Investigation of the abasic sites induced by hydrogen peroxide and methyl methanesulfonate in calf thymus DNA and BEAS-2B cells.

The primary goals of this study were to investigate the formation of abasic sites (AP sites) induced by methyl methanesulfonate (MMS) and hydrogen peroxide (H2O2), and to characterize specific types of these pro-mutagenic DNA lesions in calf thymus DNA (CT-DNA), and BEAS-2B human lung normal cell line. Furthermore, these profiles were compared with those observed in leukocytes derived from healthy controls (HC), breast cancer patients (BCP) before treatment, and 5-year survivors. Results indicated that both H2O2 and MMS induced the concentration- and time-dependent formation of AP sites in CT-DNA. To characterize the specific types of AP sites induced by H2O2 or MMS, we performed AP site cleavage assay using putrescine, T7 exonuclease (T7 Exo), and exonuclease III (Exo III). Results showed that the AP sites induced by H2O2 in CT-DNA were predominantly 5'-and 3'-nicked AP sites and no intact AP sites were detected. By contrast, the majority of AP sites generated by MMS in CT-DNA are not excisable and are classified as residual and intact AP sites. Similar approaches were performed in human BEAS-2B cells and comparable observations were confirmed in the cell-based model. Further investigation indicated that the profile of the AP sites observed in Taiwanese HC is identical to that of BEAS-2B cells treated with H2O2 whereas the pattern of AP sites detected in BCP is similar to that of CT-DNA exposed to H2O2, suggesting that these AP sites were produced primarily through reactive oxygen species (ROS) generation. More than 70 % of the AP sites in leukocytes derived from BCP were 5'-nicked and residual AP sites. Furthermore, the characteristics of the AP sites detected in 5-year survivors are comparable with the ones in HC by using putrescine cleavage assay. Overall, we speculate that deficiency in the DNA repair cascade may play a role in mediating the formation of specific types of AP sites detected in BCP.

Substituent effect on the chemical and biological properties of diisatin dihydrazone Schiff bases: DFT and docking studies.

According to the considered role of lipophilicity-hydrophobicity on organic Schiff base hydrazones, different substituents of phenyl, ethyl, and methyl groups were inserted in the synthetic strategy of diisatin dihydrazones (L1-4). The biochemical enhancement was evaluated depending on their inhibitive potential of the growth power of three human tumor cells, fungi, and bacteria. The biochemical assays assigned the effected role of different substituents of phenyl, ethyl, and methyl groups on the effectiveness of their diisatin dihydrazone reagents. The interacting modes with calf thymus DNA (i.e. Ct-DNA) were studied via viscometric and spectrophotometric titration. The organo-reagent L1 with the oxalic derivative assigned a performed inhibitive action for the examined microbes and the human tumor cell lines growing up over the terephthalic (L4) > malonic (L2) > succinic (L3) ones. From Kb = binding constant, and ∆Gb≠ = Gibb's free energy values, the binding of interaction within Ct-DNA was evaluated for all compounds (L1-4), in which L1, L3, and L4 assigned the highest reactivity referring to the covalent/non-covalent modes of interaction, as given for (L1-4), 14.32, 13.28, 10.87, and 12.41 × 107 mol-1 dm3, and -45.17, -43.24, -43.75, and -44.05 kJ mol-1, respectively. DFT and docking studies were achieved to support the current work.

Nucleic acid binding affinity and antioxidant activity of N-m-Tolyl-4-Chlorophenoxyacetohydroxamicacid.

