Interaction of 9-Methoxyluminarine with Different G-Quadruplex Topologies: Fluorescence and Circular Dichroism Studies

The interactions of G–quadruplexes of different topologies with highly fluorescent 9-methoxyluminarine ligand 9-MeLM were investigated by fluorescence and circular dichroism spectroscopy. The results showed that 9-methoxyluminarine was able to interact and did not destabilize any investigated molecular targets. The studied compound was selectively quenched by parallel c-MYC G-quadruplex DNA, whereas hybrid and antiparallel G4 topology caused only a negligible decrease in the fluorescence of the ligand. A high decrease of fluorescence of the ligand after binding with c-MYC G-quadruplex suggests that this molecule can be used as a selective probe for parallel G-quadruplexes.     

NBD‐Based Green Fluorescent Ligands for Typing of Thymine‐Related SNPs by Using an Abasic Site‐Containing Probe DNA

The binding behavior of green fluorescent ligands, derivatives of 7‐nitrobenzo‐2‐oxa‐1,3‐diazole (NBD), with DNA duplexes containing an abasic (AP) site is studied by thermal denaturation and fluorescence experiments. Among NBD derivatives, N¹‐(7‐nitrobenzo[c][1,2,5]oxadiazol‐4‐yl)propane‐1,3‐diamine (NBD‐NH₂) is found to bind selectively to the thymine base opposite an AP site in a DNA duplex with a binding affinity of 1.52×10⁶ M ^?1. From molecular modeling studies, it is suggested that the NBD moiety binds to thymine at the AP site and a protonated amino group tethered to the NBD moiety interacts with the guanine base flanking the AP site. Green fluorescent NBD‐NH₂ is successfully applied for simultaneous G>T genotyping of PCR amplification products in a single cuvette in combination with a blue fluorescent ligand, 2‐amino‐6,7‐dimethyl‐4‐hydroxypteridine (diMe‐pteridine).     

DNA interaction and cytotoxic activity of copper complex based on tridentate hydrazone derived ligand and nitrogen donor heterocycle

Tetra coordinated copper(II) complex of formula [Cu(L¹)(imid)] is synthesized using imidazole and a tridentate O,N,O hydrazone ligand L¹ prepared by condensation of 1,1,1-trifluoropentanedione and 4-chlorobenzhydrazide. The ligand and complex are characterized by UV–Visible, FTIR, NMR, mass and single crystal XRD techniques. DNA binding mode is assessed by UV absorption, fluorescence spectral and circular dichroism studies. Based on the results, it is observed that complex has preferred intercalative mode of binding with DNA. The binding constant, K_b, was found to be 4.02 ± 0.09 × 10⁴ M^− 1. Thermodynamic parameters such as ΔH, ΔS and ΔG obtained from acridine orange fluorescence displacement assay revealed that the hydrophobic and hydrogen bonding interactions are playing a major role in the binding pattern. In addition, cytotoxicity of the complex towards MCF-7 breast cancer cell line has also been assessed.     

Molecular Recognition of Parallel DNA Quadruplex d(TTAGGGT)4 by Mitoxantrone: Binding with 1:2 Stoichiometry Leading to Thermal Stabilization and Telomerase Inhibition

The interaction of the anthraquinone derivative mitoxantrone, a semisynthetic anti‐cancer drug with two non‐planar side chains, with heptamer G‐quadruplex d(TTAGGGT)₄, which contains the human telomere DNA sequence, was evaluated by differential scanning calorimetry, fluorescence Job plotting, absorption, and NMR and CD spectroscopy. Binding led to thermal stabilization of DNA (ΔT_m=13–20 °C). The spectra revealed that two mitoxantrone molecules bind externally at two sites of the DNA quadruplex as monomers, by partial insertion of the chromophore and side‐chain interaction at the grooves. The inhibition of telomerase (IC₅₀=2 μm), as determined by a TRAP assay, can be attributed to thermal stabilization of the DNA quadruplex because of the interactions with mitoxantrone. The studies revealed highly specific molecular recognition between a ligand and a parallel‐stranded G‐quadruplex; this might serve as a platform for the rational design of new drugs.     

