Presence Of Competitive Ligands Research Articles

Effective removal of manganese in graphene oxide via competitive ligands and the properties of reduced graphene oxide hydrogels and films

Reduced graphene oxide (rGO) has tremendous importance due to its potential applications in electronics, catalysis and sensors. However, the properties of rGO are often affected by the impurity of manganese ions, and their removal from graphene oxide (GO) is time and labor-consuming due to their strong interaction with oxygen-containing groups of GO. In this paper, we use citric acid, sulfosalicylic acid and tartaric acid as competitive ligands to make manganese ions removal easy. The results show that under the same purifying conditions, the presence of competitive ligands can increase the purifying efficiency, and the lowest content of manganese is only one-third of that in GO without competitive ligands. The hydrogels and films of rGO have been prepared, and their properties such as capacitance and conductivity have also been evaluated. The results show that the microstructure of hydrogels is affected by the added ligands. The hydrogels prepared from GO which is purified in the presence of sulfosalicylic acid exhibit well-defined pore structure and the capacitive performance such as rate capability and the cycling stability is the best. It is proved that adding competitive ligands is an effective approach to remove manganese impurity in GO.

Synthesis and Structure of [М(DMF)6][B10H10] (M = Zn(II), Cd(II)) as Precursors for Solid-Phase Synthesis of Trischelate Complexes [М(L)3][B10H10

The reactivity of the decahydro-closo-decaborate anion in the complexation of metals being among intermediates acids in the concept of the Pearson’s theory of hard and soft acids and bases is studied systematically in the presence of competitive ligands L. In continuing these studies, we have synthesized Zn(II) and Cd(II) complexes of the composition [М(DMF)6][B10H10] (M = Zn(II), Cd(II)) and studied them by physicochemical analyses (IR and UV spectroscopy, X-ray powder diffraction). The possibility of their use in solid-phase synthesis of the corresponding trischelate complexes [M(Bipy)3][B10H10] has been found. Complexes [М(DMF)6][B10H10] are formed by the reaction between triethylammonium closo-decaborate with metal nitrates in DMF. The compounds are isostructural with the corresponding Co(II) and Ni(II) complexes [М(DMF)6][B10H10]. Specific interactions have been found between the BH groups of the boron clusters and CH groups of the DMF molecules. According to the data of IR and UV spectroscopy, the complexes contain specific B–H···C–H interactions. The corresponding tris-chelate complexes of the metals [M(Bipy)3][B10H10] are also isostructural.

Competitive ligands facilitate dissociation of the complex of bifunctional inhibitor and protein kinase

Dissociation of the complex of a ligand and a protein usually follows the kinetic profile of the first order process and the rate of dissociation is not affected by the presence of competitive ligands. We discovered that dissociation of the complex between a bifunctional ligand and a protein kinase (the catalytic subunit of cAMP-dependent protein kinase), an enzyme possessing 2 different substrate binding sites, was accelerated (facilitated) over 50-fold in the presence of competitive ligands at higher concentrations. Structurally diverse compounds revealed >10-fold different efficiency for acceleration of dissociation of the complex. These results show that the kinetic behavior of flexible biomolecular complexes possessing two spatially separated contact areas is highly dynamic. This property of biomolecular complexes should be carefully considered for effective application of bifunctional ligands for regulation of activity of target proteins in cells.

Preliminary in vitro and in vivo evaluation of liposomal nanoparticles for passive and active tumour targeting by scintigraphic and MRI imaging.

Liposomes belong to the most desirable drug delivery systems that enable their application in MRI and SPECT by incorporating lipids chelated to MR contrast agents as well as to radioactive tracers. Here we report our observation on a novel αvβ3- targeting liposome formulation (RGD-LP) and on the corresponding non-targeting control (NT-LP) for diagnosis and molecular imaging of tumor by exploiting SPECT/MRI technology. Radiolabelling was performed with 99mTc tricarbonyl core via the ligand cysteine (Cys). Both peptide and ligand were coupled to DSPE-PEG2000-Mal lipid. Radiochemical purity and in vitro stability tested in the presence of competitive ligands for 99mTc(I) were assessed by paper chromatography. For nuclear imaging a custom SPECT system (1.5mm spatial resolution) was used and the results were correlated to the biodistribution data. Both liposomes disposed of a PEG surface density of 5.5% and were loaded with 25% of Gd-based lipid (B2286) for MRI studies. For recording MR images, cells were transferred into glass capillaries, centrifuged and placed in an agar phantom. MR images were acquired on a Bruker Avance300 spectrometer operating at 7.1 T. In vivo MRI experiments were carried out on U87MG tumor xenografts with a 1T scanner (Bruker, Icon Instrument). Radiolabelling yield (>80%), radiochemical purity (>90%), in vitro stability in the presence of excess amount of cysteine and in 10% FBS/DMEM were satisfactory to permit in vivo evaluation at normal mice. NT-LP-Cys showed higher blood retention and lower uptake at liver and spleen at later time points than RGD-LP which is in good correlation with the in vivo MRI results obtained in U87MG bearing mice. The internalization amount of Gd3+ inside the cells was of higher extent for RGD-LP at all the time points analyzed. These results could be considered as an important step in the development of tumour targeted SPECT/MRI contrast agents for cancer imaging.

