Carbon nanotubes modified with antimony nanoparticles: A novel material for electrochemical sensing

In this study, a novel material for the electrochemical determination of bisphenol A using a nanocomposite based on multi-walled carbon nanotubes modified with antimony nanoparticles has been investigated. The morphology, structure, and electrochemical performance of the nanocomposite electrodes were characterised by field emission gun scanning electron microscopy, energy-dispersive X-ray spectroscopy, and cyclic voltammetry. A scan rate study and electrochemical impedance spectroscopy showed that the bisphenol A oxidation product is adsorbed on nanocomposite electrode surface. Differential pulse voltammetry in phosphate buffer solution at pH 6, allowed the development of a method to determine bisphenol A levels in the range of 0.5-5.0 mu mol L-1, with a detection limit of 5.24 nmol L-1 (1.19 mu g L-1). (C) 2012 Elsevier Ltd. All rights reserved.

Dissecting structure-function-stability relationships of a thermostable GH5-CBM3 cellulase from Bacillus subtilis 168

Cellulases participate in a number of biological events, such as plant cell wall remodelling, nematode parasitism and microbial carbon uptake. Their ability to depolymerize crystalline cellulose is of great biotechnological interest for environmentally compatible production of fuels from lignocellulosic biomass. However, industrial use of cellulases is somewhat limited by both their low catalytic efficiency and stability. In the present study, we conducted a detailed functional and structural characterization of the thermostable BsCe15A (Bacillus subtilis cellulase 5A), which consists of a GH5 (glycoside hydrolase 5) catalytic domain fused to a CBM3 (family 3 carbohydrate-binding module). NMR structural analysis revealed that the Bacillus CBM3 represents a new subfamily, which lacks the classical calcium-binding motif, and variations in NMR frequencies in the presence of cellopentaose showed the importance of polar residues in the carbohydrate interaction. Together with the catalytic domain, the CBM3 forms a large planar surface for cellulose recognition, which conducts the substrate in a proper conformation to the active site and increases enzymatic efficiency. Notably, the manganese ion was demonstrated to have a hyper-stabilizing effect on BsCel5A, and by using deletion constructs and X-ray crystallography we determined that this effect maps to a negatively charged motif located at the opposite face of the catalytic site.

Medium Chain Fatty Acids Are Selective Peroxisome Proliferator Activated Receptor (PPAR) gamma Activators and Pan-PPAR Partial Agonists

Thiazolidinediones (TZDs) act through peroxisome proliferator activated receptor (PPAR) gamma to increase insulin sensitivity in type 2 diabetes (T2DM), but deleterious effects of these ligands mean that selective modulators with improved clinical profiles are needed. We obtained a crystal structure of PPAR gamma ligand binding domain (LBD) and found that the ligand binding pocket (LBP) is occupied by bacterial medium chain fatty acids (MCFAs). We verified that MCFAs (C8-C10) bind the PPAR gamma LBD in vitro and showed that they are low-potency partial agonists that display assay-specific actions relative to TZDs; they act as very weak partial agonists in transfections with PPAR gamma LBD, stronger partial agonists with full length PPAR gamma and exhibit full blockade of PPAR gamma phosphorylation by cyclin-dependent kinase 5 (cdk5), linked to reversal of adipose tissue insulin resistance. MCFAs that bind PPAR gamma also antagonize TZD-dependent adipogenesis in vitro. X-ray structure B-factor analysis and molecular dynamics (MD) simulations suggest that MCFAs weakly stabilize C-terminal activation helix (H) 12 relative to TZDs and this effect is highly dependent on chain length. By contrast, MCFAs preferentially stabilize the H2-H3/beta-sheet region and the helix (H) 11-H12 loop relative to TZDs and we propose that MCFA assay-specific actions are linked to their unique binding mode and suggest that it may be possible to identify selective PPAR gamma modulators with useful clinical profiles among natural products.

