From ab initio quantum mechanics to molecular neurobiology: A cation–π binding site in the nicotinic receptor

The nicotinic acetylcholine receptor is the prototype ligand-gated ion channel. A number of aromatic amino acids have been identified as contributing to the agonist binding site, suggesting that cation–π interactions may be involved in binding the quaternary ammonium group of the agonist, acetylcholine. Here we show a compelling correlation between: (i) ab initio quantum mechanical predictions of cation–π binding abilities and (ii) EC50 values for acetylcholine at the receptor for a series of tryptophan derivatives that were incorporated into the receptor by using the in vivo nonsense-suppression method for unnatural amino acid incorporation. Such a correlation is seen at one, and only one, of the aromatic residues—tryptophan-149 of the α subunit. This finding indicates that, on binding, the cationic, quaternary ammonium group of acetylcholine makes van der Waals contact with the indole side chain of α tryptophan-149, providing the most precise structural information to date on this receptor. Consistent with this model, a tethered quaternary ammonium group emanating from position α149 produces a constitutively active receptor.

Altered recognition mutants of the response regulator PhoB: A new genetic strategy for studying protein–protein interactions

Two-component regulatory systems require highly specific interactions between histidine kinase (transmitter) and response regulator (receiver) proteins. We have developed a novel genetic strategy that is based on tightly regulated synthesis of a given protein to identify domains and residues of an interacting protein that are critical for interactions between them. Using a reporter strain synthesizing the nonpartner kinase VanS under tight arabinose control and carrying a promoter-lacZ fusion activated by phospho-PhoB, we isolated altered recognition (AR) mutants of PhoB showing enhanced activation (phosphorylation) by VanS as arabinose-dependent Lac+ mutants. Changes in the PhoBAR mutants cluster in a “patch” near the proposed helix 4 of PhoB based on the CheY crystal structure (a homolog of the PhoB receiver domain) providing further evidence that helix 4 lies in the kinase-regulator interface. Based on the CheY structure, one mutant has an additional change in a region that may propagate a conformational change to helix 4. The overall genetic strategy described here may also be useful for studying interactions of other components of the vancomycin resistance and Pi signal transduction pathways, other two-component regulatory systems, and other interacting proteins. Conditionally replicative oriRR6Kγ attP “genome targeting” suicide plasmids carrying mutagenized phoB coding regions were integrated into the chromosome of a reporter strain to create mutant libraries; plasmids encoding mutant PhoB proteins were subsequently retrieved by P1-Int-Xis cloning. Finally, the use of similar genome targeting plasmids and P1-Int-Xis cloning should be generally useful for constructing genomic libraries from a wide array of organisms.

Case-only study of interactions between genetic polymorphisms of GSTM1, P1, T1 and Z1 and smoking in Parkinson's disease

Current opinion contends that complex interactions between genetic and environmental factors play a role in the etiology of Parkinson's disease (PD). Cigarette smoking is thought to reduce risk of PD, and emerging evidence suggests that genetic factors may modulate smoking's effect. We used a case-only design, an approach not previously used to study gene-environment interactions in PD, specifically to study interactions between glutathione-S-transferase (GST) gene polymorphisms and smoking in relation to PD. Four-hundred PD cases (age at onset: 60.0 +/- 10.7 years) were genotyped for common polymorphisms in GSTM1, PI, T1 and Z1 using well-established methods. Smoking exposure data were collected in face-to-face interviews. The independence of the studied GST genotypes and smoking exposure was confirmed by studying 402 healthy, aged individuals. No differences were observed in the distributions of GSTM1, T1 or Z1 polymorphisms between ever-smoked and never-smoked PD cases using logistic regression (all P > 0.43). However, GSTP1 *C haplotypes were over-represented among PD cases who ever smoked (odds ratio for interaction (ORi) = 2.00 (95% Cl: 1.11-3.60, P = 0.03)). Analysis revealed that ORi between smoking and the GSTP1-114Val carrier status increased with increasing smoking dose (P = 0.02 for trend). These data suggest that one or more GSTP1 polymorphisms may interact with cigarette smoking to influence the risk for PD. (C) 2004 Elsevier Ireland Ltd. All rights reserved.