Hydroxamic acids represent a group of weak organic acids, both naturally occurring and synthetically derived, characterized by the general formula RC(= O)N(R'OH). In this study, we investigated the binding behavior of N-m-tolyl-4-chlorophenoxyaceto hydroxamic acid with calf thymus DNA (ct-DNA) and torula yeast RNA (t-RNA) through a combination of techniques including UV-visible spectroscopy, fluorescence emission analysis, viscometry, and computational simulations using AutoDock4 software. Our findings reveal that the mode of binding between the compound and the nucleic acids is consistent with intercalation. Competitive binding experiments demonstrated that the complex competes effectively with ethidium bromide (EB) for binding to ct-DNA/t-RNA, displacing EB from its binding sites. Additionally, the introduction of the compound into the DNA-EB system resulted in a quenching of fluorescence emission peaks. Analysis of absorption spectra indicated a red shift and hypochromic shift when the compound interacted with DNA, further supporting the intercalative binding mode. The calculated binding constant (Kb) value for the compound is 6.62 × 104 M-1 and 5.40 × 103 M-1 indicating a strong interaction with ct-DNA and t-RNA respectively. We determined the Stern-Volmer constants for ct-DNA and t-RNA as 9.96 × 104 M-1 and 8.13 × 105 M-1, respectively. The binding free energy values for ct-DNA/t-RNA were calculated to be - 3.741 × 107 and - 5.425 × 108 kcal/mol, respectively. Viscometric studies corroborated the UV results, showing a continuous increase in relative viscosity of ct-DNA/t-RNA solutions with the addition of the optimal hydroxamic acid concentration. Furthermore, we assessed the antioxidant activity of the compound using DPPH-radical scavenging and ß-carotene linoleic acid assays. Gel electrophoresis results demonstrated the compound's remarkable efficacy in preventing DNA damage. Collectively, all experimental evidence supports the conclusion that N-m-tolyl-4-chlorophenoxyaceto hydroxamic acid binds to ct-DNA/t-RNA through an intercalative mechanism, which is consistent with our molecular docking simulations.

Investigation of antileishmanial, antioxidant activities, CT-DNA interaction and DFT study of novel cobalt(II) complexes derived from mesogenic aromatic amino acids based Schiff base ligands.

In the present work, new Co(II) complexes were synthesized from mesogenic aromatic amino acids based Schiff base ligands, HL1 [Methyl 2-((2-hydroxy-4-(tetradecyloxy)benzylidene)amino)-3-phenylpropanoate] and HL2 [Methyl 2-((2-hydroxy-4-(tetradecyloxy)benzylidene)amino)-3-(1H-indol-2-yl)propanoate]. The compounds were thoroughly characterised using different elemental, thermogravimetric and spectroscopic studies. The in-vitro antileishmanial efficacy of the compounds against Leishmania donovani was evaluated by MTT assay and the antioxidant activity was performed by Mensor's method. The cell viability percentage and IC50 values for both the antileishmanial and antioxidant studies revealed that the cobalt(II) complexes are comparable to the standard, amphotericin B and ascorbic acid, respectively, signifying the potential applications of the biogenic compounds. The CT-DNA interaction experiments study using photophysical techniques indicated that the cobalt(II) complexes exhibited pronounced interactions as compared to the parent ligand. The parent ligands were found to possess mesogenicity as evidenced from the polarizing optical microscope (POM) and differential scanning calorimetry (DSC). The optical band gap of the compounds, as estimated from the Tauc plot of the UV-Vis spectra, lies within the domain of optoelectronic material properties, which was further supported through Density Functional Theory (DFT) study. Moreover, DFT methods have been used to explore the ground state geometry and DFT based reactivity descriptors of the two synthesised ligands, HL1 and HL2 along with their corresponding Co(II) complexes, Co(L1)2 and Co(L2)2. Reactivity descriptors obtained from Conceptual Density Functional Theory (CDFT) analysis reveal that Co(L1)2 is the most stable and Co(L2)2 is the most electrophilic.

Analysis of the effect of baseline detection and early clearance of ct-DNA, on survival outcomes among patients with advanced EGFR-mutant non-small cell lung cancer.

BACKGROUND: To determine if circulating tumor DNA (ct-DNA) dynamics of epidermal growth factor receptor (EGFR) mutation in plasma can identify a subset of patients with EGFR-mutant (EGFR- m) non-small cell lung cancer (NSCLC) with inferior survival outcomes, we analyzed and compared survival outcomes among patients with and without baseline presence and early clearance of EGFR ct-DNA in plasma. MATERIAL AND METHODS: For 66 patients newly dia-gnosed with EGFR- m NSCLC, plasma samples were collected at baseline and 1st response assessment at 12-24 weeks for extraction of ct-DNA. Estimation of ct-DNA (EGFR exons 18, 19, 20 and 21) was done using droplet digital polymerase chain reaction (dd-PCR) on the QX200 ddPCR system (BioRad, USA). Patients with detectable EGFR ct-DNA at baseline (sample 1), with either undetectable or persistent detectable ct-DNA in sample 2 were classified as clearers and non-clearers, respectively. RESULTS: Fifty-three patients received 1st/ 2nd generation EGFR tyrosine kinase inhibitors (TKIs) and 13 received either 3rd generation TKI (osimertinib) or chemotherapy plus gefitinib. The baseline ct-DNA-positive group had more patients with extra thoracic disease (60.4 vs. 48.5%). For the entire cohort, there was no difference in median progression-free survival (PFS) among baseline ct-DNA-negative (13.57 months) vs. ct-DNA-positive patients (12.32 months) (HR 0.74). There was a significant improvement of PFS among early ct-DNA clearers vs. non-clearers (12.32 vs. 9.92 months; HR 0.57). For those treated with 1st/ 2nd generation EGFR TKIs, this improvement in median PFS among early ct-DNA clearers vs. non-clearers was more apparent (11.76 vs. 6.8 months; HR 0.34). CONCLUSIONS: Baseline detection of the presence of ct-DNA of EGFR mutation in plasma was not predictive of first-line PFS, but is associated with extra thoracic disease. Patients with EGFR mutation and persistence of ct-DNA at first follow-up have worse PFS and overall survival (OS) in comparison to those clearing the same in plasma, especially among those treated with 1st/ 2nd generation EGFR TKIs.

