ?1.6?×?10⁵ cal/mol, ?1.13?×?10³ cal/mol and ?3.80 cal/mol/deg, respectively, evidencing a spontaneous exothermic reaction. The dominant binding forces in building the nucleosome are electrostatic interactions.     

Anticancer activity,calf thymus DNA and human serum albumin binding properties of Farnesiferol C from Ferula pseudalliacea

In this study, Farnesiferol C was introduced as an anti-colon cancer agent. Its cytotoxicity was investigated on two cancer cell lines, HCT116 and CT26, and mesenchymal stem cells (MSCs) as normal cells employing MTT assay. Moreover, Farnesiferol C interactions with ct-DNA and HSA were investigated by various techniques. The IC₅₀ values of Farnesiferol C on HCT116 and CT26 cells were 42 and 46?μM, respectively, while its IC₅₀ value on MSCs cells was 92?μM, indicating that Farnesiferol C was more efficacious against cancer cell lines than normal cells. DNA competitive binding studies, viscosity and zeta potential measurements confirmed that Farnesiferol C bound to DNA through intercalation binding. HSA binding investigations revealed that there were two different binding sites for Far C on HSA with higher binding affinity in site 2 compared to site 1. Furthermore, Farnesiferol C could bind to HSA and quench its intrinsic fluorescence in a static quenching mechanism, with a distance of 2.54?nm. Competitive binding in the presence of warfarin and ibuprofen was carried out and the resulting quenching constant was strongly changed in the presence of warfarin. Consequently, Farnesiferol C most probably will be located within sub-domain IIA. In this study, molecular modeling buttressed and confirmed our laboratory results. Conclusively, we proposed that DNA is an appropriate target for Farnesiferol C. Therefore, Farnesiferol C and its semisynthetic analogues can be one of the priority innovations in research on anticancer drugs.     

The effect of dimerization on the interaction of ibuprofen drug with calf thymus DNA: Molecularmodeling and spectroscopic investigation

The interaction between the dimer structure of ibuprofen drug (D-IB) and calf thymus DNA under simulative physiological conditions was investigated with the use of Hoechst 33258 and methylene blue dye as spectral probes by the methods of UV-visible absorption, fluorescence spectroscopy, circular dichroism spectroscopy and molecular modeling study.Using the Job's plot, a single class of binding sites for theD-IB on DNA was put in evidence. The Stern–Volmer analysis of fluorescence quenching data shows the presence of both the static and dynamic quenching mechanisms. The binding constants, K_b were calculated at different temperatures, and the thermodynamic parameters ?G^°, ?H^° and ?S^° were given. The experimental results showed that D-IB molecules could bind with DNA via groove binding mode as evidenced by: I. DNA binding constant from spectrophotometric studies of the interaction of D-IB with DNA is comparable to groove binding drugs. II. Competitive fluorimetric studies with Hoechst 33258 have shown that D-IB exhibits the ability of this complex to displace with DNA-bounded Hoechst, indicating that it binds to DNA in strong competition with Hoechst for the groove binding. III. There is no significantly change in the absorption of the MB-DNA system upon adding the D-IB, indicates that MB molecules are not released from the DNA helix after addition of the D-IB and are indicative of a non-intercalative mode of binding. IV. Small changes in DNA viscosity in the presence of D-IB, indicating weak link to DNA, which is consistent with DNA groove binding. As well as, induced CD spectral changes, and the docking results revealed that groove mechanism is followed by D-IB to bind with DNA.     