GQ-16, a Novel Peroxisome Proliferator-activated Receptor gamma (PPAR gamma) Ligand, Promotes Insulin Sensitization without Weight Gain

The recent discovery that peroxisome proliferator-activated receptor gamma (PPAR gamma) targeted anti-diabetic drugs function by inhibiting Cdk5-mediated phosphorylation of the receptor has provided a new viewpoint to evaluate and perhaps develop improved insulin-sensitizing agents. Herein we report the development of a novel thiazolidinedione that retains similar anti-diabetic efficacy as rosiglitazone in mice yet does not elicit weight gain or edema, common side effects associated with full PPAR gamma activation. Further characterization of this compound shows GQ-16 to be an effective inhibitor of Cdk5-mediated phosphorylation of PPAR gamma. The structure of GQ-16 bound to PPAR gamma demonstrates that the compound utilizes a binding mode distinct from other reported PPAR gamma ligands, although it does share some structural features with other partial agonists, such as MRL-24 and PA-082, that have similarly been reported to dissociate insulin sensitization from weight gain. Hydrogen/deuterium exchange studies reveal that GQ-16 strongly stabilizes the beta-sheet region of the receptor, presumably explaining the compound's efficacy in inhibiting Cdk5-mediated phosphorylation of Ser-273. Molecular dynamics simulations suggest that the partial agonist activity of GQ-16 results from the compound's weak ability to stabilize helix 12 in its active conformation. Our results suggest that the emerging model, whereby "ideal" PPAR gamma-based therapeutics stabilize the beta-sheet/Ser-273 region and inhibit Cdk5-mediated phosphorylation while minimally invoking adipogenesis and classical agonism, is indeed a valid framework to develop improved PPAR gamma modulators that retain antidiabetic actions while minimizing untoward effects.

Ligand- and Structure-Based Drug Design Strategies and PPAR delta/alpha Selectivity

Peroxisome-proliferator-activated receptors are a class of nuclear receptors with three subtypes: a, ? and d. Their main function is regulating gene transcription related to lipid and carbohydrate metabolism. Currently, there are no peroxisome-proliferator-activated receptors d drugs being marketed. In this work, we studied a data set of 70 compounds with a and d activity. Three partial least square models were created, and molecular docking studies were performed to understand the main reasons for peroxisome-proliferator-activated receptors d selectivity. The obtained results showed that some molecular descriptors (log P, hydration energy, steric and polar properties) are related to the main interactions that can direct ligands to a particular peroxisome-proliferator-activated receptors subtype.

Mode of Peroxisome Proliferator-Activated Receptor gamma Activation by Luteolin

The peroxisome proliferator-activated receptor gamma (PPAR gamma) is a target for treatment of type II diabetes and other conditions. PPAR gamma full agonists, such as thiazolidinediones (TZDs), are effective insulin sensitizers and anti-inflammatory agents, but their use is limited by adverse side effects. Luteolin is a flavonoid with anti-inflammatory actions that binds PPAR gamma but, unlike TZDs, does not promote adipocyte differentiation. However, previous reports suggested variously that luteolin is a PPAR gamma agonist or an antagonist. We show that luteolin exhibits weak partial agonist/antagonist activity in transfections, inhibits several PPAR gamma target genes in 3T3-L1 cells (LPL, ORL1, and CEBP alpha) and PPAR gamma-dependent adipogenesis, but activates GLUT4 to a similar degree as rosiglitazone, implying gene-specific partial agonism. The crystal structure of the PPAR gamma ligand-binding domain (LBD) reveals that luteolin occupies a buried ligand-binding pocket (LBP) but binds an inactive PPAR gamma LBD conformer and occupies a space near the beta-sheet region far from the activation helix (H12), consistent with partial agonist/antagonist actions. A single myristic acid molecule simultaneously binds the LBP, suggesting that luteolin may cooperate with other ligands to bind PPAR gamma, and molecular dynamics simulations show that luteolin and myristic acid cooperate to stabilize the Omega-loop among H2', H3, and the beta-sheet region. It is noteworthy that luteolin strongly suppresses hypertonicity-induced release of the pro-inflammatory interleukin-8 from human corneal epithelial cells and reverses reductions in transepithelial electrical resistance. This effect is PPAR gamma-dependent. We propose that activities of luteolin are related to its singular binding mode, that anti-inflammatory activity does not require H12 stabilization, and that our structure can be useful in developing safe selective PPAR gamma modulators.