The adsorption of phenolic and organotin compounds by clays and cation exchanged clays

Quaternary ammonium exchanged laponites (Quat-laponites) show selectivity in the adsorption of phenols and chlorinated phenols. Strong adsorbate-adsorbent interactions are indicated by adsorption isotherms. Adsorption of phenols and chlorinated phenols by Quat-smectites is greater than that by the Bi Quat-Smectites prepared in this study. It is thought that the quaternary ammonium exchanged smectite components of the Bi Quat-smectites interact with each other (adsorbent-adsorbent interactions) reducing the number of sites available for adsorbate-adsorbent interactions. Solidification/stabilisation studies of 2-chlorophenol show that a blend of ground granulated blast furnace slag and ordinary Portland cement attenuates 2-chlorophenol more effectively than ordinary Portland cement alone. Tetramethyl ammonium- (TMA-) and tetramethyl phosphonium- (TMP-) montmorillonites were exposed to solutions of phenol or chlorinated phenols. TMP- montmorillonite was the better adsorbent and preferentially adsorbed 4-chlorophenol over phenol. Hydration of the interlayer cations occurs to a greater extent in the TMA-montmorillonite than the TMP-montmorillonite restricting interlayer adsorption. Contrary to that observed for phenols and chlorinated phenols, the Quat-smectites were ineffective as adsorbents for triphenyltin hydroxide and bis(tributyltin) oxide at room temperature. Under microwave conditions, only bis(tributyltin) oxide was adsorbed by the quaternary ammonium exchanged smectites. Bis(tributyltin) oxide was adsorbed from ethanol on the surface of the smectite clays at room temperature and under microwave conditions. The adsorbate-adsorbent interactions were weak. Adsorption is accompanied by conversion of bis(tributyltin) oxide to a different tin(IV) species and the release of sodium cations from the montmorillonite interlayer region. Attempts to introduce conditions suitable for charge transfer interactions between synthesised quaternary ammonium compounds and 2,4,6-trichlorophenol are documented. Transition metal complex exchanged clays adsorb 2,4,6-trichlorophenol and phenol. Strong adsorbate-adsorbent interactions (Type I isotherms) occur when the adsorbate is 2,4,6-trichlorophenol and when the adsorbent is [Fe(bipy)3]2+ exchanged montmorillonite or [Co(bipy)3]3+ exchanged montmorillonite. The 2,2'-bipyridyl ligands of the adsorbents are electron rich and the 2,4,6-trichlorophenol is electron deficient. This may have enhanced adsorbate-adsorbent interactions.

Interactions of different arsenic species with thols: Chemical and biological implications

Arsenic has been classified as a group I carcinogen. It has been ranked number one in the CERCLA priority list of hazardous substances due to its frequency, toxicity and potential for human exposure. Paradoxically, arsenic has been employed as a successful chemotherapeutic agent for acute promyelocytic leukemia and has found some success in multiple myeloma. Since arsenic toxicity and efficacy is species dependent, a speciation method, based on the complementary use of reverse phase and cation exchange chromatography, was developed. Inductively coupled plasma mass spectrometer (ICP-MS), as an element specific detector, and electrospray ionization mass spectrometer (ESI-MS), as a molecule specific detector, were employed. Low detection limits in the µg. L−1 range on the ICP-MS and mg. L−1 range on the ESI-MS were obtained. The developed methods were validated against each other through the use of a Deming plot. With the developed speciation method, the effects of both pH on the stability of As species and reduced glutathione (GSH) concentration on the formation and stability of arsenic glutathione complexes were studied. To identify arsenicals in multiple myeloma (MM) cell lines post arsenic trioxide (ATO) and darinaparsin (DAR) incubation, an extraction method based on the use of ultrasonic probe was developed. Extraction tools and solvents were evaluated and the effect of GSH concentration on the quantitation of arsenic glutathione (As-GSH) complexes in MM cell extracts was studied. The developed method was employed for the identification of metabolites in DAR incubated cell lines where the effect of extraction pH, DAR incubation concentration and incubation time on the relative distribution of the As metabolites was assessed. A new arsenic species, dimethyarsinothioyl glutathione (DMMTA V-GS), a pentavalent thiolated arsenical, was identified in the cell extracts through the use of liquid chromatography tandem mass spectrometry. The formation of the new metabolite in the extracts was dependent on the decomposition of s-dimethylarsino glutathione (DMA(GS)). These results have major implications in both the medical and toxicological fields of As because they involve the metabolism of a chemotherapeutic agent and the role sulfur compounds play in this mechanism.