Interaction of procarbazine with calf thymus DNA—a biophysical and molecular docking study

Interaction of procarbazine (PCZ) with calf thymus DNA was studied using biophysical and molecular docking studies. Procarbazine was to interact with DNA with a binding constant of 6.52 × 10³ M⁻¹ as calculated using ultraviolet‐visible spectroscopy. To find out the binding mode, molecular docking was performed that predicted PCZ to interact with DNA through groove binding mode with binding affinity of −6.7 kcal/mole. To confirm the groove binding nature, different experiments were performed. Dye displacement assays confirmed the non‐intercalative binding mode. Procarbazine displaced Hoechst dye from the minor groove of DNA while it was unable to displace intercalating dyes. There was no increase in the viscosity of DNA solution in presence of PCZ. Also, negligible change in the secondary structure of DNA was observed in presence of PCZ as evident by circular dichroism spectra. Procarbazine caused decrease in the melting temperature of DNA possibly because of decrease in the stability of DNA caused by groove binding interaction of PCZ with DNA.     

Two homologous domains of similar structure but different stability in the yeast linker histone, Hho1p

The Saccharomyces cerevisiae homologue of the linker histone H1, Hho1p, has two domains that are similar in sequence to the globular domain of H1 (and variants such as H5). It is an open question whether both domains are functional and whether they play similar structural roles. Preliminary structural studies showed that the two isolated domains, GI and GII, differ significantly in stability. In 10 mM sodium phosphate (pH 7), the GI domain, like the globular domains of H1 and H5, GH1 and GH5, was stably folded, whereas GII was largely unstructured. However, at high concentrations of large tetrahedral anions (phosphate, sulphate, perchlorate), which might mimic the charge-screening effects of DNA phosphate groups, GII was folded. In view of the potential significance of these observations in relation to the role of Hho1p, we have now determined the structures of its GI and GII domains by NMR spectroscopy under conditions in which GII (like GI) is folded. The backbone r.m.s.d. over the ordered residues is 0.43 A for GI and 0.97 A for GII. Both structures show the "winged-helix" fold typical of GH1 and GH5 and are very similar to each other, with an r.m.s.d. over the structured regions of 1.3 A, although there are distinct differences. The potential for GII to adopt a structure similar to that of GI when Hho1p is bound to chromatin in vivo suggests that both globular domains might be functional. Whether Hho1p performs a structural role by bridging two nucleosomes remains to be determined.     

Voltammetric behavior of complexation of salbutamol with calf thymus DNA and its analytical application

The interaction of salbutamol (Sal), an animal growth promoter, with DNA was investigated by differential pulse voltammetry (DPV), cyclic voltammetry (CV), and fluorescence spectroscopy. An irreversible reduction was observed from the cyclic voltammograms, and the reaction mechanism involved a one-electron change irreversible oxidation. In the presence of DNA, the DPV peak current decreased and the Sal peak shifted to higher potentials, indicating that Sal interacted with DNA to form an intercalation Sal–DNA complex. In addition, reaction binding parameters were extracted from the DPV data with the use of the multivariate curve resolution–alternating least squares (MCR–ALS) method; the binding constant and ratio were found to be (2.0 ± 0.5) × 10⁵ M⁻¹ and 1:1, respectively. Quantitative voltammetric analysis of Sal was performed in the concentration range of 3.02 × 10⁻⁶ to 1.23 × 10⁻⁴ mol L⁻¹, and it was found that the detection limit was 5.11 × 10⁻⁷ mol L⁻¹ in the presence of 1.00 × 10⁻⁶ mol L⁻¹ DNA. The method was applied for the determination of Sal in spiked urine and human serum samples, and the calibration was successfully verified.     

In vitro interaction between calf thymus DNA and Escherichia coli LPS in the presence of divalent cation Ca2+

