Point Mutations of Nicotinic Receptor α1 Subunit Reveal New Molecular Features of G153S Slow-Channel Myasthenia

Slow-channel congenital myasthenic syndromes (SCCMSs) are rare genetic diseases caused by mutations in muscle nicotinic acetylcholine receptor (nAChR) subunits. Most of the known SCCMS-associated mutations localize at the transmembrane region near the ion pore. Only two SCCMS point mutations are at the extracellular domains near the acetylcholine binding site, α1(G153S) being one of them. In this work, a combination of molecular dynamics, targeted mutagenesis, fluorescent Ca²⁺ imaging and patch-clamp electrophysiology has been applied to G153S mutant muscle nAChR to investigate the role of hydrogen bonds formed by Ser 153 with C-loop residues near the acetylcholine-binding site. Introduction of L199T mutation to the C-loop in the vicinity of Ser 153 changed hydrogen bonds distribution, decreased acetylcholine potency (EC₅₀ 2607 vs. 146 nM) of the double mutant and decay kinetics of acetylcholine-evoked cytoplasmic Ca²⁺ rise (τ 14.2 ± 0.3 vs. 34.0 ± 0.4 s). These results shed light on molecular mechanisms of nAChR activation-desensitization and on the involvement of such mechanisms in channelopathy genesis.     

A computational model of the nicotinic acetylcholine binding site

We have derived a model of the nicotinic acetylcholine binding site. This was accomplished by using three known agonists (acetylcholine, nicotine and epibatidine) as templates around which polypeptide side chains, found to be part of the receptor cavity from published molecular biology studies, are allowed to flow freely in molecular dynamics simulations and mold themselves around these templates. The resulting supramolecular complex should thus be a complement, both in terms of steric effects as well as electronic effects, to the agonists and it should be a good estimation of the true receptor cavity structure. The shapes of those minireceptor cavities equilibrated rapidly on the simulation time scale and their structural congruence is very high, implying that a satisfactory model of the nicotinic acetylcholine binding site has been achieved. The computational methodology was internally tested against two rigid and specific antagonists (dihydro--erytroidine and erysoidine), that are expected to give rise to a somewhat differently shaped binding site compared to that derived from the agonists. Using these antagonists as templates there were structural reorganizations of the initial receptor cavities leading to distinctly different cavities compared to agonists. This indicates that adequate times and temperatures were used in our computational protocols to achieve equilibrium structures for the agonists. Overall, both minireceptor geometries for agonists and antagonists are similar with the exception of one amino acid (ARG209).     

First synthesis of (±)-vertilecanin A

5-[(Hydroxy)(phenyl)methyl]pyridine-2-carboxylic acid ((±)-vertilecanin A) was prepared from nicotinic acid in four steps with an overall yield of 29%. Published in Russian in Izvestiya Akademii Nauk. Seriya Khimicheskaya, No. 10, pp. 2389–2390, October     

MENTHOR,a database system for the storage and retrieval of three-dimensional molecular structures and associated data searchable by substructural,biologic, physical,or geometric properties

Summary MENTHOR is a database system for the storage and retrieval of three-dimensional coordinate and charge information on molecules as well as of traditional biological and physical properties. Our molecular graphics system retrieves from MENTHOR structural information in individual molecules and receptor map/macromolecular binding site hypotheses. Substructural searches of MENTHOR are used to find starting coordinates for molecular modeling and traditional database searches of MENTHOR identify compounds for which modeling is needed. It also forms the data to be searched with ALLADDIN, our substructure/geometric search program. MENTHOR expedites molecular modeling by organizing previous work and facilitating transmission of information between individuals. Examples from modeling of D-2 receptor agonists are shown.     