Renal Drug Transporters – Clinical Significance and Role in Drug-Drug Interactions

Thesis (Ph.D.)--University of Washington, 2016-07

Probing the interactions between ionic liquids and water: experimental and quantum chemical approach

For an adequate choice or design of ionic liquids, the knowledge of their interaction with other solutes and solvents is an essential feature for predicting the reactivity and selectivity of systems involving these compounds. In this work, the activity coefficient of water in several imidazolium-based ionic liquids with the common cation 1-butyl-3-methylimidazolium was measured at 298.2 K. To contribute to a deeper insight into the interaction between ionic liquids and water, COSMO-RS was used to predict the activity coefficient of water in the studied ionic liquids along with the excess enthalpies. The results showed good agreement between experimental and predicted activity coefficient of water in ionic liquids and that the interaction of water and ionic liquids was strongly influenced by the hydrogen bonding of the anion with water. Accordingly, the intensity of interaction of the anions with water can be ranked as the following: [CF3SO3](-) < [SCN](-) < [TFA](-) < Br(-) < [TOS](-) < Cl(-) < [CH3SO3](-) [DMP](-) < [Ac](-). In addition, fluorination and aromatization of anions are shown to reduce their interaction with water. The effect of temperature on the activity coefficient of water at infinite dilution was measured by inverse gas chromatography and predicted by COSMO-RS. Further analysis based on COSMO-RS provided information on the nature of hydrogen bonding between water and anion as well as the possibility of anion-water complex formation.

Understanding the cation specific effects on the aqueous solubility of amino acids: from mono to polyvalent cations

The interactions established by mono and polyvalent cations in natural media have important implications on the structure formation, function and physico-chemical behavior of biomolecules, playing therefore a critical role in biochemical processes. In order to further elucidate the molecular phenomena behind the cation specific effects in biological environments, and clarify the influence of the charge of the ions, solubility measurements and molecular dynamics simulations were performed for aqueous solutions of three amino acids (alanine, valine and isoleucine), in the presence of a series of inorganic salts comprising mono-, di- and trivalent cations (LiCl, Li2SO4, K2SO4, CaCl2, AlCl3 and Al-2(SO4)(3)). The evidence gathered indicates that the mechanism by which (salting-in inducing) polyvalent cations affect the solubility of amino acids in aqueous solutions is different from that of monovalent cations. A consistent and refined molecular description of the effect of the cation on the solubility of amino acids based on specific interactions of the cations with the negatively charged moieties of the biomolecules is here proposed.

Interactions of pyridinium, pyrrolidinium or piperidinium based ionic liquids with water: measurements and COSMO-RS modelling

Looking for a better knowledge concerning water and ionic liquids (ILs) interactions, a systematic study of the activity coefficients of water in pyridinium, pyrrolidinium and piperidinium-based ILs at 298.2 K is here presented based on water activity measurements. Additionally, the study of the structural effects of the pyridinium-based cation is also pursued. The results show that non-aromatic ILs are interacting more with water than aromatic ones, and among the ortho, meta and para isomers of 1-butyl-methylpyridinium chloride, the ortho position confers a more hydrophilic character to that specific IL. The physicalchemistry of the solutions was interpreted based on dissociation constants, natural bond orbitals and excess enthalpies providing a sound basis for the interpretation of the experimental observations. These results show that hydrogen bonding controls the behavior of these systems, being the anion-water one of the most relevant interactions, but modulated by the anionecation interactions.

Iis--integrated Interactome System: A Web-based Platform For The Annotation, Analysis And Visualization Of Protein-metabolite-gene-drug Interactions By Integrating A Variety Of Data Sources And Tools.