With increasing addition of Escherichia coli LPS to calf thymus DNA, both dissolved in CaCl2, absorption maxima of DNA at 260 nm decreased gradually with the appearance of isosbastic points at both ends of spectra, which implied some binding between DNA and LPS. Hill plot of absorbance data showed that the binding interaction was positive cooperative in nature. For any fixed concentration of DNA and LPS, extent of interaction increased as concentration of CaCl2 was raised from 1.0 to 100 mM, signifying the electrostatic nature of the interaction, mediated through Ca2+ ion. Stepwise addition of EDTA, a chelating agent for divalent cations, to DNA-LPS bound complex gradually reversed the spectral shift with increase in absorbance at 260 nm, which implied opening up of the complex, that is, reversible nature of the interaction. Circular dichroism spectral changes of DNA by the addition of LPS indicated partial transition of DNA from B to A form. Isothermal titration calorimetric (ITC) study showed that the DNA-LPS binding was an exothermic and enthalpy-driven phenomenon. Moreover, in the presence of 100 mM CaCl2, binding constant of the interaction was found to be 2.6 x 10(4) M(-1) and 3.1 x 10(4) M(-1) from the analysis of Hill plot and ITC result, respectively. DNA-melting study showed that the LPS binding had increased the melting temperature of DNA, indicating more stabilization of DNA double helix. The binding of LPS to DNA made the complex resistant to digestion with endonucleases EcoRI and DNase I.     

Multispectroscopic studies on the interaction of a copper(ii) complex of ibuprofen drug with calf thymus DNA

The interaction of copper(II)–ibuprofenato complex with calf thymus DNA (ct-DNA) has been explored following, UV-visible spectrophotometry, fluorescence measurement, dynamic viscosity measurements, and circular dichroism spectroscopy. In spectrophotometric studies of ct-DNA it was found that [Cu(ibp)₂]₂ can form a complex with double-helical DNA. The association constant of [Cu(ibp)₂]₂ with DNA from UV-Vis study was found to be 6.19 × 10⁴ L mol^?1. The values of K_f from fluorescence measurement clearly underscore the high affinity of [Cu(ibp)₂]₂ to DNA. The experimental results showed that the conformational changes in DNA helix induced by [Cu(ibp)₂]₂ are the reason for the fluorescence quenching of the DNA-Hoechst system. In addition, the fluorescence emission spectra of intercalated methylene blue (MB) with increasing concentrations of [Cu(ibp)₂]₂ represented a significant increase of MB intensity as to release MB from MB-DNA system. The results of circular dichroism (CD) suggested that copper(II)–ibuprofenato complex can change the conformation of DNA. In addition, the results of viscosity measurements suggest that copper(II)–ibuprofenato complex may bind with non-classical intercalative mode. From spectroscopic and hydrodynamic studies, it has been found that [Cu(ibp)₂]₂ interacts with DNA by partial intercalation mode which contains intercalation and groove properties.     

Biological evaluation,molecular docking and DNA interaction studies of coordination compounds gleaned from a pyrazolone incorporated ligand

Abstract

In this work, we have synthesized a few novel mononuclear complexes of Cu(II), Co(II), Ni(II) and Zn(II) using a pyrazolone-derived Schiff base ligand. They were characterized by spectroscopic and analytical methods. The elemental analyses, UV-Vis, magnetic moment values and molar conductance of the complexes reveal that the complexes adopt an octahedral arrangement around the central metal ions. The interaction of complexes with CT-DNA was studied by absorption spectral titration and viscosity measurements. The observed data show that the complexes bind with CT-DNA via an intercalation mode. Efficient pUC18 DNA cleavage ability of the synthesized compounds was explored by gel electrophoresis. The antimicrobial activity of these compounds against a set of bacterial and fungal strains reveals that the complexes exhibit better activity than the free ligand. Moreover, all the complexes were evaluated against two cancer (HeLa and HepG2) and one normal (NHDF) cell lines. The data were compared with cisplatin. Anti–inflammatory activity has been experimentally validated which proves that theoretical predictions concur with the experimental results. In addition, molecular docking studies have been performed to consider the nature of binding mode and binding affinity of these compounds with DNA (1BNA) and protein (3hb5). These studies reveal that the mode of binding is intercalation and the complexes have higher binding energy scores than the free ligand.     