Exploring the Interaction of Curaxin CBL0137 with G-Quadruplex DNA Oligomers

Curaxins and especially the second-generation derivative curaxin CBL0137 have important antitumor activities in multiple cancers such as glioblastoma, melanoma and others. Although most of the authors suggest that their mechanism of action comes from the activation of p53 and inactivation of NF-kB by targeting FACT, there is evidence supporting the involvement of DNA binding in their antitumor activity. In this work, the DNA binding properties of curaxin CBL0137 with model quadruplex DNA oligomers were studied by ¹H NMR, CD, fluorescence and molecular modeling. We provided molecular details of the interaction of curaxin with two G-quadruplex structures, the single repeat of human telomere d(TTAGGGT)₄ and the c-myc promoter Pu22 sequence. We also performed ¹H and ³¹P NMR experiments were also performed in order to investigate the interaction with duplex DNA models. Our data support the hypothesis that the interaction of curaxin with G-quadruplex may provide a novel insight into the DNA-binding properties of CBL0137, and it will be helpful for the design of novel selective DNA-targeting curaxin analogues.     

Stripping of Rh–Sn–Cl‐loaded alkylated 8‐hydroxyquiniline with Na2SO3–HCl

Acidic rhodium(III) chloride solutions pretreated with SnCl₂ at a ratio Sn(II) : Rh(III) = 3 are contacted with an alkylated 8‐hydroxyquinoline (Kelex 100) to prepare loaded organics which subsequently are subjected to stripping with Na₂SO₃–HCl solutions. Rh(III) was found to be stripped selectively involving the following reaction: where H₂Q is the protonated form of kelex 100. Direct characterization of the stripped complex and slope analysis confirmed this stripping reaction. Equilibrium was reached within 20 min. A kinetic analysis found the stripping process to be irreversible and to be controlled most probably by the slow release of SnCl₃^? ligand from the intermediate Rh–Q–SO₃–SnCl₃ organic complex. Copyright © 2004 Society of Chemical Industry     

Discovery of New E‐Selectin Inhibitors by Virtual Screening,Fluorescence Binding Assays,and STD NMR Experiments

E‐selectin is an endothelial protein that participates in the adhesion of metastatic cancer cells, and is therefore a relevant target for antitumor therapeutic intervention. In this work, virtual screening was used to identify new E‐selectin inhibitors from a subset of drug‐like molecules retrieved from the ZINC database, including the physiological ligand sLe^x as reference structure (PDB ID: 1G1T ). Four hits were chosen and subjected to molecular dynamics simulations and fluorescence binding assays, which led to the determination of experimental dissociation constants between 333 and 1012 μm . The candidate with the highest affinity was studied by saturation transfer difference (STD) NMR experiments and complete relaxation and conformational exchange matrix analysis of saturation transfer (CORCEMA‐ST), aimed at identifying the preferable binding mode with E‐selectin. Our results revealed that this new inhibitor binds more strongly than sLe^x in the E‐selectin binding site, in good agreement with simulation predictions. These properties will prove valuable for the future design of drugs that target E‐selectin.     

Multivalent Display and Receptor‐Mediated Endocytosis of Transferrin on Virus‐Like Particles

The structurally regular and stable self‐assembled capsids derived from viruses can be used as scaffolds for the display of multiple copies of cell‐ and tissue‐targeting molecules and therapeutic agents in a convenient and well‐defined manner. The human iron‐transfer protein transferrin, a high affinity ligand for receptors upregulated in a variety of cancers, has been arrayed on the exterior surface of the protein capsid of bacteriophage Qβ. Selective oxidation of the sialic acid residues on the glycan chains of transferrin was followed by introduction of a terminal alkyne functionality through an oxime linkage. Attachment of the protein to azide‐functionalized Qβ capsid particles in an orientation allowing access to the receptor binding site was accomplished by the Cu^I‐catalyzed azide–alkyne cycloaddition (CuAAC) click reaction. Transferrin conjugation to Qβ particles allowed specific recognition by transferrin receptors and cellular internalization through clathrin‐mediated endocytosis, as determined by fluorescence microscopy on cells expressing GFP‐labeled clathrin light chains. By testing Qβ particles bearing different numbers of transferrin molecules, it was demonstrated that cellular uptake was proportional to ligand density, but that internalization was inhibited by equivalent concentrations of free transferrin. These results suggest that cell targeting with transferrin can be improved by local concentration (avidity) effects.     