Competitive displacement of cofilin can promote actin filament severing

Cofilin is an essential actin filament severing protein that functions in the dynamic remodeling of the actin cytoskeleton. Filament severing activity is most efficient at sub-stoichiometric cofilin binding densities (i.e. <1cofilinperactin filament subunit), and peaks when the number density of boundaries (i.e. junctions) between bare and cofilin-decorated segments is maximal. A model in which local topological and mechanical discontinuities lead to preferential fragmentation at boundaries accounts for available experimental data, including direct visualization of cofilin and actin during real-time severing events. The boundary-severing model predicts that ligands (e.g. other actin-binding proteins) that compete with cofilin for actin filament binding and modulate cofilin occupancy on filaments will alter the bare-decorated segment boundary density, and thus, the filament severing activity of cofilin. Here, we directly test this model prediction by evaluating the effects of phalloidin and myosin, two ligands that compete with cofilin for filament binding, on the actin filament binding and severing activities of cofilin. Our experiments demonstrate that competitive displacement of cofilin lowers cofilin occupancy and promotes severing when initial cofilin occupancy is high (i.e. >50%). Even in the presence of competitive ligands, maximum severing activity occurs when cofilin-decorated boundary density is highest, consistent with preferential fragmentation at boundaries. We propose a general “severodyne” framework for the modulation of cofilin-mediated actin filament severing by small molecule or actin-binding protein ligands that compete with cofilin for actin filament binding.

Validation of Alexa-647-ATP as a powerful tool to study P2X receptor ligand binding and desensitization

Ion channel opening and desensitization is a fundamental process in neurotransmission. The ATP-gated P2X1 receptor (P2X1R) shows rapid and long-lasting desensitization upon agonist binding. This makes the electrophysiological investigation of its desensitization process, agonist unbinding, and recovery from desensitization a challenging task. Here, we show that the fluorescent agonist Alexa-647-ATP is a potent agonist at the P2X1R and a versatile tool to directly visualize agonist binding and unbinding. We demonstrate that the long-lasting desensitization of the P2X1R is due to both slow unbinding of agonist from the desensitized receptor and agonist mediated receptor internalization. Furthermore, the unbinding of the agonist Alexa-647-ATP from the desensitized receptor is accelerated in the continuous presence of competitive ligand. Modeling of our data indicates that three agonist molecules are required to drive the receptor into desensitization. Direct visualization of ligand unbinding from the desensitized receptor demonstrates the cooperativity of this process.

Exploring avidity: understanding the potential gains in functional affinity and target residence time of bivalent and heterobivalent ligands

Bivalent ligands are increasingly important therapeutic agents. Although the naturally occurring antibodies are predominant, it is becoming more common to combine different antibody fragments or even low molecular weight compounds to generate heterobivalent ligands. Such ligands exhibit markedly increased affinity (i.e. avidity) and target residence time when both pharmacophores can bind simultaneously to their target sites. This is because binding of one pharmacophore forces the second tethered one to stay close to its corresponding site. This 'forced proximity' favours its binding and rebinding (once dissociated) to that site. However, rebinding will also take place when the diffusion of freshly dissociated ligands is merely slowed down. The present differential equation-based simulations explore the way both situations affect ligand binding. Both delay the attainment of binding equilibrium (resulting in steep saturation curves) and also increase the target residence time. Competitive ligands are able to interfere in a concentration-dependent manner, although much higher concentrations are required in the 'forced proximity' situation. Also, it is only in that situation that the ligand shows increased affinity. These simulations shed light on two practical consequences. Depending on the pharmacokinetic half-life of the bivalent ligand in the body, it may not have sufficient time to achieve equilibrium with the target. This will result in lower potency than expected, although it would have significant advantages in terms of residence time. In in vitro experiments, the manifestation of steep saturation curves and of accelerated dissociation in the presence of competitive ligands could mistakenly be interpreted as evidence for non-competitive, allosteric interactions.

