A fluorescent sensor for 2,3-bisphosphoglycerate using a europium tetra-N-oxide bis-bipyridine complex for both binding and signaling purposes

Host 1 was designed and synthesized as a fluorescent sensor for 2,3-bisphosphoglycerate (BPG, 3). The design features a tris-functionalized triethylbenzene core to preorganize binding groups. The three cationic moieties, a tetra-N-oxide bipyridine-europium complex and two ammonium groups, were included to complement the three anionic functionalities on the guest. Beyond acting as a binding site, the europium complex was used to signal binding of the guest through modification of the charge transfer emission. A 1:1 complex with BPG was determined in 50 % methanol/acetonitrile with a K(a) of 6.7 x 10(5) mol(-1) by monitoring the reduction of the fluorescence signal upon guest addition. In the titration of related glycolytic intermediates lacking a second phosphate (4-6) into host 1, 2:1 host to guest binding was observed. Similarly, control compound 2, which lacks the ammonium groups, binds BPG and 4-6 in a 2:1 fashion. Also, phenylphosphate 7 binds to host 1 in a 1:1 stoichiometry with a K(a) over three times less than 3.     

Disruption of protein-protein interactions: design of a synthetic receptor that blocks the binding of cytochrome c to cytochrome c peroxidase

Synthetic receptor 1 has been found via fluorescence titration to compete effectively with cytochrome c peroxidase for binding cytochrome c (Cc), forming 1:1 Cc:1 complex with a binding constant of (3 +/- 1) x 10(8) M-1, and to disrupt Cc: Apaf-1 complex, a key adduct in apoptosis.     

Binding of amino acids into a novel multiresponsive ferrocene receptor having an ene backbone

[reaction: see text] Receptor 1 featuring two open arms, multipoint binding sites, and unsaturated linkers on a ferrocene platform shows strong 1:1 binding to unprotected alpha-amino acids (UV-vis, fluorescence, CV, ITC, NMR, and ESI-MS). NMR and ESI-MS studies suggest an encapsulative binding mode involving the alpha,beta-unsaturated carbonyl residue (site for -NH3+, interaction A) and the terminal -OH groups (site for -COO-, interaction B).     

Energetics of the human Tel-22 quadruplex-telomestatin interaction: a molecular dynamics study

The formation and stabilization of telomeric quadruplexes has been shown to inhibit the activity of telomerase, thus establishing telomeric DNA quadruplex as an attractive target for cancer therapeutic intervention. In this context, telomestatin, a G-quadruplex-specific ligand known to bind and stabilize G-quadruplex, is of great interest. Knowledge of the three-dimensional structure of telomeric quadruplex and its complex with telomestatin in solution is a prerequisite for structure-based rational drug design. Here, we report the relative stabilities of human telomeric quadruplex (AG3[T2AG3]3) structures under K+ ion conditions and their binding interaction with telomestatin, as determined by molecular dynamics simulations followed by energy calculations. The energetics study shows that, in the presence of K+ ions, mixed hybrid-type Tel-22 quadruplex conformations are more stable than other conformations. The binding free energy for quadruplex-telomestatin interactions suggests that 1:2 binding is favored over 1:1 binding. To further substantiate our results, we also calculated the change in solvent-accessible surface area (DeltaSASA) and heat capacity (DeltaCp) associated with 1:1 and 1:2 binding modes. The extensive investigation performed for quadruplex-telomestatin interaction will assist in understanding the parameters influencing the quadruplex-ligand interaction and will serve as a platform for rational drug design.     

Sulfate‐Selective Binding and Sensing of a Fluorescent [3]Rotaxane Host System

The chloride‐templated synthesis of a novel [3]rotaxane, capable of binding anionic guests, and incorporating a naphthalene group for fluorescence sensing is reported. Extensive ¹H NMR titration studies were used to probe the anion binding selectivity of the system. The rotaxane selectively recognises sulfate, undergoing an induced conformational change upon sulfate binding to form a 1:1 stoichiometric sandwich‐type complex, concomitant with significant quenching of the fluorescence. Binding of mono‐anionic guests results in the formation of a 2:1 stoichiometric guest–host complex, and a modest enhancement of the emission. Addition of an excess of sulfate in non‐competitive solvent also results in a 2:1 emissive complex.     

