Based on the specific folic acid-folate receptor (FA-FR) interaction, macromolecular FR can bind with FA-linked DNA-small molecule chimeras, which can prevent enzymolysis by exonuclease III (Exo III), enabling a novel fluorescence biosensor for FR to be developed using quinaldine red as a G-quadruplex-binding probe.

Hydroxycinnamic acid derivatives (HCAs) are a group of naturally occurring polyphenolic compounds which possess various pharmacological activities. In this work, the interactions of bovine serum albumin (BSA) with six HCA derivatives, including chlorogenic acid (CHA), caffeic acid (CFA), m-coumaric acid (m-CA), p-coumaric acid (p-CA), ferulic acid (FA) and sinapic acid ...

Our group previously developed a multifunctional, targeted cancer therapeutic based on Generation 5 (G5) polyamidoamine (PAMAM) dendrimers. In those studies we conjugated the targeting molecule folic acid (FA) and the chemotherapeutic drug methotrexate (MTX) sequentially. This complex macromolecule was shown to selectively bind and kill KB tumor cells that over-express ...

Theophylline, popular diuretic, is used to treat asthma and bronchospasm. In blood it forms complexes with albumin, which is also the main transporter of fatty acids. The aim of the present study was to describe the influence of fatty acids (FA) on binding of theophylline (Th) to human serum albumin ...

High-risk strains of human papillomaviruses (HPVs) cause nearly all cases of cervical cancer as well as a growing number of head and neck cancers. The oncogenicity of these viruses can be attributed to the activities of their two primary oncoproteins, E6 and E7. The E6 protein has among its functions ...

Neuronal nicotinic acetylcholine receptor (nAChR) agonists active at the alpha-7 (α-7) receptor subtype are potential therapeutics for cognitive deficits in schizophrenia, Alzheimer's disease and other mental disorders. SSR180711, an α-7 selective partial agonist, has been shown to improve preclinical cognition. A novel PET radioligand, (11) C-Chiba1001, is a close analogue ...

A series of pyrazoloquinoline analogs have been synthesized and shown to bind to PDE10 with high affinity. From the SAR study and our lead optimization efforts, compounds 16 and 27 were found to possess potent oral antipsychotic activity in the MK-801 induced hyperactive rat model.

Disrupted in Schizophrenia-1 (DISC1) plays essential roles in neuronal proliferation, neuronal migration and axon guidance, and has been implicated in schizophrenia. DISC1 forms a functional complex with Nuclear Distribution Element Like protein-1 (NDEL1), a key component that regulates microtubule organization during cell division and neuronal migration. DISC1 mutations at the ...

Cognitive functioning is based on binding processes, by which different features and elements of neurocognition are integrated and coordinated. Binding is an essential ingredient of, for instance, Gestalt perception. We have implemented a paradigm of causality perception based on the work of Albert Michotte, in which 2 identical discs move ...

Dopamine D2 receptor homodimers might be of particular importance in the pathophysiology of schizophrenia and, thus, serve as promising target proteins for the discovery of atypical antipsychotics. A highly attractive approach to investigate and control GPCR dimerization may be provided by the exploration and characterization of bivalent ligands, which can ...

The β3-adrenegic receptor (β3-AR) selectivity over β1- and β2-ARs has been the most important aspect for successful therapeutic agents for obesity and type-II diabetes, as the concomitant activation of β1- and β2-ARs would lead to undesirable side effects such as increased heart rate. In order to explore the structural basis ...

Conventional determination of agonist efficacy at G-protein coupled receptors is measured by stimulation of guanosine-5'-γ-thiotriphosphate (GTPγS) binding. We analysed the role of guanosine diphosphate (GDP) in the process of activation of the M₂ muscarinic acetylcholine receptor and provide evidence that negative cooperativity between agonist and GDP binding is an alternative ...

Curcumin is a well-known component of the cook seasoning and traditional herb turmeric (Curcuma longa), which has been reported to prevent obesity. However, the mechanism still remains to be determined. In this study, curcumin is found to be an effective inhibitor of fatty acid synthase (FAS), and its effects on ...

The binding of orally administered imidafenacin, used to treat overactive bladders, to muscarinic receptors in rat tissue was characterized based on pharmacokinetics. The binding in six tissues including bladder tissue was measured using [N-methyl-(3)H] scopolamine methyl chloride ([(3)H]NMS). Pharmacokinetic parameters were estimated from measurements of the concentration of imidafenacin in ...

