A hypothesis to account for the selective and diverse actions of neonicotinoid insecticides at their molecular targets,nicotinic acetylcholine receptors: catch and release in hydrogen bond networks

The low mammalian toxicity of neonicotinoid insecticides has been shown to be attributable, at least in part, to their selective actions on insect nicotinic acetylcholine receptors (nAChRs). There are multiple nAChRs in insects and a wealth of neonicotinoid chemicals. Studies to date have discribed a wide range of effects on nAChRs, notably partial agonist, super agonist and antagonist actions. Both the diversity of the neonicotinoid actions and their selectivity for insect over vertebrate nAChRs are the result of physicochemical and steric interactions at their molecular targets (nAChRs). In such interactions, the formation and breakage of hydrogen bond (HB) networks plays a key role. Therefore the loss or gain of even a single HB resulting from either structural changes in neonicotinoids, or the amino acid sequence of a particular nAChR subunit, could result in a drastic modification of neonicotinoid actions. In addition to the amino acid residues, the backbone carbonyl of nAChRs may also be involved in the formation of HB networks with neonicotinoids.     

The pore domain of the nicotinic acetylcholine receptor: molecular modeling, pore dimensions, and electrostatics.

The pore domain of the nicotinic acetylcholine receptor has been modeled as a bundle of five kinked M2 helices. Models were generated via molecular dynamics simulations incorporating restraints derived from 9-A resolution cryoelectron microscopy data (Unwin, 1993; 1995), and from mutagenesis data that identify channel-lining side chains. Thus, these models conform to current experimental data but will require revision as higher resolution data become available. Models of the open and closed states of a homopentameric alpha 7 pore are compared. The minimum radius of the closed-state model is less than 2 A; the minimum radius of the open-state models is approximately 6 A. It is suggested that the presence of "bound" water molecules within the pore may reduce the effective minimum radii below these values by up to approximately 3 A. Poisson-Boltzmann calculations are used to obtain a first approximation to the potential energy of a monovalent cation as it moves along the pore axis. The differences in electrostatic potential energy profiles between the open-state models of alpha 7 and of a mutant of alpha 7 are consistent with the experimentally observed change in ion selectivity from cationic to anionic. Models of the open state of the heteropentameric Torpedo nicotinic acetylcholine receptor pore domain are also described. Relatively small differences in pore radius and electrostatic potential energy profiles are seen when the Torpedo and alpha 7 models are compared.     

Interactions between tachykinins and diverse,human nicotinic acetylcholine receptor subtypes

Nicotinic acetylcholine receptors (nAChR) are diverse members of the ligand-gated ion channel superfamily of neurotransmitter receptors and play critical roles in chemical signaling throughout the nervous system. Reports of effects of substance P (SP) on nAChR function prompted us to investigate interactions between several tachykinins and human nAChR subtypes using clonal cell lines as simple experimental models. Acute exposure to SP inhibits carbamylcholine- or nicotinestimulated function measured using⁸⁶Rb⁺ efflux assays of human ganglionic (α3β4) nAChR expressed in SH-SY5Y neuroblastoma cells (IC₅₀∼2.3 μM) or of human muscle-type (α1β1γδ) nAChR expressed in TE671/RD clonal cells (IC₅₀∼21 μM). SP also acutely blocks function of rat ganglionic nAChR expressed in PC12 pheochromocytoma cells (IC₅₀∼2.1 μM). Neurokinin A and eledoisin inhibit function (extrapolated IC₅₀ values between 60 and 160 μM) of human muscle-type or ganglionic nAChR, but neurokinin B does not, and neither human nAChR is as sensitive as PC12 cell α3β4-nAChR to eledoisin or neurokinin A inhibition. At concentrations that produce blockade of nAChR function, SP fails to affect binding of [³H]acetylcholine to human muscle-type or ganglionic nAChR. SP-mediated blockade of rat or human ganglionic nAChR function is insurmountable by increasing agonist concentrations. Collectively, these results indicate that tachykinins act noncompetitively to inhibit human nAChR function with potencies that vary across tachykinins and nAChR subtypes. They also indicate that tachykinin actions at nAChR could further contribute to complex cross-talk between nicotinic cholinergic and tachykinin signals in regulation of nervous system activity.     