Interaction between antitumor drugs and a double-stranded oligonucleotide studied by electrospray ionization mass spectrometry

Electrospray ionization mass spectrometry was used to investigate the complex formation between a double-stranded oligonucleotide and various antitumor drugs belonging to two categories: intercalators (ethidium bromide, amsacrine and ascididemin) and minor groove binders (Hoechst 33258, netropsin, distamycin A, berenil and DAPI). The goal of this study was to determine whether the relative intensities in the mass spectra reflect the relative abundances of the species in the solution phase. The full-scan mass spectra suggest non-specific binding for the intercalators and specific binding for the minor groove binders. The preferential stoichiometries adopted by each minor groove binder were determined by studying the influence of the drug concentration on the spectra. We obtained 2:1 > 1:1 for distamycin, 1:1 > 2:1 for Hoechst 33258 and DAPI and only the 1 : 1 complex for netropsin and berenil. These features reflect their known behavior in solution. The compared tandem mass spectra of the 1 : 1 complexes with Hoechst 33258 and netropsin, when correlated with published crystallographic data, suggest the possibility of inferring some structural information. The relative binding affinities of the drug for the considered duplex were deduced with two by two competition experiments, assuming that the relative intensities reflect the composition of the solution phase. The obtained affinity scale is netropsin > distamycin A > DAPI > Hoechst 33258 > berenil. These examples show some of the potential uses of mass spectrometry as a useful tool for the characterization of specific drug binding to DNA, and possibly a rapid drug screening method requiring small amounts of materials.     

A Designed Inhibitor of a CLC Antiporter Blocks Function through a Unique Binding Mode

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Highlights? OADS is a known small-molecule inhibitor of a CLC antiporter ? OADS specifically inhibits the ClC-ec1 antiporter but not the ClC-1 channel ? Photoaffinity labeling and mass spectrometry have localized OADS binding to two discrete sites ? The unique binding mode and lipid dependence of OADS suggest potential mechanisms of action     

Weak C-H/pi interaction participates in the diastereoselectivity of a host-guest complex in the presence of six strong hydrogen bonds

[structure: see text] We report a study of the interaction between methylmethanetriacetic acid (MMTA) and a tripodal amidopyridine receptor 1, where the geometry of the binding is in part governed by a weak C-H/pi interaction in the presence of six strong N(O)-H.O(N) hydrogen bonds. There are two possible binding geometries for the 1:1 complex 1.MMTA; combining computational and experimental evidence we demonstrate that the endo binding mode is more favorable as the result of a C-H/pi interaction.     

Noncovalent interactions within a synthetic receptor can reinforce guest binding

Structural and thermodynamic data are presented on the binding properties of anion receptors containing two covalently linked cyclopeptide subunits that bind sulfate and iodide anions with micromolar affinity in aqueous solution. A synchrotron X-ray crystal structure of the sulfate complex of one receptor revealed that the anion is bound between the peptide rings of the biscyclopeptide. Intimate intramolecular contacts between the nonpolar surfaces of the proline rings of the individual receptor moieties in the complex suggest that hydrophobic interactions within the receptor that do not directly involve the guest contribute to complex stability. This finding is supported by a microcalorimetric analysis of the solvent dependence of complex stability, which showed that increasing the water content of the solvent has only a weak influence on the Gibbs energy of binding. Hence, the increasing amount of energy required for desolvating the binding partners in solutions containing more water is almost compensated by the increasingly favorable hydrophobic interactions. Further observations that suggest that guest-induced intra-receptor interactions contribute to guest binding are (i) anion binding of a monomeric cyclopeptide lacking the covalent linkage between the two rings leads to the formation of 2:1 complexes; (ii) in the crystal structure of the 2:1 iodide complex of this monotopic receptor, a similar arrangement of the two cyclopeptide rings has been found as in the sulfate complex of the biscyclopeptide; (iii) complex formation of the monomeric cyclopeptide in aqueous solution is highly cooperative with a large stability constant corresponding to the formation of the 2:1 complexes from relatively instable 1:1 complexes; (iv) the monomeric cyclopeptide forms only 1:1 anion complexes in DMSO where hydrophobic interactions do not take place; and (v) introducing polar hydroxy groups on the proline rings of the monomeric cyclopeptide disrupts cooperativity causing the formation of only 1:1 complexes even in aqueous solution. Taken together these observations demonstrate that, in addition to direct receptor-substrate interactions, noncovalent interactions between the two subunits of such biscyclopeptides contribute significantly to anion complex stability. Reinforcement of molecular recognition through intra-receptor interactions should be an attractive new strategy to boost host-guest affinities.     