Towards a new strategy of a chitosan‐based molecularly imprinted membrane for removal of 4‐nitrophenol in real water samples

The issue of water contaminants, which affects human and environmental health, is not trivial. It is thus paramount to find new cheap and user friendly ways to detect and remove them from the environment. Here, the synthesis of a green chitosan (CS ) based molecularly imprinted membrane for the detection and quantification of 4‐nitrophenol (4‐NO₂Ph ) in aqueous media is proposed. The concentration of 4‐NO₂Ph in a water solution was measured by HPLC analysis. CS as a functional polymer, 4‐NO₂Ph as template, 4‐[(4‐hydroxy)phenylazo]benzenesulfonic acid as ligand, and glutaraldehyde as crosslinker in the presence of polyethylene glycol as porogen were used. The membrane was characterized by SEM and Fourier transform IR analyses, which confirmed the CS and polyethylene glycol backbone of the membrane. Kinetic studies of the detection system were performed by using pseudo‐first‐order and pseudo‐second‐order models. Then, the binding efficiency between 195.33 µmol L^?1 and 9235.55 µmol L^?1 of 4‐NO₂Ph was evaluated, finding a maximum adsorption of 723.25 µmol 4‐NO₂Ph per gram of membrane consistent with the Q _max calculated from the Langmuir isotherm. The selectivity of the membrane versus three phenolic competitor molecules, sharing very similar molecular structure to 4‐NO₂Ph , was demonstrated. Finally, the applicability of the membrane to real‐world samples was evaluated, by using drinking water spiked with 7.19 µmol L^?1 of 4‐NO₂Ph , obtaining a removal efficiency of 70.6%. © 2017 Society of Chemical Industry     

DNA and copper (II) governed fluorescence tuning of phenanthroline possessing Ru(II) complex. Interplay of electrostatic interactions

Tuning of the fluorescence properties of [Ru (bpy) ₂ (L₁)] ^2 + (1) [L₁ = 2-(2-methoxyphenyl)-1H–imidazo [4, 5-f] Kumar et al. (2010), Rawle et al. (1992) phenanthroline] by DNA and/or Cu^2 + ion have been investigated. Fluorescence of 1 gets enhanced upon binding to DNA owing to electrostatic interactions. The binding constant of 8.4 × 10⁴ has been evaluated. Cu^2 + ions though unable to alter the fluorescence intensity of free 1 but successfully sequestrate the DNA bound 1, evidently decreasing its fluorescence intensity. The reversibility of the sequestration process was ascertained by the recovery of the quenched fluorescence intensity via introduction of equimolar EDTA to the buffer system of DNA bound 1 and Cu^2 +.     

Small-Molecule Activators of Glucose-6-phosphate Dehydrogenase (G6PD) Bridging the Dimer Interface

We recently identified AG1, a small-molecule activator that functions by promoting oligomerization of glucose-6-phosphate dehydrogenase (G6PD) to the catalytically competent forms. Biochemical experiments indicate that the activation of G6PD by the original hit molecule (AG1) is noncovalent and that one C₂-symmetric region of the G6PD homodimer is important for ligand function. Consequently, the disulfide in AG1 is not required for activation of G6PD, and a number of analogues were prepared without this reactive moiety. Our study supports a mechanism of action whereby AG1 bridges the dimer interface at the structural nicotinamide adenine dinucleotide phosphate (NADP⁺) binding sites of two interacting G6PD monomers. Small molecules that promote G6PD oligomerization have the potential to provide a first-in-class treatment for G6PD deficiency. This general strategy could be applied to other enzyme deficiencies in which control of oligomerization can enhance enzymatic activity and/or stability.     