Pyrazolone-fused combretastatins and their precursors: synthesis,cytotoxicity, antitubulin activity and molecular modeling studies

A series of pyrazolone-fused combretastatins and precursors were synthesized and their cytotoxicity as well as antitubulin potential was evaluated. The hydrazide 9f and the pyrazolone-fused combretastatins 12a, 12b and 12c were highly cytotoxic against various tumor cell lines including cisplatin resistant cells. The same compounds were also the best inhibitors of tubulin polymerization. Molecular modeling results showed that they bind the colchicine binding site at the tubulin heterodimer. The hydrazide 9f arrested HeLa cells in the G2/M phase of the cell cycle and strongly affected cell shape and microtubule network.     

Synthesis, binding studies and molecular modeling of novel cannabinoid receptor ligands

In the present work, we report upon the design, synthesis and biological evaluation of new anandamide derivatives obtained by modifications of the fatty acyl chain and/or of the ethanolamide 'tail'. The compounds are of the general formula: 6-(substituted-phenyl)/naphthyl-4-oxohex-5-enoic acid N-substituted amide and 7-naphthyl-5-oxohept-6-enoicacid N-substituted amide. The novel compounds had been evaluated for their binding affinity to CB1/CB2 cannabinoid receptors, binding studies showed that some of the newly developed compounds have measurable affinity and selectivity for the CB2 receptor. Compounds XI and XVIII showed the highest binding affinity for CB2 receptor. None of the compounds exhibited inhibitory activity towards anandamide hydrolysis, thus arguing in favor of their enzymatic stability. The structure-activity relationship has been extensively studied through a tailor-made homological model using constrained docking in addition to pharmacophore analysis, both feature and field based.     

Cytotoxic ribonucleases: molecular weapons and their targets

Many ribonucleases (RNases) are highly cytotoxic. In some cases, they attack selectively malignant cells, triggering apoptotic response, and therefore are considered as alternative chemotherapeutic drugs. Factors that determine the cytotoxicity of RNases, primarily of those of microbial origin, are reviewed here. These factors include catalytic activity, ability to escape natural inhibitors, stability, and efficiency of internalization. The latter is, in turn, determined by positive charge on the molecule and interaction with cell membrane. Cellular targets and molecular determinants of RNases decisive for their cytotoxic action are characterized.     

The alpha7 nicotinic acetylcholine receptor: molecular modelling, electrostatics, and energetics

The structure of a homopentameric alpha7 nicotinic acetylcholine receptor is modelled by combining structural information from two sources: the X-ray structure of a water soluble acetylcholine binding protein from Lymnea stagnalis, and the electron microscopy derived structure of the transmembrane domain of the Torpedo nicotinic receptor. The alpha7 nicotinic receptor model is generated by simultaneously optimising: (i) chain connectivity, (ii) avoidance of stereochemically unfavourable contacts, and (iii) contact between the beta1-beta2 and M2-M3 loops that have been suggested to be involved in transmission of conformational change between the extracellular and transmembrane domains. A Gaussian network model was used to predict patterns of residue mobility in the alpha7 model. The results of these calculations suggested a flexibility gradient along the transmembrane domain, with the extracellular end of the domain more flexible that the intracellular end. Poisson-Boltzmann (PB) energy calculations and atomistic (molecular dynamics) simulations were used to estimate the free energy profile of a Na+ ion as a function of position along the axis of the pore-lining M2 helix bundle of the transmembrane domain. Both types of calculation suggested a significant energy barrier to exist in the centre of the (closed) pore, consistent with a "hydrophobic gating" model. Estimations of the PB energy profile as a function of ionic strength suggest a role of the extracellular domain in determining the cation selectivity of the alpha7 nicotinic receptor. These studies illustrate how molecular models of members of the nicotinic receptor superfamily of channels may be used to study structure-function relationships.     