Probing the structure of DNA aptamers with a classic heterocycle

DNA aptamers are synthetic, single-stranded DNA oligonucleotides selected by SELEX methods for their binding with specific ligands. Here we present ethidium binding results for three related DNA aptamers (PDB code: 1OLD, 1DB6, and 2ARG)that bind L-argininamide (L-Arm). The ligand bound form of each aptamer's structure has been reported and each are found to be composed primarily of two domains consisting of a stem helical region and a loop domain that forms a binding pocket for the cognate ligand. Previous thermodynamic experiments demonstrated that the DNA aptamer 1OLD undergoes a large conformational ordering upon binding to L-Arm. Here we extend those linkage binding studies by examining the binding of the heterocyclic intercalator ethidium to each of the three aptamers by fluorescence and absorption spectrophotometric titrations. Our results reveal that ethidium binds to each aptamer with DeltaG degree's in the range of -8.7 to -9.4 kcal/mol. The stoichiometry of binding is 2:1 for each aptamer and is quantitatively diminished in the presence of L-Arm as is the overall fluorescence intensity of ethidium. Together, these results demonstrate that a portion of the bound ethidium is excluded from the aptamer in the presence of a saturating amount of L-Arm. These results demonstrate the utility of ethidium and related compounds for the probing of non-conventional DNA structures and reveal an interesting fundamental thermodynamic linkage in DNA aptamers. Results are discussed in the context of the thermodynamic stability and structure of each of the aptamers examined.     

Molecular dynamics study on the interaction of a mithramycin dimer with a decanucleotide duplex

The complex of a minor groove binding drug mithramycin (MTR) and the self-complementary d(TAGCTAGCTA) 10-mer duplex was investigated by molecular dynamics (MD) simulations using the AMBER 7.0 suite of programs. There is one disaccharide and trisaccharide segment projecting from opposite ends of an aglycone chromophore of MTR. A MTR dimer complex (MTR)2Mg2+ is formed in the presence of a coordinated ion Mg2+. A NMR solution structure of two (MTR)2Mg2+ complexes bound with one DNA duplex, namely, the 2:1 duplex complex, was taken as the starting structure for the MD simulation. The partial charge on each atom was calculated using the multiple-RESP fitting procedure, and all of the missing parameters in the Parm99 force field used were adapted comparably from the literature. The length of the MD simulation was 5 ns, and the binding free energy for the formation of a 1:1 or 2:1 duplex complex was determined from the last 4 ns of the simulation. The binding free energies were decomposed to components of the contributions from different energy types, and the changes in the helical parameters of the bound DNA duplex plus the glycosidic linkages between sugar residues of the bound MTR dimer were determined. It was found that binding of the first (MTR)2Mg2+ complex with the DNA duplex to form a 1:1 duplex complex does not cause stiffening of the duplex especially in the unoccupied site of the duplex. However, the overall flexibility of the DNA duplex is reduced substantially once the second (MTR)2Mg2+ complex is bound with the unoccupied site to form the 2:1 duplex complex. The van der Waals interactions were found to be dominant in the central part of the DNA duplex where sugar residues from each bound (MTR)2Mg2+ complex were inwardly pointing and the corresponding minor groove was widened.     