Radiolabeling of PAMAM dendrimers conjugated to a pyridine-N-oxide DOTA analog with 111In: Optimization of reaction conditions and biodistribution

Polyamidoamine dendrimers (PAMAMs) of generations 1 (G1) and 4 (G4) were conjugated with a bifunctional pyridine-N-oxide DOTA analog, 10-[(4-carboxy-1-oxidopyridin-2-yl)methyl]-1,4,7,10-tetraazacyclododecane-1,4,7-triacetic acid (H4do3a-pyNO−C), through the pyridine-4-carboxylic acid group, and the conjugates were radiolabeled with indium-111. Reaction conditions for the radiolabelling were optimized. Both radiolabeled conjugates, G1-[111In(do3a-pyNO−C)] and G4-[111In(do3a-pyNO−C)], were kinetically stable for at least 48h after preparation; in the presence of competitive ligands, the radiochemical purity of the conjugates slightly decreased (4–7%) over the same time period. The preclinical pharmacokinetics of both agents were evaluated. Biodistribution and elimination in rats were more favorable for the G1-[111In(do3a-pyNO−C)] conjugate than G4-[111In(do3a-pyNO−C)] conjugate. However, the G1-[111In(do3a-pyNO−C)] conjugate was rapidly eliminated from the body, mainly through urine, while, significant and long-term radioactivity uptake in the liver and kidney was observed for the G4-[111In(do3a-pyNO−C)] conjugate.

Noni as an anxiolytic and sedative: A mechanism involving its gamma-aminobutyric acidergic effects

Noni (Morinda citrifolia) is increasing in worldwide popularity as a food or dietary supplement with versatile health benefits. The aim of this study was to investigate the effects of Noni fruit on anxiety symptoms in vitro. To this end, a competitive GABAa receptor-binding assay was developed. Our preliminary study indicates that the methanol crude extract of Noni fruit showed significant affinity to the gamma-aminobutyric acid A (GABAa) inhibitory neurotransmitter receptors, and displayed 75% binding inhibition of the agonist radioligand [3H] muscimol at a concentration of 100 microg/ml. Further experiments demonstrated that the MeOH extract, and its BuOH and H2O partitions, exhibited IC50 values of 22.8, 27.2, and 17.1 microg/ml, respectively, in the GABAa-binding assay. Experimental results with Noni fruit indicate the presence of competitive ligand(s), which may bind to the GABAa receptor as an agonist, and thus induce its anxiolytic and sedative effects. The study provides an in vitro rationale for one of Noni's versatile and traditional uses. In addition, an HPLC fingerprint profile of the methanolic extract of Noni fruit has been established for quality control purpose.

Electrochemical behaviour of iron-complexes in presence of competitive ligands: A strategy for optimization of current density

Triethanolamine and D-gluconate both form alkali stable complexes with iron(III)- and iron(II)-ions. The redox behaviour of solutions containing one type of ligand is compared with solutions containing both ligands in excess compared with the iron(III)-concentration. Spectrophotometric investigations and experiments with cyclic voltammetry yield information about follow-up reactions proceeding during the cathodic reduction of the iron(III)-triethanolamine complex when D-gluconate is present in addition. The results indicate that the Fe(III)-triethanolamine complex is the predominant species as long as c(triethanolamine) exceeds c(Fe(III)) and thus the cathodic current density is mainly determined by the concentration of this complex. When the concentration of triethanolamine is lowered to 20% of the iron concentration then iron(III)–D-gluconate is the major species present in the solution. Then, besides ligand exchange reactions, an increase in cathodic current density indicates a redox reaction between Fe(II)-triethanolamine and Fe(III)–D-gluconate similar to the Ecat-type in cyclic voltammetry. Such conditions are of particular interest for the optimization of mediator systems with regard to the current density attainable at given iron(III)-concentration. The results show that a combination of different ligands can lead to an addition of the specific advantages of each metal-complex mediator.