High-throughput screening of physical, genetic and chemical-genetic interactions brings important perspectives in the Systems Biology field, as the analysis of these interactions provides new insights into protein/gene function, cellular metabolic variations and the validation of therapeutic targets and drug design. However, such analysis depends on a pipeline connecting different tools that can automatically integrate data from diverse sources and result in a more comprehensive dataset that can be properly interpreted. We describe here the Integrated Interactome System (IIS), an integrative platform with a web-based interface for the annotation, analysis and visualization of the interaction profiles of proteins/genes, metabolites and drugs of interest. IIS works in four connected modules: (i) Submission module, which receives raw data derived from Sanger sequencing (e.g. two-hybrid system); (ii) Search module, which enables the user to search for the processed reads to be assembled into contigs/singlets, or for lists of proteins/genes, metabolites and drugs of interest, and add them to the project; (iii) Annotation module, which assigns annotations from several databases for the contigs/singlets or lists of proteins/genes, generating tables with automatic annotation that can be manually curated; and (iv) Interactome module, which maps the contigs/singlets or the uploaded lists to entries in our integrated database, building networks that gather novel identified interactions, protein and metabolite expression/concentration levels, subcellular localization and computed topological metrics, GO biological processes and KEGG pathways enrichment. This module generates a XGMML file that can be imported into Cytoscape or be visualized directly on the web. We have developed IIS by the integration of diverse databases following the need of appropriate tools for a systematic analysis of physical, genetic and chemical-genetic interactions. IIS was validated with yeast two-hybrid, proteomics and metabolomics datasets, but it is also extendable to other datasets. IIS is freely available online at: http://www.lge.ibi.unicamp.br/lnbio/IIS/.

Ant-caterpillar Antagonism At The Community Level: Interhabitat Variation Of Tritrophic Interactions In A Neotropical Savanna.

Ant foraging on foliage can substantially affect how phytophagous insects use host plants and represents a high predation risk for caterpillars, which are important folivores. Ant-plant-herbivore interactions are especially pervasive in cerrado savanna due to continuous ant visitation to liquid food sources on foliage (extrafloral nectaries, insect honeydew). While searching for liquid rewards on plants, aggressive ants frequently attack or kill insect herbivores, decreasing their numbers. Because ants vary in diet and aggressiveness, their effect on herbivores also varies. Additionally, the differential occurrence of ant attractants (plant and insect exudates) on foliage produces variable levels of ant foraging within local floras and among localities. Here, we investigate how variation of ant communities and of traits among host plant species (presence or absence of ant attractants) can change the effect of carnivores (predatory ants) on herbivore communities (caterpillars) in a cerrado savanna landscape. We sampled caterpillars and foliage-foraging ants in four cerrado localities (70-460 km apart). We found that: (i) caterpillar infestation was negatively related with ant visitation to plants; (ii) this relationship depended on local ant abundance and species composition, and on local preference by ants for plants with liquid attractants; (iii) this was not related to local plant richness or plant size; (iv) the relationship between the presence of ant attractants and caterpillar abundance varied among sites from negative to neutral; and (v) caterpillars feeding on plants with ant attractants are more resistant to ant predation than those feeding on plants lacking attractants. Liquid food on foliage mediates host plant quality for lepidopterans by promoting generalized ant-caterpillar antagonism. Our study in cerrado shows that the negative effects of generalist predatory ants on herbivores are detectable at a community level, affecting patterns of abundance and host plant use by lepidopterans. The magnitude of ant-induced effects on caterpillar occurrence across the cerrado landscape may depend on how ants use plants locally and how they respond to liquid food on plants at different habitats. This study enhances the relevance of plant-ant and ant-herbivore interactions in cerrado and highlights the importance of a tritrophic perspective in this ant-rich environment.

Contrasting Effects Of Land Use Intensity And Exotic Host Plants On The Specialization Of Interactions In Plant-herbivore Networks.

Human land use tends to decrease the diversity of native plant species and facilitate the invasion and establishment of exotic ones. Such changes in land use and plant community composition usually have negative impacts on the assemblages of native herbivorous insects. Highly specialized herbivores are expected to be especially sensitive to land use intensification and the presence of exotic plant species because they are neither capable of consuming alternative plant species of the native flora nor exotic plant species. Therefore, higher levels of land use intensity might reduce the proportion of highly specialized herbivores, which ultimately would lead to changes in the specialization of interactions in plant-herbivore networks. This study investigates the community-wide effects of land use intensity on the degree of specialization of 72 plant-herbivore networks, including effects mediated by the increase in the proportion of exotic plant species. Contrary to our expectation, the net effect of land use intensity on network specialization was positive. However, this positive effect of land use intensity was partially canceled by an opposite effect of the proportion of exotic plant species on network specialization. When we analyzed networks composed exclusively of endophagous herbivores separately from those composed exclusively of exophagous herbivores, we found that only endophages showed a consistent change in network specialization at higher land use levels. Altogether, these results indicate that land use intensity is an important ecological driver of network specialization, by way of reducing the local host range of herbivore guilds with highly specialized feeding habits. However, because the effect of land use intensity is offset by an opposite effect owing to the proportion of exotic host species, the net effect of land use in a given herbivore assemblage will likely depend on the extent of the replacement of native host species with exotic ones.