An exclusive fluoride receptor: fluoride-induced proton transfer to a quinoline-based thiourea

A new quinoline-based tripodal thiourea has been synthesized, which exclusively binds fluoride anion in DMSO, showing no affinity for other anions including chloride, bromide, iodide, perchlorate, nitrate, and hydrogen sulfate. As investigated by ¹H NMR, the receptor forms both 1:1 and 1:2 complexes yielding binding constants of 2.32(3) (in log β₁) and 4.39(4) (in log β₂), respectively. The quinoline groups are protonated by fluoride-induced proton transfer from the solution to the host molecule. The 1:2 binding is due to the interactions of one fluoride with NH binding sites of urea sites and another fluoride with secondary ⁺NH binding sites within the tripodal pocket. The formation of both 1:1 and 1:2 complexes has been confirmed by theoretical calculations based on density functional theory (DFT).     

Intercalation of a Heterocyclic Ligand between Quartets in a G-Rich Tetrahelical Structure

YES G-rich oligonucleotide VK2 folds into an AGCGA-quadruplex tetrahelical structure distinct and significantly different from G-quadruplexes, even though it contains four G₃ tracts. Herein, a bis-quinolinium ligand 360A with high affinity for G-quadruplex structures and selective telomerase inhibition is shown to strongly bind to VK2. Upon binding, 360A does not induce a conformational switch from VK2 to an expected G-quadruplex. In contrast, NMR structural study revealed formation of a well-defined VK2–360A complex with a 1:1 binding stoichiometry, in which 360A intercalates between GAGA- and GCGC-quartets in the central cavity of VK2. This is the first high-resolution structure of a G-quadruplex ligand intercalating into a G-rich tetrahelical fold. This unique mode of ligand binding into tetrahelical DNA architecture offers insights into the stabilization of an AGCGA-quadruplex by a heterocyclic ligand and provides guidelines for rational design of novel VK2 binding molecules with selectivity for different DNA secondary structures.     

Design and synthesis of a unique ditopic macrocyclic fluorescent receptor containing furan ring as a spacer for the recognition of dicarboxylic acids

A macrocyclic fluorescent receptor was designed and synthesised and the binding study with three different types of dicarboxylic acids was performed with the receptor being found to have appreciable association constants. Downfield shifts of specific amide protons in 1:1 binding by ¹H NMR and the quenching in the fluorescence spectra reveal strong binding and thus unambiguously support the complexation of the receptor 1 with dicarboxylic acids.     

Artificial receptors that provides a preorganized hydrophobic environment: a biomimetic approach to dopamine recognition in water

[structure: see text] The recognition of dopamine in water has been achieved with tripodal oxazoline-based artificial receptors, capable of providing a preorganized hydrophobic environment by rational design, which mimics a hydrophobic pocket predicted for a human D2 receptor. The receptors show an amphiphilic nature owing to the presence of hydrophilic sulfonate groups at the periphery of the tripodal oxazoline ligands, which seems to contribute in forming the preorganized hydrophobic environment. The artificial receptors recognized dopamine hydrochloride in water with reasonable selectivity among various organoammonium guests examined. The observed binding behavior of the receptors was explained by evoking guest inclusion in the preorganized hydrophobic pocket-like environment and not by simple ion-pairing interactions. The rationally predicted 1:1 inclusion binding mode was supported by binding studies such as with a reference receptor that cannot provide a similar binding pocket, Job and VT-NMR experiments, electrospray ionization mass analysis, and guest selectivity data. This study implies that an effective hydrophobic environment can be generated even from an acyclic, small molecular artificial receptor. Such a preorganized hydrophobic environment, as being utilized in biological systems, can be effectively used as a complementary binding force for the recognition of organoammonium guests such as dopamine hydrochloride in water.     