Evolutionary Fate of the Opine Synthesis Genes in the Arachis L. Genomes.

Naturally transgenic plants are plants that have undergone Agrobacterium-mediated transformation under natural conditions without human involvement. Among Arachis hypogaea L., A. duranensis Krapov. & W.C. Greg, A. ipaensis Krapov. & W.C. Greg, A. monticola Krapov. & Rigoni, and A. stenosperma Krapov. & W.C. Greg are known to contain sequences derived from the T-DNA of "Agrobacterium". In the present study, using molecular genetics and bioinformatic methods, we characterized natural transgenes in 18 new species from six sections of the genus Arachis. We found that small fragments of genes for enzymes of the agropine synthesis pathway were preserved only in some of the studied samples and were lost in the majority of the species during evolution. At the same time, genes, similar to cucumopine synthases (cus-like), remained intact in almost all of the investigated species. In cultivated peanuts, they are expressed in a tissue-specific manner. We demonstrated the intraspecific variability of the structure and expression of the cus-like gene in cultivated peanuts. The described diversity of gene sequences horizontally transferred from Agrobacterium to plants helps to shed light on the phylogeny of species of the genus Arachis and track possible hybridization events. Data on the ability of certain species to hybridize are useful for planning breeding schemes aimed at transferring valuable traits from wild species into cultivated peanuts.

Synthesis, characterisation, and in vitro antiparasitic activity of new flavanoidal tetrazinan-6'-ones and their binding study with calf thymus DNA using molecular modelling and spectroscopic techniques.

With the developing resistance to traditional antiparasitic medications, the purpose of this study was to efficiently develop a series of six noble flavanoidal tetrazinane-6'-one derivatives by a one-pot reaction pathway. FT-IR, 1HNMR, 13CNMR, and Mass spectra were employed for the structural elucidation of the synthesized compounds (7-12). Clinostomum complanatum, a parasite infection model that has been well-established, demonstrated that all the synthesized compounds are potent antiparasitic agents. DNA is the main target for various medicinal compounds. As a result, thestudy of how small molecules attach to DNA has received a lot of attention. In the present study, we have performed various biophysical techniques to determine the mode of binding of synthesized compounds (7-12) with calf thymus DNA (ct-DNA). It was observed from the UV-visible absorbance and fluorescence spectra that all synthesized compounds (7-12) form complexes with the ct-DNA. The value of binding constant (Kb) was obtained to be in the range of 4.36---24.50 × 103 M - 1 at 298 K. Competitive displacement assay with ethidium bromide (EB), CD spectral analysis, viscosity measurements, and in silico molecular docking confirmed that ligands (7-12) incorporate with ct-DNA through groove binding only. Molecular docking studies were performed for all synthesized compounds with the calf thymus DNA and it was found that all the newly synthesized compounds strongly bind with the chain B of DNA in the minor groove with the value of binding energy in the range of -8.54 to -9.04 kcal per mole and several hydrogen bonding interactions.

Organoruthenium metallocycle induced mutation in gld-1 tumor suppression gene in JK1466 strain and appreciable lifespan expansion.