Developing High Field MRI Contrast Agents by Tuning the Rotational Dynamics: Bisaqua GdAAZTA-based Dendrimers

Monomeric and dimeric AAZTA-based bifunctional chelators (AAZTA=6-amino-6-methylperhydro-1,4-diazepine tetraacetic acid) were attached to different generations (G0, G1 and G2) of ethylenediamine-cored PAMAM dendrimers (PAMAM=polyamidoamine) to obtain a series of six dendrimeric systems with 4 to 32 chelates at the periphery. These Gd^III-loaded dendrimers have molecular weight ranging from 3.5 to 25 kDa, thus allowing a systematic investigation on the changes in relaxivity (r₁) with the variation of the rotational dynamics following the increase in molecular size. Variable-temperature ¹⁷O NMR (on the dimeric building block Gd₂ L2 ) and ¹H Nuclear Magnetic Relaxation Dispersion measurements at different temperatures indicate that the water exchange lifetime (τ_M∼90 ns) of the two inner sphere water molecules does not represent a limiting factor to the relaxivity of the systems. The r₁ values at 1.5 T (60 MHz) and 298 K increases from 10.2 mM⁻¹ s⁻¹ for the monomer Gd L1 to 31.4 mM⁻¹ s⁻¹ for the dendrimer Gd₃₂ G2-32 (+308 %). However, the relaxivity (per Gd) does not show a linear dependence on the molecular mass, but rather the enhancement tends to attenuate markedly for larger systems. This effect has been attributed to the growing decrease in correlation between local rotational motions and global molecular tumbling.     

Binding properties of polyamidoamine dendrimers

Dendrimers are globular, hyperbranched polymers possessing a high concentration of surface functional groups and internal cavities. These unique features make them good host molecules for small ligands. To reveal relationships between dendrimer size and its encapsulating properties, the interactions of the fourth and the sixth generations of polyamidoamine dendrimers (PAMAM G4 and PAMAM G6) with a fluorescent dye 1‐anilinonaphthalene‐8‐sulfonate (ANS) were studied. Because ANS is a fluorescent molecule and its fluorescence is very sensitive to changes in its microenvironment, it was possible to use spectrofluorometric methods to evaluate the interactions with dendrimers. A double fluorometric titration method was used to estimate a binding constant and the number of binding centers. There were two types of dendrimer binding centers characterized by different affinity towards ANS. For PAMAM G4, the values of K_b and n for low‐affinity and high‐affinity sites equaled to 2.6 × 10⁵, 0.60 and 3.70 × 10⁶, 0.34, respectively, whereas in the case of PAMAM G6, these values equaled to 1.2 × 10⁵, 76.34 and 1.38 × 10⁶, 22.73. It was observed that the size of the dendrimer had a strong impact on the number of ANS molecules that interacted with dendrimers and their location within the macromolecule. © 2006 Wiley Periodicals, Inc. J Appl Polym Sci 103: 2036–2040, 2007     

Studies on interaction of Cr(III) polypyridyl complexes with DNA

Two chromium(III) complexes containing derivatives of 1,10-phenanthroline were synthesized and characterized using different spectroscopic methods. The interaction of the synthesized complexes with DNA was performed. From the absorption titration data, the binding constant for the complexes [Cr(imiphen)₂Cl₂]⁺ (1) and [Cr(furphen)₂Cl₂]⁺ (2) with DNA were found to be 5.69 ± 0.3 × 10⁴ M^− 1 for 1 and 3.7 ± 0.2 × 10⁴ M^− 1 for 2, respectively. The viscosity studies show that both the complexes bind to DNA in a partial intercalative mode. The results of competitive binding assay reveal that both complexes displace EB from DNA marginally. Gel electrophoresis data show that both the complexes 1 and 2 requires co-reagent to induce DNA cleavage but complex 2 exhibit better nuclease activity compared to complex 1. The results from this study describe the role of ligand structure in the binding and cleavage of DNA as furan based ligand exhibit better cleaving ability compared to imidazole moiety.     