Activation of M1 muscarinic receptors occurs through orthosteric and allosteric binding sites. To identify critical residues, site-directed mutagenesis and chimeric receptors were evaluated in functional calcium mobilization assays to compare orthosteric agonists, acetylcholine and xanomeline, M1 allosteric agonists AC-42 (4-n-butyl-1-[4-(2-methylphenyl)-4-oxo-1-butyl]-piperidine hydrogen chloride), TBPB (1-[1'-(2-methylbenzyl)-1,4'-bipiperidin-4-yl]-1,3-dihydro-2H-benzimidazol-2-one), and the clozapine metabolite N-desmethylclozapine. A ...

Postoperative morbidity of pancreaticoduodenectomy remains high and is mainly related to postoperative pancreatic fistula. Peng et al. (J Gastrointest Surg 2003;7:898-900; Am J Surg 2002;183:283-285; Ann Surg 2007;245:692-298) recently described binding pancreaticojejunostomy and reported a zero percent rate of pancreatic fistula. The aim of this study was to compare postoperative ...

In the search for new chemical entities that interact with G-proteincoupled receptors (GPCRs), assays that quantify efficacy and affinity are employed. Traditional methods for measuring affinity involve radiolabeled ligands. To address the need for homogeneous biochemical fluorescent assays to characterize orthosteric ligand affinity and dissociation rates, we have developed a ...

We characterized contribution of N-oxide metabolites [1-methyl-4-piperidyl diphenylpropoxyacetate N-oxide (M-1) and 1-methyl-4-piperidyl benzilate N-oxide (M-2)] to the binding of muscarinic receptors in relation to the pharmacokinetics of propiverine in rats. The in vitro muscarinic receptor binding activity of M-2 was equipotent to that of propiverine, whereas M-1 was much less ...

N-desmethylclozapine (NDMC) has been reported to display partial agonism at the human recombinant and rat native M(1) mAChR, a property suggested to contribute to the clinical efficacy of clozapine. However, the profile of action of NDMC at the human native M(1) mAChR has not been reported. The effect of NDMC ...

Muscarinic acetylcholine receptors (mAChRs) of rat cerebral cortex were evaluated using a tissue segment radioligand binding assay. [(3)H]-Quinuclidinyl benzilate (QNB, a hydrophobic ligand) specifically bound to mAChRs in the cortex segments. The total mAChRs level was approximately 2,000 fmol/mg protein, which was estimated after incubation for 120 min at 37 ...

The present study was undertaken to examine whether distigmine, a therapeutic agent used to treat detrusor underactivity, binds directly to muscarinic and nicotinic receptors. We used radioreceptor binding assays and compared the effects of distigmine with those of neostigmine and donepedil. The inhibitory effect of distigmine on the blood acetylcholinesterase ...

Muscarinic acetylcholine (M) and adrenergic (AR) receptors mediate gastrointestinal motility. Using radioligand binding assays and real-time polymerase chain reaction, the densities of binding sites and mRNA levels of M₂, M₃, α₂(AD)- and β₂-AR were compared in muscle tissues from the abomasal fundus, pylorus, duodenum, caecum, and external loop of the ...

Ligand binding to G protein-coupled receptors is a complex process that involves sequential receptor conformational changes, ligand translocation, and possibly ligand-induced receptor oligomerization. Binding events at muscarinic acetylcholine receptors are usually interpreted from radioligand binding studies in terms of two-step ligand-induced receptor isomerization. We report here, using a combination of ...

Muscarinic acetylcholine receptor (mAChR)-mediated guanine nucleotide-binding regulatory protein (G protein) activation and the functional interaction between receptors and the respective G proteins were investigated using an agonist-induced [(35)S]guanosine-5'-O-(gamma-thiotriphosphate) ([(35)S]GTPgammaS)-binding approach in membranes of 3 native equine airway segments (trachea, bronchus and lung), which differ tremendously in mAChR density and subtype ...

Point mutations and molecular modeling have been used to study the activation of the M(1) muscarinic acetylcholine receptor (mAChR) by the functionally selective agonists 4-n-butyl-1-[4-(2-methylphenyl)-4-oxo-1-butyl]-piperidine (AC-42), and 1-[3-(4-butyl-1-piperidinyl)propyl]-3,4-dihydro-2(1H)-quinolinone (77-LH-28-1), comparing them with N-desmethylclozapine (NDMC) and acetylcholine (ACh). Unlike NDMC and ACh, the activities of AC-42 and 77-LH-28-1 were undiminished by ...