Dissecting the binding mechanism of the linker histone in live cells: an integrated FRAP analysis

The linker histone H1 has a fundamental role in DNA compaction. Although models for H1 binding generally involve the H1 C‐terminal tail and sites S1 and S2 within the H1 globular domain, there is debate about the importance of these binding regions and almost nothing is known about how they work together. Using a novel fluorescence recovery after photobleaching (FRAP) procedure, we have measured the affinities of these regions individually, in pairs, and in the full molecule to demonstrate for the first time that binding among several combinations is cooperative in live cells. Our analysis reveals two preferred H1 binding pathways and we find evidence for a novel conformational change required by both. These results paint a complex, highly dynamic picture of H1–chromatin binding, with a significant fraction of H1 molecules only partially bound in metastable states that can be readily competed against. We anticipate the methods we have developed here will be broadly applicable, particularly for deciphering the binding kinetics of other nuclear proteins that, similar to H1, interact with and modify chromatin.     

Experimental and computational studies on the effects of valganciclovir as an antiviral drug on calf thymus DNA

DNA-binding properties of an antiviral drug, valganciclovir (valcyte) was studied by using emission, absorption, circular dichroism, viscosity, differential pulse voltammetry, fluorescence techniques, and computational studies. The drug bound to calf thymus DNA (ct-DNA) in a groove-binding mode. The calculated binding constant of UV-vis, K_a, is comparable to groove-binding drugs. Competitive fluorimetric studies with Hoechst 33258 showed that valcyte could displace the DNA-bound Hoechst 33258. The drug could not displace intercalated methylene blue from DNA double helix. Furthermore, the induced detectable changes in the CD spectrum of ct-DNA as well as changes in its viscosity confirm the groove-binding mode. In addition, an integrated molecular docking was employed to further investigate the binding interactions between valcyte and calf thymus DNA.     

Thermodynamic study of interaction of TSPP,CoTsPc, and FeTsPc with calf thymus DNA

The interaction of two water soluble phthalocyanines, cobalt(II) 4,4′,4″,4‴-tetrasulfo-phthalocyanine (CoTsPc) and iron(II) 4,4′,4″,4‴-tetrasulfo-phthalocyanine (FeTsPc), and one water soluble porphyrin, tetra sodium mesotetrakis(p-sulfophenyl)porphyrin (TSPP), with calf thymus DNA has been studied by UV-Vis spectroscopy at five different temperatures (20, 25, 30, 35, and 40°C). The optical absorption spectra of these materials were analyzed to obtain binding constants and stoichiometries using SQUAD software. The results show that the best fitting corresponds to a 1: 1 complex model between a base pair of DNA and these materials. All of the studied porphyrin and phthalocyanines showed strong electrolyte effect, and increasing NaCl concentration induced self-aggregation of these materials. Published in Russian in Biokhimiya, 2007, Vol. 72, No. 6, pp. 799–804.     

Comparative study of two cephalosporin antibiotics binding to calf thymus DNA by multispectroscopy,electrochemistry, and molecular docking

The over‐use of antibiotics has caused a number of problems such as contamination of antibiotic residues and virus resistance, and therefore has attracted global attention. In this study, spectroscopic techniques and molecular docking were employed to predict conformational changes and binding interaction between two cephalosporins (cefaclor and cefixime) and calf thymus DNA (ctDNA). Fluorescence and UV–vis spectra suggested that static quenching was predominant and cephalosporin bound to the groove region of ctDNA. Binding parameters calculated by the Stern–Volmer and Scatchard equations showed that cephalosporin bound to ctDNA with a binding affinity in the order of 10³ L mol^?1. Thermodynamic parameters further indicated that the reaction was a spontaneous process driven by enthalpy and entropy, and that the main binding force was an electrostatic force. The effects of iodide, denaturant, thermal denaturation and pH on a cephalosporin–Hoechst–DNA complex were also studied, and the results confirmed that cephalosporin bound to the groove area of DNA. Finally, these results were further confirmed by molecular docking and electrochemical studies.     