Sex steroids and their receptors: molecular actions on brain cells

Sex steroids exert actions of paramount importance on brain cells. They contribute to shape the central nervous system during embryo development. They modulate the formation and the turnover of the interconnections between neurons. They control the function of glial cells. And they do it through a signaling machinery that is apparently simple, but that hides a level of complexity that has been unveiled only in part. Different receptor isoforms, different interactions between receptors and co-regulators, chains of events originating at the cell membrane and leading to effects in the nucleus (or the other way around) all interact to determine selective modulations of brain cells. All these actions end up in phenomenal effects on brain function that change through adolescence, pregnancy, adulthood, up to menopause and ageing. Many of these actions are relevant for degenerative processes and research may offer soon new strategies to counteract these diseases.     

Homology modeling and molecular dynamics simulations of transmembrane domain structure of human neuronal nicotinic acetylcholine receptor

A three-dimensional model of the transmembrane domain of a neuronal-type nicotinic acetylcholine receptor (nAChR), (alpha4)2(beta2)3, was constructed from a homology structure of the muscle-type nAChR recently determined by cryo-electron microscopy. The neuronal channel model was embedded in a fully hydrated DMPC lipid bilayer, and molecular-dynamics simulations were performed for 5 ns. A comparative analysis of the neuronal- versus muscle-type nAChR models revealed many conserved pore-lining residues, but an important difference was found near the periplasmic mouth of the pore. A flickering salt-bridge of alpha4-E266 with its adjacent beta2-K260 was observed in the neuronal-type channel during the course of the molecular-dynamics simulations. The narrowest region, with a pore radius of approximately 2 A formed by the salt-bridges, does not seem to be the restriction site for a continuous water passage. Instead, two hydrophobic rings, formed by alpha4-V259, alpha4-L263, and the homologous residues in the beta2-subunits, act as the gates for water flow, even though the region has a slightly larger pore radius. The model offers new insight into the water transport across the (alpha4)2(beta2)3 nAChR channel, and may lead to a better understanding of the structures, dynamics, and functions of this family of ion channels.     

Tea polyphenols, their biological effects and potential molecular targets

Tea is the most popular beverage in the world, second only to water. Tea contains an infusion of the leaves from the Camellia sinensis plant rich in polyphenolic compounds known as catechins, the most abundant of which is (-)-EGCG. Although tea has been consumed for centuries, it has only recently been studied extensively as a health-promoting beverage that may act to prevent a number of chronic diseases and cancers. The results of several investigations indicate that green tea consumption may be of modest benefit in reducing the plasma concentration of cholesterol and preventing atherosclerosis. Additionally, the cancer-preventive effects of green tea are widely supported by results from epidemiological, cell culture, animal and clinical studies. In vitro cell culture studies show that tea polyphenols potently induce apoptotic cell death and cell cycle arrest in tumor cells but not in their normal cell counterparts. Green tea polyphenols were shown to affect several biological pathways, including growth factor-mediated pathway, the mitogen-activated protein (MAP) kinase-dependent pathway, and ubiquitin/proteasome degradation pathways. Various animal studies have revealed that treatment with green tea inhibits tumor incidence and multiplicity in different organ sites such as skin, lung, liver, stomach, mammary gland and colon. Recently, phase I and II clinical trials have been conducted to explore the anticancer effects of green tea in humans. A major challenge of cancer prevention is to integrate new molecular findings into clinical practice. Therefore, identification of more molecular targets and biomarkers for tea polyphenols is essential for improving the design of green tea trials and will greatly assist in a better understanding of the mechanisms underlying its anti-cancer activity.     