A library of robust ghrelin receptor mutants with single substitutions at 22 positions in the main ligand-binding pocket was employed to map binding sites for six different agonists: two peptides (the 28-amino-acid octanoylated endogenous ligand ghrelin and the hexapeptide growth hormone secretagogue GHRP-6) plus four nonpeptide agonists-the original benzolactam L-692,429 ...

We showed previously that RGS8 directly binds to the third intracellular loop (i3L) of the M1 muscarinic acetylcholine receptor using the sequence MPRR at the N-terminus of RGS8 and specifically inhibits signal transduction. Here, we identified spinophilin (SPL) as an RGS8-interacting protein. We found that the SPL-binding site of RGS8 ...

Decreased muscarinic M1 receptor (CHRM1) mRNA has been reported in Brodmann's area (BA) 6 from subjects with schizophrenia. We have extended this study by measuring levels of CHRM1 ([(3)H]pirenzepine binding), CHRM3 ([(3)H]4-DAMP binding), the transcription factor SP1 and the CHRM1 downstream target beta-site APP-cleaving enzyme 1 (BACE1) in BA 6 ...

Many G protein-coupled receptors (GPCRs) possess allosteric binding sites distinct from the orthosteric site utilized by their cognate ligands, but most GPCR allosteric modulators reported to date lack signaling efficacy in their own right. McN-A-343 (4-(N-(3-chlorophenyl)carbamoyloxy)-2-butynyltrimethylammonium chloride) is a functionally selective muscarinic acetylcholine receptor (mAChR) partial agonist that can also ...

Studies with 4-[3-[(1,1-dimethylethyl)amino]2-hydroxypropoxy]-1,3-dihydro-2H-benzimidazol-2-one hydrochloride (CGP 12177) at the human beta1-adrenoceptor have provided evidence for two binding modes or conformations that have markedly different pharmacological properties. Here, key transmembrane residues (Asp104, Asp138, Ser228, Ser229, Ser232, Phe341, Asn344 and Asn363) have been mutated to provide structural insights into the nature of these ...

We explored the interaction of a nitrogen mustard derivative of acetylcholine with the human M(2) muscarinic receptor expressed in Chinese hamster ovary cells using the muscarinic radioligand, [3H]N-methylscopolamine (NMS). Acetylcholine mustard caused a concentration-dependent, first-order loss of [3H]NMS binding at 37 degrees C, with the half-maximal rate constant occurring at ...

The M4 muscarinic acetylcholine (ACh) receptor (mAChR) is a potential therapeutic target but characterized by a lack of subtype-selective ligands. We recently generated "designer receptors exclusively activated by a designer drug" (DREADDs), which contained mutations of two conserved orthosteric-site residues (Y113C/A203G in the M4 mAChR) that caused a loss of ...

The M(2) muscarinic receptor has two topographically distinct sites: the orthosteric site and an allosteric site recognized by compounds such as gallamine. It also can exhibit cooperative effects in the binding of orthosteric ligands, presumably to the orthosteric sites within an oligomer. Such effects would be difficult to interpret, however, ...

BACKGROUND AND PURPOSE: In human airways, muscarinic acetylcholine receptors (mAChRs) exert a predominant role in the control of airways resistance and anti-muscarinic agents are currently included in the pharmacological treatment of chronic obstructive pulmonary disease (COPD). However, the development of more effective mAChR antagonists is hampered by considerable species variability ...

GPCRs (G-protein-coupled receptors) exist in a spontaneous equilibrium between active and inactive conformations that are stabilized by agonists and inverse agonists respectively. Because ligand binding of agonists and inverse agonists often occurs in a competitive manner, one can assume an overlap between both binding sites. Only a few studies report ...

Using a selective Galpha(q/11) protein antibody capture guanosine 5'-O-(3-[35S]thio)triphosphate ([35S]GTPgammaS) binding approach, it has been possible to perform a quantitative pharmacological examination of the functional activity of the M(1) muscarinic acetylcholine receptor (mAChR) in membranes prepared from human postmortem cerebral cortex. Oxotremorine-M caused a > or = 2-fold increase in ...