Four Ru(II) complexes (A2-A5) were synthesized from the reaction of coumarin Schiff base ligands (7da2-tsc, 7da3-mtsc, 7da4-etsc and 7da5-ptsc) with [RuHCl(CO)(PPh3)3]. The compounds were characterized by FT-IR, UV-Vis, 1H, 13C and 31P NMR, mass spectrometry and crystallographic analysis. Calf Thymus DNA (CT-DNA) binding studies revealed the intercalative mode of binding of the complexes with DNA. The results of Bovine serum albumin (BSA) binding studies established the interaction between BSA followed static quenching mechanism. The cytotoxic effects of the complexes and the ligands were evaluated against breast (MCF-7 and MDA-MB-231) and lung carcinoma cell lines (A549 and NCI-H460) using MTT assay. Complex A4 demonstrated potent cytotoxic effects on both breast and lung cancer cells. Furthermore, morphological observations and FACS analysis showed the decrease in cell density by complex A4 by induced morphological changes and apoptotic body formation and cell death in both breast and lung cancer cells. Moreover, the invertebrate model Caenorhabditis elegans was employed to assess the in vivo anticancer activity of compound A4. The findings indicated that the treatment with A4 reduced tumor development and significantly extended organismal lifespan by 64 % in the tumoral strain JK1466 without adversely affecting essential physiological functions of the worm. Additionally, A4 demonstrated an upregulation of two crucial antioxidant defense genes. Overall, these results suggested that the compound A4 can be a potential candidate with novel chemotherapeutic applications.

Peptidic Compound as DNA Binding Agent: In Silico Fragment-based Design, Machine Learning, Molecular Modeling, Synthesis, and DNA Binding Evaluation.

BACKGROUND: Cancer remains a global burden, with increasing mortality rates. Current cancer treatments involve controlling the transcription of malignant DNA genes, either directly or indirectly. DNA exhibits various structural forms, including the G-quadruplex (G4), a secondary structure in guanine-rich regions. G4 plays a crucial role in cellular processes by regulating gene expression and telomerase function. Researchers have recently identified G4-stabilizing binding agents as promising anti-cancer compounds. Additionally, peptides have emerged as effective anticancer pharmaceuticals due to their ability to form multiple hydrogen bonds, electrostatic interactions, and van der Waals forces. These properties enable peptides to bind to specific areas of DNA chains selectively. However, despite these advancements, designing G4-binding peptides remains challenging due to a lack of comprehensive information. OBJECTIVE: In our present study, we employed an in silico fragment-based approach to design G4- binding peptides. This innovative method combines machine learning classification, molecular docking, and dynamics simulation. METHODS: AutoDock Vina and Gromacs performed molecular docking and MD simulation, respectively. The machine learning algorithm was implemented by Scikit-learn. Peptide synthesis was performed using the SPPS method. The DNA binding affinity was measured by applying spectrophotometric titration. RESULTS: As a result of this approach, we identified a high-scoring peptide (p10; sequence: YWRWR). The association constant (Ka) between p10 and the ctDNA double helix chain was 4.45 × 105 M-1. Molecular modeling studies revealed that p10 could form a stable complex with the G4 surface. CONCLUSION: The obtained Ka value of 4.45 × 105 M-1 indicates favorable interactions. Our findings highlight the role of machine learning and molecular modeling approaches in designing new G4-binding peptides. Further research in this field could lead to targeted treatments that exploit the unique properties of G4 structures.

Targeting HER2 in Gastroesophageal Adenocarcinoma: Molecular Features and Updates in Clinical Practice.

Gastroesophageal adenocarcinoma (GEA) is one of the principal causes of death related to cancer globally. Human epidermal growth factor receptor 2 (HER2) is a tyrosine kinase receptor which is found to be overexpressed or amplified in approximately 20% of GEA cases. In GEA, the identification of HER2-positive status is crucial to activate a specific anti-HER2 targeted therapy. The landmark ToGA trial demonstrated the superiority of adding trastuzumab to platinum-based chemotherapy, becoming the first-line standard of treatment. However, unlike breast cancer, the efficacy of other anti-HER2 drugs, such as lapatinib, pertuzumab, and T-DM1, has failed to improve outcomes in advanced and locally advanced resectable GEA. Recently, the combination of trastuzumab with pembrolizumab, along with chemotherapy, and the development of trastuzumab deruxtecan, with its specific bystander activity, demonstrated improved outcomes, renewing attention in the treatment of this disease. This review will summarise historical and emerging therapies for the treatment of HER2-positive GEA, with a section dedicated to the HER2 molecular pathway and the use of novel blood biomarkers, such as circulating tumour DNA and circulating tumour cells, which may be helpful in the future to guide treatment decisions.