Molecular docking,molecular modeling,and molecular dynamics studies of azaisoflavone as dual COX-2 inhibitors and TP receptor antagonists

Designed multi-target ligand (DML) is an emerging strategy for the development of new drugs and involves the engagement of multiple targets with the same moiety. In the context of NSAIDs it has been suggested that targeting the thromboxane prostanoid (TP) receptor along with cyclooxygenase-2 (COX-2) may help to overcome cardiovascular (CVS) complications associated with COXIBs. In the present work, azaisoflavones were studied for their COX-2 and TP receptor binding activities using structure based drug design (SBDD) techniques. Flavonoids were selected as a starting point based on their known COX-2 inhibitory and TP receptor antagonist activity. Iterative design and docking studies resulted in the evolution of a new class scaffold replacing the benzopyran-4-one ring of flavonoids with quinolin-4-one. The docking and binding parameters of these new compounds are found to be promising in comparison to those of selective COX-2 inhibitors, such as SC-558 and celecoxib. Owing to the lack of structural information, a model for the TP receptor was generated using a threading base alignment method with loop optimization performed using an ab initio method. The model generated was validated against known antagonists for TP receptor using docking/MMGBSA. Finally, the molecules that were designed for selective COX-2 inhibition were docked into the active site of the TP receptor. Iterative structural modifications and docking on these molecules generated a series which displays optimum docking scores and binding interaction for both targets. Molecular dynamics studies on a known TP receptor antagonist and a designed molecule show that both molecules remain in contact with protein throughout the simulation and interact in similar binding modes.

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Synthesis,antibacterial studies,and molecular modeling studies of 3,4-dihydropyrimidinone compounds

The syntheses of dihydropyrimidinones (DHPMs) using solvent-free grindstone chemistry method. All the synthesized compounds exhibited significant activity against pathogenic bacteria. The current effort has been developed to obtain new DHPM derivatives that focus on the bacterial ribosomal A site RNA as a drug target. Molecular docking simulation analysis was applied to confirm the target specificity of DHPMs. The crystal structure of bacterial 16S rRNA and human 40S rRNA was taken as receptors for docking. Finally, the docking score, binding site interaction analysis revealed that DHPMs exhibit more specificity towards 16S rRNA than known antibiotic amikacin. Accordingly, targeting the bacterial ribosomal A site RNA with potential drug leads promises to overcome the bacterial drug resistance. Even though, anti-neoplastic activities of DHPMs were also predicted through prediction of activity spectra for substances (PASS) tool. Further, the results establish that the DHPMs can serve as perfect leads against bacterial drug resistance.     

Synthesis,biological activity and multiscale molecular modeling studies for coumaryl-carboxamide derivatives as selective carbonic anhydrase IX inhibitors

New coumaryl-carboxamide derivatives with the thiourea moiety as a linker between the alkyl chains and/or the heterocycle nucleus were synthesized and their inhibitory activity against the human carbonic anhydrase (hCA) isoforms hCA I, II, VII and IX were evaluated. While the hCA I, II and VII isoforms were not inhibited by the investigated compounds, the tumour-associated isoform hCA IX was inhibited in the high nanomolar range. 2-Oxo-N-((2-(pyrrolidin-1-yl)ethyl)carbamothioyl)-2H-chromene-3-carboxamide (e11) exhibited a selective inhibitory action against hCA IX with the K_i of 107.9?nM. In order to better understand the inhibitory profiles of studied molecules, multiscale molecular modeling approaches were used. Different molecular docking algorithms were used to investigate binding poses and predicted binding energies of studied compounds at the active sites of the CA I, II, VII and IX isoforms.     

N-haloacetylimino neonicotinoids: potency and molecular recognition at the insect nicotinic receptor

This structure-activity relationship study for neonicotinoids with an N-haloacetylimino pharmacophore identifies several candidate compounds showing outstanding insecticidal potency and consequently leads to establishing their molecular recognition at an insect nicotinic receptor structural model, wherein the neonicotinoid halogen atoms (fluorine, chlorine, bromine, and iodine) variously interact with the receptor loops C-D interfacial niche via H-bonding and/or hydrophobic interactions.     

A structural biology perspective on bioactive small molecules and their plant targets

Structural biology efforts in recent years have generated numerous co-crystal structures of bioactive small molecules interacting with their plant targets. These studies include the targets of various phytohormones, pathogen-derived effectors, herbicides and other bioactive compounds. Here we discuss that this collection of structures contains excellent examples of nine collective observations: molecular glues, allostery, inhibitors, molecular mimicry, promiscuous binding sites, unexpected electron densities, natural selection at atomic resolution, and applications in structure-guided mutagenesis and small molecule design.     