Chloride binding by a polyimidazolium macrocycle detected via fluorescence,NMR, and X-ray crystallography

Herein, we describe a macrocyclic polyimidazolium receptor that is preorganized for the binding of anionic guests, and particularly chloride. Additionally, diphenylimidazolium units were incorporated into this structure to enhance photophysical properties that were exploited for signal transduction of binding. In subsequent fluorescence binding studies, this receptor was found to bind a range of halides as well as phosphate with high affinity (K_a=1.8×10⁴, and 1.5×10⁴ for phosphate and chloride, respectively) in a competitive solvent mixture (1:1 water/acetonitrile). Results under these conditions were fitted to 1:1 binding curves, and indicated modest selectivity of the host for phosphate and chloride over other halides. Binding studies were also performed using ¹H NMR spectroscopy, during which the imidazolium C–H signal was observed to shift downfield upon titration with anions. These experiments were run in less polar solvent (1:9 water/acetonitrile), and could not be fitted to a 1:1 binding curve, suggesting higher order aggregates in this environment. Binding was further probed in the solid state by obtaining an X-ray crystal structure of receptor–iodide complex. In the resulting structure, two iodides were found to bind through interactions with two polyimidazolium hydrogens each. These results show that the described macrocycle is effective for anion-binding in competitive solvent, with modest selectivity for chloride over other halides, and that the nature of the binding interactions varies depending upon the solvent environment.     

Efficient synthesis and anion recognition of a colorimetric preorganized tripodal thiourea compound

A preorganized colorimetric tripodal thiourea receptor was synthesized in high yield utilizing a thiol-ene reaction as a primary step. The interaction of the receptor with dihydrogen phosphate, acetate, chloride, and fluoride anions was investigated using UV–vis. and ¹H NMR spectroscopic titration techniques. The binding stoichiometry of the receptor with dihydrogen phosphate and acetate was found to be 1:2, and 1:1 with chloride and fluoride. The binding constants for the receptor and dihydrogen phosphate, acetate, chloride, and fluoride were determined using HypNMR2008 and HypSpec.     

Synthesis and Evaluation of a Cyclophane Receptor for Acetic Acid

A bicyclic cyclophane ( 2 ) containing one pyridine nitrogen and four amide N-H groups oriented toward the interior of the cavity was synthesized. The binding constants of various carboxylic acids with 2 were measured by UV/Vis spectroscopy. Acetic acid bound to 2 with a K a of 980 - 90 M m 1 in chloroform while branched carboxylic acids showed significantly lower binding. The data indicate that acetic acid was bound within the cavity of 2 . Only one acetic acid binds to two control hosts, whereas 2 shows definitive 1:1 binding. The results suggest that selectivity in the binding of carboxylic acids can be achieved via size constraints dictated by the receptor cavity, and that the same size restrictions lead to only one carboxylic acid bound to the cyclophane. The crystal structure of 2 is reported.     

Polyoxometalate binding to human serum albumin: a thermodynamic and spectroscopic approach

The molecular recognition of polyoxometalates by human serum albumin is studied using two different polyoxometalates (POMs) at pH 7.5. The results are compared with those obtained at pH 3.5 and 9.0. At pH 7.5, both POMs strongly interact with the protein with different binding behaviors. The Keggin shaped POM, [H(2)W(12)O(40)](6-) (H2W12), specifically binds the protein, forming a complex with a 1:1 stoichiometry with Ka = 2.9 x 10(6) M(-1). The binding constant decreased dramatically with the increase of the ionic strength, thus indicating a mostly electrostatic binding process. Isothermal titration calorimetry (ITC) experiments show that the binding is an enthalpically driven exothermic process. For the wheel shaped POM [NaP(5)W(30)O(110)](14-) (P5W30), there are up to five binding sites on the protein. Increasing the ionic strength changes the binding behavior significantly, leading to a simple exothermic process, with several binding sites. Competitive binding experiments indicate that the two POMs share one common binding site. In addition, they show the existence of another important binding site for P5W30. The two POMs exhibit different binding dependences on the pH. The combination of the experimental results with the knowledge of the surface map of the protein in its N-B conformation transition domain leads to the proposal for the probable binding site of POMs. The present work reveals a protein conformation change upon P5W30 binding, a new feature not explicitly documented in previous studies.     

Self-assembly of a hydrophobic groove

Heptakis(2,6-di-O-methyl)-β-cyclodextrin interacts with 5,15-diphenylporphine to produce a 2:1 complex in dimethyl sulfoxide. This complex possesses a hydrophobic groove that circumscribes the metal binding site of the porphyrin moiety.