Salicylic acid and jasmonic acid biosynthetic pathways are simultaneously activated in transgenic Arabidopsis expressing the rolB/C gene from Ipomoea batatas.

The Agrobacterium rhizogenes root oncogenic locus (rol) genes interfere with hormone balance by altering their synthesis and/or recognition, giving rise to varied impacts on the physiological characteristics of plants and cell cultures. The homolog of the rolB and rolC genes from Ipomoea batatas, named Ib-rolB/C, similarly induces morphological and physiological alterations in transgenic Arabidopsis thaliana; however, its role in plant hormonal homeostasis has not been previously defined. In this study, we found that external application of salicylic acid (SA) and methyl jasmonate (MeJA) significantly upregulated Ib-rolB/C in detached I. batatas leaves. Furthermore, heterologous expression of Ib-rolB/C in A. thaliana markedly enhanced the accumulation of SA and MeJA, and to a lesser extent, elevated abscisic acid (ABA) levels, through the modulation of genes specific to hormone biosynthesis. Even though the RolB/RolC homolog protein has a notable structural resemblance to the RolB protein from A. rhizogenes, it exhibits a distinct localization pattern, predominantly residing in the cytoplasm and certain discrete subcellular structures, instead of the nucleus. Consequently, the functions of RolB/RolC in both naturally and artificially transgenic plants are linked to changes in the hormonal state of the cells, though the underlying signaling pathways remain to be elucidated.

Pharmaceutical manifestation of Knoevenagel condensed metal (II) complexes through virtual, in vitro and in vivo assessments.

Sulphur containing compounds possess a great deal of interest due to wide range of beneficial activities towards biotic species. This work also deals with the study of biological examination of newly synthesized sulphur containing Cu(II) and Zn(II) complexes derived from (E)-4-(phenylimino)-3-((E)-1-(phenylimino)ethyl)pent-2-ene-1-thiol Schiff bases. Moreover, the DNA nuclease efficiency of the synthesized metal complexes is studied by UV absorption studies, Fluorescence studies, Viscosity and CV titrations which confirm the intercalative mode of binding. Pharmacokinetic studies and drug like activity of these compounds are screened with the help of SWISS ADME online freeware. Their morphological nature is corroborated by various spectral techniques. Optimized geometry and biologically accessible nature of the synthesized compounds are investigated by Gaussian 09 W software. Interestingly, molecular docking studies are carried out against cancer DNA and 6J10 cancer cell. Anti-inflammatory and in vitro antioxidant activities have been studied to validate the theoretical prediction. Based on these preliminary pharmacological activities, the in vitro cytotoxicity and in vivo antitumor activities are examined using MCF-7, HeLa, Hep-2, HepG-2 and Ehrlich ascites carcinoma (EAC) cell lines. All the above examinations reveal that the nitro substituted transition metal complexes possess higher biological bustle.Communicated by Ramaswamy H. Sarma.

Cu(II) and Zn(II) complexes incorporating Knoevenagel ß-diketimine Schiff basesThe quantum mechanical examinations using Gaussian 09W softwareInvestigation of possible interactions with DNA by Molecular docking studiesIn silico, in vitro and in vivo analyses of the synthesized compounds.

Unleashing the binding interaction of chrysin-Cu(II) complex with the biomacromolecular targets: further studies of cell cytotoxicity and radical scavenging properties.

Flavonoids are significant dietary components and have ability to coordinate with metal ions to produce novel drug discovery leads that are superior to those of the parent flavonoids. Here, in this report, we have synthesized chrysin-Cu(II) complex (as per reported article) and characterized it further with different analytical techniques. The synthesized complex was evaluated for radical scavenging and cell cytotoxicity studies where it exhibited enhanced activity as compared to bare chrysin. The interaction studies of the complex with ct-DNA (Kb ⁓ 105 M-1), human serum albumin (HSA) and ovalbumin (Kb ⁓ 104 M-1) were evaluated using multi-spectroscopic and molecular docking studies. Groove binding mode with ct-DNA was observed as confirmed from competitive displacement studies, viscosity measurement, melting temperature estimation and docking analyses. The complex exhibited comparatively higher affinity towards ct-DNA which indicated it efficient transportation by the carrier proteins and controlled release in the target DNA.Communicated by Ramaswamy H. Sarma.

Molecular interaction of antiviral drug penciclovir with DNA and HSA insights from experimental and docking studies.