We measured the influence of gallamine on the functional responses and binding properties of selected agonists at the M(2) muscarinic receptor and analyzed the data within the context of the allosteric ternary complex model. Our analysis showed that gallamine modified agonist affinity without influencing efficacy. To explain this behavior, we ...

In the presence of neostigmine (0.1 microM), S-isopetasin competitively antagonized cumulative acetylcholine-induced contractions in guinea pig trachealis, because the slope [1.18+/-0.15 (n=6)] of Schild's plot did not significantly differ from unity. The pA2 value of S-isopetasin was calculated to be 4.62+/-0.05 (n=18). The receptor binding assay for muscarinic receptors of ...

A group of small proteins, designated as muscarinic toxins (MTs), have been isolated from the venom of African green mamba (Dendroaspis angusticeps) and documented to bind selectively to individual muscarinic acetylcholine receptor (mAChR) subtypes. These components have less been reported to be isolated from other snake venoms. In this study, ...

The study presents a docking analysis for the interaction capabilities of some muscarinic receptors (i.e., M(1), M(2), and M(5)) whose full-length models were previously generated by us. In detail, the docking simulations involved a dataset of 30 agonists, taken from the literature, including a first series of oxathiolane/dioxolane congeners and ...

Two stereoselective parallel divergent four-step procedures to obtain all four enantiomeric forms of N,N,N-trimethyl(6-methyl-1,4-dioxan-2-yl)methanaminium iodide were developed. Enantiomeric purity was determined by quantitative (1)H NMR spectroscopy in the presence of the chiral shift reagent (+)-MTPA. The biological profile of the obtained compounds was evaluated at all muscarinic receptor subtypes by ...

It is almost 60 years since Heinz Schild devised a method that allowed measurement of a genuine physical quantity, the equilibrium constant for binding of a competitive antagonist. The clever bit was that the measurements could be made using responses from tissues despite the fact that little or nothing was ...

In 2005, the first evidence was obtained that the cannabinoid CB(1) receptor contains an allosteric binding site. The site can be recognized by synthetic small molecules, which display a markedly divergent effect on orthosteric ligand affinity versus efficacy; these small molecules are allosteric enhancers of agonist binding affinity and allosteric ...

A series of xanomeline analogs were synthesized and evaluated for binding at the M(1) muscarinic acetylcholine receptor (M(1) receptor). Specifically, compounds that substitute the O-hexyl chain of xanomeline with polar, ionizable, or conformationally restricted moieties were assessed for their ability to bind to the M(1) receptor in a wash-resistant manner ...

Gallamine and tacrine are allosteric antagonists at muscarinic M2 acetylcholine receptors and inhibitors of acetylcholinesterase. At both acetylcholine-binding proteins, gallamine and tacrine are known to occupy two different binding sites: in M2 receptors within the allosteric binding area and in acetylcholinesterase at its catalytic and its peripheral site. To find ...

In recurrent airway obstruction (RAO) or heaves, bronchospasm has been attributed to enhanced cholinergic activity. However, the expression and function of muscarinic acetylcholine receptors (mAChR) and their signaling components are not yet known. Thus, we examined the expression, subtype distribution and postreceptor signaling pathways of mAChR in the peripheral lung, ...

RGS2 and RGS5 are inhibitors of G-protein signaling belonging to the R4/B subfamily of RGS proteins. We here show that RGS2 is a much more potent attenuator of M1 muscarinic receptor signaling than RGS5. We hypothesize that this difference is mediated by variation in their ability to constitutively associate with ...

Alanine substitution mutagenesis has been used to investigate residues that make up the roof and floor of the muscarinic binding pocket and regulate ligand access. We mutated the amino acids in the second extracellular loop of the M1 muscarinic acetylcholine receptor that are homologous to the cis-retinal contact residues in ...

Ala substitution scanning mutagenesis has been used to probe the functional role of amino acids in transmembrane (TM) domain 2 of the M1 muscarinic acetylcholine receptor, and of the highly conserved Asn43 in TM1. The mutation of Asn43, Asn61, and Leu64 caused an enhanced ACh affinity phenotype. Interpreted using a ...

Muscarinic acetylcholine receptors (mAChRs) are prototypical Family A G protein coupled-receptors. The five mAChR subtypes are widespread throughout the periphery and the central nervous system and, accordingly, are widely involved in a variety of both physiological and pathophysiological processes. There currently remains an unmet need for better therapeutic agents that ...