Deciphering the intercalative binding modes of benzoyl peroxide with calf thymus DNA

The binding of benzoyl peroxide (BPO), a flour brightener, with calf thymus DNA (ctDNA) was predicted by molecular simulation, and this were confirmed using multi‐spectroscopic techniques and a chemometrics algorithm. The molecular docking result showed that BPO could insert into the base pairs of ctDNA, and the adenine bases were the preferential binding sites which were validated by the analysis of Fourier transform infrared spectra. The mode of binding of BPO with ctDNA was an intercalation as supported by the results from ctDNA melting and viscosity measurements, iodide quenching effects and competitive binding investigations. The circular dichroism and DNA cleavage assays indicated that BPO induced a conformational change from B‐like DNA structure towards to A‐like form, but did not lead to significant damage in the DNA. The complexation was driven mainly by hydrogen bonds and hydrophobic interactions. Moreover, the ultraviolet–visible (UV–vis) spectroscopic data matrix was resolved by a multivariate curve resolution–alternating least–squares algorithm. The equilibrium concentration profiles for the components (BPO, ctDNA and BPO–ctDNA complex) were extracted from the highly overlapping composite response to quantitatively monitor the BPO–ctDNA interaction. This study has provided insights into the mechanism of the interaction of BPO with ctDNA and potential hazards of the food additive.     

One‐pot synthesis and characterization CdTe:Zn2+ quantum dots and its molecular interaction with calf thymus DNA

Tremendous research efforts have been dedicated to fabricating high‐quality Zn‐doped CdTe quantum dots (QDs) for any potential biomedical applications. In particular, the correlation of issues regarding how QDs interact with DNA is of greatest importance. Herein, a pH‐responsive study of the interactions between CdTe:Zn²⁺ quantum dots with 4 different sizes and calf thymus DNA (ctDNA) was conducted using multispectroscopic techniques and electrochemical investigation. Fluorescence studies revealed that this interaction process is predominantly a static process and groove binding was the main binding mode for CdTe:Zn²⁺ QDs to ctDNA. The calculated negative values of enthalpy (?45.06 kJ mol^?1) and entropy (?133.62 J mol^?1 K^?1) with temperature changes indicated that the hydrogen bonds and van der Waals interactions played major roles in the reaction. Furthermore, circular dichroism spectroscopy and Fourier transform infrared spectrometry analyses indicate that the normal conformation of ctDNA is discombobulated by CdTe:Zn²⁺ QDs. In addition, the electrochemical behavior of the affinity of CdTe:Zn²⁺ QDs for ctDNA agreed well with the results obtained from fluorescence experiments. This study might be meaningful for understanding the molecular binding mechanism of QDs for DNA and provides a basis for QD‐labeled systems.     

Synthesis of Zn(II)-cloxacillin sodium complex and study of its interaction with calf thymus DNA

In this paper, a solid complex of cloxacillin sodium (CS) with Zn(II) was prepared by coprecipitation and characterized by UV, fluorescence, IR, and thermal spectra. Furthermore, the nature of the complex has been studied by ¹H-NMR and ¹³C-NMR spectroscopy. The influence of Zn(II) on the combination of CS and calf thymus DNA (CT DNA) was studied using fluorescence spectrophotometry, and the formation of binary CS-Zn(II) and CS-CT DNA complexes and ternary CS-Zn(II)-CT DNA complex was studied. The results show that the fluorescence intensity of CS can be quenched in the presence of Zn(II) or DNA. In the presence of Zn(II), the fluorescence quenching action of DNA on CS was strongly enhanced. Based on the fluorescence intensity, the formation constants of CS-Zn(II) and CS-CT DNA complexes were calculated, and the mechanism of interaction between CS, Zn(II), and DNA is discussed. Published in Russian in Biokhimiya, 2007, Vol. 72, No. 2, pp. 184–193.