One approach to accelerate the availability of new cancer drugs is to test drugs approved for other conditions as anticancer agents. During recent decades, penciclovir (PNV) has been frequently utilized as a potent antiviral drug, in particular against infections caused by herpes viruses. Many antivirals interact with DNA and change their expression level, so determining the binding mode is of great importance. In our laboratory, we have focused our attention to design improved drugs that target cellular DNA, to understand the mechanism of action at the molecular level, and also to investigate the effect of antiviral drugs as anticancer agents. The results of ct-DNA-PNV interactions at physiological pH using fluorescence spectroscopy, UV-visible absorption spectroscopy, and molecular modeling reveal this drug binds well to ct-DNA through groove binding. The circular dichroism measurements displayed that PNV caused a slight change in the DNA structure which affirmed that the binding of PNV with the DNA occurs through the groove mode. Besides, multi-spectroscopic and molecular docking were used to evaluate how PNV interacts with human serum albumin under physiological conditions. The findings of fluorescence quenching suggested that static quenching was involved in the spontaneous development of HSA-PNV complex through hydrophobic force. The docking simulation results validated the findings of spectroscopic techniques.Communicated by Ramaswamy H. Sarma.

Quercetin-phenylalanine 3d-transition metal-based {Co(II), Ni(II) & Cu(II)} intercalative therapeutic agents: DNA & BSA interaction studies in vitro and cleavage activity.

New Quercetin-phenylalanine metal-based therapeutic agents of the formulation [Qu(Phe)M(II).(H2O)2].NO3 where M(II) = Co(II) and Ni(II) and [Qu(Phe)Cu(II).(H2O)2] were synthesized and their structure was predicted by IR, UV-vis, EPR and ESI-MS spectroscopic techniques. The bio-molecular interaction studies of the Quercetin-phenylalanine complexes, 1-3 with ct-DNA and BSA were performed using a battery of complimentary biophysical techniques. The corroborative results of these experiments revealed strong binding propensity via electrostatic interactions probably through minor grove binding towards ct-DNA, therapeutic target. The binding affinity of Quercetin-phenylalanine complexes 1-3 was quantified by determining binding constants values, Kb, Ksv, and the magnitude of binding propensity followed the order 3 > 1 > 2, implicating the preferential binding of Cu(II) complex 3 with ct-DNA. The cleavage studies were performed with complexes using gel electrophoretic mobility assay. The complexes 1-3 demonstrated efficient cleaving ability by the hydrolytic cleavage pathway involving hydroxyl (OH) radicals. BSA binding profile of Quercetin-phenylalanine metal therapeutics 1-3 was studied in order to understand the drug carrier potential of these compounds and found that complex 3 was capable of binding preferentially with BSA as compared to other complexes.

Non-covalent binding interaction of bioactive coumarin esculetin with calf thymus DNA and yeast transfer RNA: A detailed investigation to decipher the binding affinities, binding location, interacting forces and structural alterations at a molecular level.

Esculetin is a well-known coumarin derivative found abundantly in nature possessing an extensive array of pharmacological and therapeutic properties. Consequently, to comprehend its molecular recognition mechanism, our objective is to conduct a complete investigation of its interactions with the nucleic acid, specifically ct-DNA, and t-RNA, using spectroscopic and computational techniques. The intrinsic fluorescence of esculetin is quenched when it interacts with ct-DNA and t-RNA, and this occurs through a static quenching mechanism. The thermodynamic parameters demonstrated that the interaction is influenced by hydrogen bonding and weak van der Waals forces. CD and FT-IR results revealed no conformational changes in ct-DNA and t-RNA structure on binding with esculetin. Furthermore, competitive displacement assay with ethidium bromide, melting temperature, viscosity measurement, and potassium iodide quenching experiments, reflected that esculetin probably binds to the minor groove of ct-DNA. The molecular docking results provided further confirmation for the spectroscopic findings, including the binding location of esculetin and binding energies of esculetin complexes with ct-DNA and t-RNA. Molecular dynamics simulation studies demonstrated the conformational stability and flexibility of nucleic acids.

Biophysical insights on the interaction of anticoagulant drug dicoumarol with calf thymus-DNA: deciphering the binding mode and binding force with thermodynamics.