Synthesis and structural characterization of a disulphide-bridged tetranuclear palladium(II) complex derived from 3,5-diacetyl 1,2,4-triazole bis(4-ethylthiosemicarbazone)

A novel tetranuclear [Pd₄(μ₂-η²-S₂)(H₂L¹)₂] complex, where H₂L¹ is the anion of 3,5-diacetyl-1,2,4-triazole bis(4-ethylthiosemicarbazone), was synthesized and structurally characterized by single crystal X-ray diffraction. The molecular structure shows an unexpected μ₂-η²-S₂ bridge, which is shared by all the four palladium atoms. Each trideprotonate 3,5-diacetyl-1,2,4-triazole bis(4-ethylthiosemicarbazone) ligand acts as a hexadentate in a symmetrical manner bridging two metal ions through the two adjacent nitrogen atoms of the central triazole ring. The molecular packing is characterized by hydrogen bond interactions to form a 3D supramolecular architecture with channels running down the c¹ axis in which the lattice water molecules are encapsulated.     

Methyl,Ethyl, Propyl,Butyl: Futile But Not for Water,as the Correlation of Structure and Thermodynamic Signature Shows in a Congeneric Series of Thermolysin Inhibitors

Water is ubiquitously present in any biological system and has therefore to be regarded as an additional binding partner in the protein–ligand binding process. Upon complex formation, a new solvent‐exposed surface is generated and water molecules from the first solvation layer will arrange around this newly formed surface. So far, the influence of such water arrangements on the ligand binding properties is unknown. In this study, the binding modes of nine congeneric phosphonamidate‐type inhibitors with systematically varied, size‐increasing hydrophobic P₂′ substituents (from methyl to phenylethyl) addressing the hydrophobic, solvent‐exposed S₂′ pocket of thermolysin were analyzed by high‐resolution crystal structures and correlated with their thermodynamic binding profiles as measured by isothermal titration calorimetry. Overall, ΔΔG spreads over 7.0 kJ mol^?1, ΔΔH varies by 15.8 kJ mol^?1, and ?TΔΔS by 12.1 kJ mol^?1. Throughout the series, these changes correlate remarkably well with the geometric differences of water molecules arranged adjacent to the P₂′ substituents. Ligands with medium‐sized P₂′ substituents exhibit highest affinities, presumably because of their optimal solvation patterns around these complexes. The addition, removal, or rearrangement of even a single methyl group can result in a strong modulation of the adjacent water network pattern shifting from enthalpy to entropy‐driven binding. In conclusion, the quality of a water network assembled around a protein–ligand complex influences the enthalpy/entropy signature and can even modulate affinity to a surprising extent.     

Studies on thermodynamic nature of steroselectivity for ruthenium(II) polypyridyl complex binding to DNA

A pair of ruthenium(II) complex isomers, Δ- and Λ-[Ru(bpy)₂(p-mpip)]²⁺ {bpy = 2,2′-bipyridine, p-mpip = 2-(4-methylphenyl)imidazo[4,5-f]1,10-phenanthroline} have been synthesized and characterized. Thermodyanmics of binding of the two isomers to calf thymus DNA (CT DNA) has been investigated by isothermal titration calorimetry (ITC). It is rare that the Λ isomer binds to DNA more strongly than the Δ isomer, which has been verified by viscosity measurements, and can be explained by their different thermodynamic driving forces of DNA binding for the first time. This suggests that the dominant factor governing the stereoselectivity of DNA binding of metal complex may be the different thermodynamic driving forces between interactions of its different isomer with DNA. This new finding may be very helpful to understand the nature of steroselective DNA binding of small chiral molecules, and be useful to the development of DNA molecular probes and new DNA targeting therapeutic drugs.     