Agonist–PPARγ interactions: Molecular modeling study with docking approach

Docking simulation of 18 agonists with the ligand binding pocket (LBP) of PPARγ has been performed. The binding conformations and binding affinities of these agonists were obtained by use of the flexible docking protocol FlexX. Test compound calculations indicated that FlexX can reproduce the binding conformation of the crystal structure (root mean square deviation = 1.43 Å); moreover, the predicted binding affinities correlate well with the activities of these agonists. The interaction model and pharmacophore of PPARγ agonists were derived and the difference in biologic activities of these agonists can be well explained. The PPARγ agonists must have both polar head and the hydrophobic tail, which form hydrogen bonds and hydrophobic contacts with hydrophilic and hydrophobic regions of the LBP of PPARγ, respectively. In addition, a suitable linker is also necessary. © 2003 Wiley Periodicals, Inc. Int J Quantum Chem 93: 405–410, 2003     

An improved nicotinic pharmacophore and a stereoselective CoMFA-model for nicotinic agonists acting at the central nicotinic acetylcholine receptors labelled by [3H]-N-methylcarbamylcholine

A study of a series of compounds with agonistic effect at the alpha4beta2 nicotinic acetylcholine receptors resulted in an improved pharmacophore model as well as a CoMFA model. The pharmacophore was composed of three pharmacophoric elements: (1) a site point (a) corresponding to a protonated nitrogen atom, (2) a site point (b) corresponding to an electronegative atom capable of forming a hydrogen bond, and (3) the centre of a heteroaromatic ring or a C=O bond (c). The pharmacophoric elements were related by the following parameters: (a-b) 7.3-8.0 A, (a-c) 6.5-7.4 A, and the angle between the two distance vectors (delta bac) 30.4-35.8 degrees. In addition to this, a stereoselective CoMFA model was developed, which showed good predictability even for compound classes not present in the training set.     

3 D-QSAR Analysis of Agonists of nAChRs： Epibatidine Analogues

A 3 D-QSAR about nAChRs agonists epibatidine analogues was performed using theCoMFA and CoMSIA. The correlation coefficients were R2cv = 0.546, R2cv = 0.907 in CoMFA andR2cv = 0.655, R2,~ = 0.962 in CoMSIA of the final model. The prediction using the final models tothe test set was r2 = 0.675 in CoMFA and r2 = 0.462 in CoMSIA. This model will be useful in thedesign of novel compounds with high affinity.     

Synthesis and Crystal Structure of Tetraimidazolium Dinicotinatooctamolybdate

1INTRODUCTIONPolyoxometallates(POM)areofgreatinterestnotonlyfortheirstructuralvarietybutalsotheirunexpectedproperties[1,2].Theseinorganicanionshavebeenwidelyinvestigatedasdiversascatalysis,biology,medicine,magnetochemistryandmaterialscience[3～7].Thestudiesonthecoordinatedpoly-oxometallateswithorganicligandsmighthelptounderstandtherolesofpolyoxometallatesoncatalysis.Todate,aseriesofoctamolybdateswhichcanbedescribedas[Mo8O26X2]2n－4(nisthenormalchargeofthecoordinatedbaseX)havebeenreport…     

Feed additives in animal nutrition: Quantification of a new adrenergic drug by hyphenated techniques

Zilpaterol, (±)‐trans‐4,5,6,7‐tetrahydro‐7‐hydroxy‐6‐(isopropylamino) imidazo [4,5,1‐jk]‐[1]benzazepin‐2(1H)‐one, is an adrenergic agonist drug licensed in Mexico and South Africa as feed additive for cattle at slaughter age. In contrast, the EU legislative framework prohibits the use of beta‐agonists as growth promoters to produce lean meat. In this study, the compound was quantified as its TMS derivative both by gas chromatography‐mass spectrometry (GC‐MS) with a quadrupole detector (QUADR) and by GC‐MS/MS with an ion trap detector (ITD), in Electron Impact (EI) ionization. Subsequently, an extraction and clean‐up procedure for the identification of Zilpaterol in commercial feeds was set up. Different extraction solvents and matrices were considered for their influence on Zilpaterol yield. The analytical method led to good recoveries and limits of detection and quantification in feeds well below the 5–20 μg/g dose proposed for animal nutrition.