Acetaminophen interacts with human hemoglobin: optical,physical and molecular modeling studies

Acetaminophen, a widely used analgesic and antipyretic drug has ample affinity to bind globular proteins. Here, we have illustrated a substantive study pertaining to the interaction of acetaminophen with human hemoglobin (HHb). Different spectroscopic (absorption, fluorescence, and circular dichroism (CD) spectroscopy), calorimetric, and molecular docking techniques have been employed in this study. Acetaminophen-induced graded alterations in absorbance and fluorescence of HHb confirm their interaction. Analysis of fluorescence quenching at different temperature and data obtained from isothermal titration calorimetry indicate that the interaction is static and the HHb has one binding site for the drug. The negative values of Gibbs energy change (ΔG⁰) and enthalpy changes (ΔH⁰) and positive value of entropy change (ΔS⁰) strongly suggest that it is entropy-driven spontaneous and exothermic reaction. The reaction involves hydrophobic pocket of the protein which is further stabilized by hydrogen bonding as evidenced from ANS and sucrose binding studies. These findings were also supported by molecular docking simulation study using AutoDock 4.2. The interaction influences structural integrity as well as functional properties of HHb as evidenced by CD spectroscopy, increased rate of co-oxidation and decreased esterase activity of HHb. So, from these findings, we may conclude that acetaminophen interacts with HHb through hydrophobic and hydrogen bonding, and the interaction perturbs the structural and functional properties of HHb.     

3D-QSAR, homology modeling, and molecular docking studies on spiropiperidines analogues as agonists of nociceptin/orphanin FQ receptor

The nociceptin/orphanin FQ receptor (NOP) has been implicated in a wide range of biological functions, including pain, anxiety, depression and drug abuse. Especially, its agonists have a great potential to be developed into anxiolytics. However, the crystal structure of NOP is still not available. In the present work, both structure-based and ligand-based modeling methods have been used to achieve a comprehensive understanding on 67N-substituted spiropiperidine analogues as NOP agonists. The comparative molecular-field analysis method was performed to formulate a reasonable 3D-QSAR model (cross-validated coefficient q(2)=0.819 and conventional r(2)=0.950), whose robustness and predictability were further verified by leave-eight-out, Y-randomization, and external test-set validations. The excellent performance of CoMFA to the affinity differences among these compounds was attributed to the contributions of electrostatic/hydrogen-bonding and steric/hydrophobic interactions, which was supported by the Surflex-Dock and CDOCKER molecular-docking simulations based on the 3D model of NOP built by the homology modeling method. The CoMFA contour maps and the molecular docking simulations were integrated to propose a binding mode for the spiropiperidine analogues at the binding site of NOP.     

Insulin receptor binding kinetics: modeling and simulation studies

Biological actions of insulin regulate glucose metabolism and other essential physiological functions. Binding of insulin to its cell surface receptor initiates signal transduction pathways that mediate cellular responses. Thus, it is of great interest to understand the mechanisms underlying insulin receptor binding kinetics. Interestingly, negative cooperative interactions are observed at high insulin concentrations while positive cooperativity may be present at low insulin concentrations. Clearly, insulin receptor binding kinetics cannot be simply explained by a classical bimolecular reaction. Mature insulin receptors have a dimeric structure capable of binding two molecules of insulin. The binding affinity of the receptor for the second insulin molecule is significantly lower than for the first bound insulin molecule. In addition, insulin receptor aggregation occurs in response to ligand binding and aggregation may also influence binding kinetics. In this study, we develop a mathematical model for insulin receptor binding kinetics that explicitly represents the divalent nature of the insulin receptor and incorporates receptor aggregation into the kinetic model. Model parameters are based upon published data where available. Computer simulations with our model are capable of reproducing both negative and positive cooperativity at the appropriate insulin concentrations. This model may be a useful tool for helping to understand the mechanisms underlying insulin receptor binding and the coupling of receptor binding to downstream signaling events.