Synthesis, Supramolecular Architecture and DNA Binding Studies of a Novel Heterotrinuclear FeIII/CoII/FeIII Complex: [Co(phen)2][Fe(phen)(CN)4]2·4H2O (phen?=?1,10-phenanthroline)

A novel cyano-bridged heterotrinuclear Fe^III/Co^II/Fe^III complex, [Co(phen)₂][Fe(phen)(CN)₄]₂·4H₂O (1) (phen = 1,10-phenanthroline), has been synthesized under solvothermal conditions, and characterized by elemental analysis, FI-IR spectrum and single-crystal X-ray diffraction. The structure analysis reveals complex 1 is constructed into a 3D supramolecular architecture through abundant intermolecular non-covalent interactions. According to the supramolecular self-assembly of complex 1, five types of π–π stacking models of phenanthroline ligands, hydrogen bonds formed by four lattice water molecules, and C–H···N weak interactions involving cyano groups are all observed. Electronic absorption spectroscopy and fluorescence titration studies of the interaction between complex 1 and calf thymus DNA are suggestive of an intercalative binding mode with an intrinsic binding constant of 7.95 × 10³ M⁻¹ and a linear Stern–Volmer quenching constant of 1.033 × 10⁵ M⁻¹.     

Stimuli-Responsive and Reversible Nanoassemblies of G-Triplexes

G-triplex (G3) structures formed with three consecutive G-tracts have recently been identified as a new emerging guanine-rich DNA fold. There could likely be a wide range of biological functions for G3s as occurring for G-quadruplex (G4) structures formed with four consecutive G-tracts. However, in comparison to the many reports on G4 nanoassemblies that organize monomers together in a controllable manner, G3-favored nanoassemblies have yet to be explored. In this work, we found that a natural alkaloid of sanguinarine can serve as a dynamic ligand glue to reversibly switch the dimeric nanoassemblies of the thrombin binding aptamer G3 (TBA-G3). The glue planarity was considered to be a crucial factor for realizing this switching. More importantly, external stimuli including pH, sulfite, O₂ and H₂O₂ can be employed as common regulators to easily modulate the glue's adhesivity for constructing and destructing the G3 nanoassemblies as a result of the ligand converting between isoforms. However, this assembly behavior does not occur with the counterpart TBA-G4. Our work demonstrates that higher-order G3 nanoassemblies can be reversibly operated by manipulating ligand adhesivity. This provides an alternative understanding of the unique behavior of guanine-rich sequences and focuses attention on the G3 fold since the nanoassembly event investigated herein might occur in living cells.     

Binding of Sulpiride to Seric Albumins

The aim of this work was to study the interaction of sulpiride with human serum albumin (HSA) and bovine serum albumin (BSA) through the fluorescence quenching technique. As sulpiride molecules emit fluorescence, we have developed a simple mathematical model to discriminate the quencher fluorescence from the albumin fluorescence in the solution where they interact. Sulpiride is an antipsychotic used in the treatment of several psychiatric disorders. We selectively excited the fluorescence of tryptophan residues with 290 nm wavelength and observed the quenching by titrating HSA and BSA solutions with sulpiride. Stern-Volmer graphs were plotted and quenching constants were estimated. Results showed that sulpiride form complexes with both albumins. Estimated association constants for the interaction sulpiride–HSA were 2.20 (±0.08) × 10⁴ M⁻¹, at 37 °C, and 5.46 (±0.20) × 10⁴ M⁻¹, at 25 °C. Those for the interaction sulpiride-BSA are 0.44 (±0.01) × 10⁴ M⁻¹, at 37 °C and 2.17 (±0.04) × 10⁴ M⁻¹, at 25 °C. The quenching intensity of BSA, which contains two tryptophan residues in the peptide chain, was found to be higher than that of HSA, what suggests that the primary binding site for sulpiride in albumin should be located next to the sub domain IB of the protein structure.