Graphene Oxide: A Carrier For Pharmaceuticals And A Scaffold For Cell Interactions.

During the last ten years, graphene oxide has been explored in many applications due to its remarkable electroconductivity, thermal properties and mobility of charge carriers, among other properties. As discussed in this review, the literature suggests that a total characterization of graphene oxide must be conducted because oxidation debris (synthesis impurities) present in the graphene oxides could act as a graphene oxide surfactant, stabilizing aqueous dispersions. It is also important to note that the structure models of graphene oxide need to be revisited because of significant implications for its chemical composition and its direct covalent functionalization. Another aspect that is discussed is the need to consider graphene oxide surface chemistry. The hemolysis assay is recommended as a reliable test for the preliminary assessment of graphene oxide toxicity, biocompatibility and cell membrane interaction. More recently, graphene oxide has been extensively explored for drug delivery applications. An important increase in research efforts in this emerging field is clearly represented by the hundreds of related publications per year, including some reviews. Many studies have been performed to explore the graphene oxide properties that enable it to deliver more than one activity simultaneously and to combine multidrug systems with photothermal therapy, indicating that graphene oxide is an attractive tool to overcome hurdles in cancer therapies. Some strategic aspects of the application of these materials in cancer treatment are also discussed. In vitro studies have indicated that graphene oxide can also promote stem cell adhesion, growth and differentiation, and this review discusses the recent and pertinent findings regarding graphene oxide as a valuable nanomaterial for stem cell research in medicine. The protein corona is a key concept in nanomedicine and nanotoxicology because it provides a biomolecular identity for nanomaterials in a biological environment. Understanding protein corona-nanomaterial interactions and their influence on cellular responses is a challenging task at the nanobiointerface. New aspects and developments in this area are discussed.

Interplay Of Weak Interactions In The Atom-by-atom Condensation Of Xenon Within Quantum Boxes.

Condensation processes are of key importance in nature and play a fundamental role in chemistry and physics. Owing to size effects at the nanoscale, it is conceptually desired to experimentally probe the dependence of condensate structure on the number of constituents one by one. Here we present an approach to study a condensation process atom-by-atom with the scanning tunnelling microscope, which provides a direct real-space access with atomic precision to the aggregates formed in atomically defined 'quantum boxes'. Our analysis reveals the subtle interplay of competing directional and nondirectional interactions in the emergence of structure and provides unprecedented input for the structural comparison with quantum mechanical models. This approach focuses on-but is not limited to-the model case of xenon condensation and goes significantly beyond the well-established statistical size analysis of clusters in atomic or molecular beams by mass spectrometry.

Cation size effects-modified phase and PTCR development in Er3+ and Ca2+ co-doped BaTiO3 ceramics during sintering

Development of the positive temperature coefficient of resistivity (PTCR) in Er3+ and Ca2+ co-doped ferroelectric BaTiO3 was studied in this work, with Er3+ being used to act as a donor doping. Irrespective of all the materials showing high densities after sintering at 1200 to 1300 ºC, these revealed insulator at the lowest sintering temperature, changing to semiconducting and PTCR-type materials only when the sintering temperature was further increased. Observations from X-ray diffraction help correlating this effect with phase development in this formulated (Ba,Ca,Er)TiO3 system, considering the formation of initially two separated major (Ba,Ca)TiO3- and minor (Ca,Er)TiO3-based compounds, as a consequence of cation size-induced stress energy effects. Thus, appearance and enhancement here of the semiconducting and PTCR responses towards higher sintering temperatures particularly involve the incorporation of Er3+ into the major phase, rendering finally possible the generation and "percolative-like" migration of electrons throughout the whole material.