The biological activity of drugs is exhibited due to their interactions with bio-receptors. Dicoumarol (DIC) is a natural hydroxycoumarin and a well-known anticoagulant. DNA is the genetic material and one of the targets of numerous drugs. The interaction of DIC with calf-thymus DNA (ct-DNA) has been studied using different biophysical techniques and docking studies. The binding constant in the order of 103 to 104 M-1 was observed from spectroscopic studies. Thermodynamic studies at 4 different temperatures revealed the spontaneity of the interaction with the entropy-driven process. Marker displacement studies with competitive markers of intercalators (ethidium bromide) and groove binders (Hoechst 33258) confirmed the groove-binding nature of DIC in DNA. The groove-binding mode of DIC was complemented by different studies like viscosity measurements, DNA melting, and the effect of KI on the binding. A minor perturbation in the DNA viscosity and no significant change in the DNA melting temperature (Tm) after binding with DIC further confirms the groove binding mode. The effect of KI on the DIC and DIC-DNA system suggested the absence of DIC intercalation. The absence of significant electrostatic force was revealed from the ionic-strength effect study. Binding-induced conformational variation in ct-DNA was absent in circular dichroism studies. Molecular docking studies suggested the position of DIC within the minor groove of ct-DNA, covering three base pairs long. The outcome of this report may help in understanding the pharmacodynamics and pharmacokinetics of dicoumarol analogs and related molecules.Communicated by Ramaswamy H. Sarma.

Nicotiana noctiflora Hook. Genome Contains Two Cellular T-DNAs with Functional Genes.

Agrobacterium (Rhizobium)-mediated transformation leads to the formation of crown galls or hairy roots on infected plants. These effects develop due to the activity of T-DNA genes, gathered on a big plasmid, acquired from agrobacteria during horizontal gene transfer. However, a lot of plant species are known to contain such sequences, called cellular T-DNAs (cT-DNAs), and maintain normal phenotypes. Some of the genes remain intact, which leads to the conclusion of their functional role in plants. In this study, we present a comprehensive analysis of the cT-DNAs in the Nicotiana noctiflora Hook. genome, including gene expression and opine identification. Deep sequencing of the Nicotiana noctiflora genome revealed the presence of two different cT-DNAs, NnT-DNA1 and NnT-DNA2, which contain the intact genes iaaM, iaaH, acs, orf13, orf13a, and orf14. According to the expression analysis results, all these genes are most active in roots in comparison with other organs, which is consistent with data on cT-DNA gene expression in other plant species. We also used genetic engineering approaches and HPTLC and HPLC-MS methods to investigate the product of the acs gene (agrocinopine synthase), which turned out to be similar to agrocinopine A. Overall, this study expands our knowledge of cT-DNAs in plants and brings us closer to understanding their possible functions. Further research of cT-DNAs in different species and their functional implications could contribute to advancements in plant genetics and potentially unveil novel traits with practical applications in agriculture and other fields.

Extensive natural Agrobacterium-induced transformation in the genus Camellia.

MAIN CONCLUSION: The genus Camellia underwent extensive natural transformation by Agrobacterium. Over a period of 15 million years, at least 12 different inserts accumulated in 72 investigated Camellia species. Like a wide variety of other wild and cultivated plants, Camellia species carry cellular T-DNA sequences (cT-DNAs) in their nuclear genomes, resulting from natural Agrobacterium-mediated transformation. Short and long DNA sequencing reads of 435 accessions belonging to 72 Camellia species (representing 12 out of 14 sections) were investigated for the occurrence of cT-DNA insertions. In all, 12 different cT-DNAs were recovered, either completely or partially, called CaTA to CaTL. Divergence analysis of internal cT-DNA repeats revealed that the insertion events span a period from 0.075 to 15 Mio years ago, and yielded an average transformation frequency of one event per 1.25 Mio years. The two oldest inserts, CaTA and CaTD, have been modified by spontaneous deletions and inversions, and by insertion of various plant sequences. In those cases where enough accessions were available (C. japonica, C. oleifera, C. chekiangoleosa, C. sasanqua and C. pitardii), the younger cT-DNA inserts showed a patchy distribution among different accessions of each species, indicating that they are not genetically fixed. It could be shown that Camellia breeding has led to intersectional transfer of cT-DNAs. Altogether, the cT-DNAs cover 374 kb, and carry 47 open reading frames (ORFs). Two Camellia cT-DNA genes, CaTH-orf358 and CaTK-orf8, represent new types of T-DNA genes. With its large number of cT-DNA sequences, the genus Camellia constitutes an interesting model for the study of natural Agrobacterium transformants.