Enzyme Microheterogeneous Hydration and Stabilization in Supercritical Carbon Dioxide

Supercritical carbon dioxide is a promising green-chemistry solvent for many enzyme-catalyzed chemical reactions, yet the striking stability of some enzymes in such unconventional environments is not well understood. Here, we investigate the stabilization of the Candida antarctica Lipase B (CALB) in supercritical carbon dioxide-water biphasic systems using molecular dynamics simulations. The preservation of the enzyme structure and optimal activity depend on the presence of small amounts of water in the supercritical dispersing medium. When the protein is at least partially hydrated, water molecules bind to specific sites on the enzyme surface and prevent carbon dioxide from penetrating its catalytic core. Strikingly, water and supercritical carbon dioxide cover the protein surface quite heterogeneously. In the first solvation layer, the hydrophilic residues at the surface of the protein are able to pin down patches of water, whereas carbon dioxide solvates preferentially hydrophobic surface residues. In the outer solvation shells, water molecules tend to cluster predominantly on top of the larger water patches of the first solvation layer instead of spreading evenly around the remainder of the protein surface. For CALB, this exposes the substrate-binding region of the enzyme to carbon dioxide, possibly facilitating diffusion of nonpolar substrates into the catalytic funnel. Therefore, by means of microheterogeneous solvation, enhanced accessibility of hydrophobic substrates to the active site can be achieved, while preserving the functional structure of the enzyme. Our results provide a molecular picture on the nature of the stability of proteins in nonaqueous media.

Structural Insights into Human Peroxisome Proliferator Activated Receptor Delta (PPAR-Delta) Selective Ligand Binding

Peroxisome proliferator activated receptors (PPARs delta, alpha and gamma) are closely related transcription factors that exert distinct effects on fatty acid and glucose metabolism, cardiac disease, inflammatory response and other processes. Several groups developed PPAR subtype specific modulators to trigger desirable effects of particular PPARs without harmful side effects associated with activation of other subtypes. Presently, however, many compounds that bind to one of the PPARs cross-react with others and rational strategies to obtain highly selective PPAR modulators are far from clear. GW0742 is a synthetic ligand that binds PPAR delta more than 300-fold more tightly than PPAR alpha or PPAR gamma but the structural basis of PPAR delta: GW0742 interactions and reasons for strong selectivity are not clear. Here we report the crystal structure of the PPAR delta:GW0742 complex. Comparisons of the PPAR delta:GW0742 complex with published structures of PPARs in complex with alpha and gamma selective agonists and pan agonists suggests that two residues (Val312 and Ile328) in the buried hormone binding pocket play special roles in PPAR delta selective binding and experimental and computational analysis of effects of mutations in these residues confirms this and suggests that bulky substituents that line the PPAR alpha and gamma ligand binding pockets as structural barriers for GW0742 binding. This analysis suggests general strategies for selective PPAR delta ligand design.

Electronic Structure Calculations in a 2D SixGe1-x Alloy Under an Applied Electric Field.

The recent advances and promises in nanoscience and nanotechnology have been focused on hexagonal materials, mainly on carbon-based nanostructures. Recently, new candidates have been raised, where the greatest efforts are devoted to a new hexagonal and buckled material made of silicon, named Silicene. This new material presents an energy gap due to spin-orbit interaction of approximately 1.5 meV, where the measurement of quantum spin Hall effect(QSHE) can be made experimentally. Some investigations also show that the QSHE in 2D low-buckled hexagonal structures of germanium is present. Since the similarities, and at the same time the differences, between Si and Ge, over the years, have motivated a lot of investigations in these materials. In this work we performed systematic investigations on the electronic structure and band topology in both ordered and disordered SixGe1-x alloys monolayer with 2D honeycomb geometry by first-principles calculations. We show that an applied electric field can tune the gap size for both alloys. However, as a function of electric field, the disordered alloy presents a W-shaped behavior, similarly to the pure Si or Ge, whereas for the ordered alloy a V-shaped behavior is observed.

Evaluation of degree of conversion and hardness of dental composites photo-activated with different light guide tips

Objective: The aim of this study was to evaluate the degree of conversion and hardness of different composite resins, photo-activated for 40 s with two different light guide tips, fiber optic and polymer. Methods: Five specimens were made for each group evaluated. The percentage of unreacted carbon double bonds (% C=C) was determined from the ratio of absorbance intensities of aliphatic C=C (peak at 1637 cm-1) against internal standard before and after curing of the specimen: aromatic C-C (peak at 1610 cm-1). The Vickers hardness measurements were performed in a universal testing machine. A 50 gf load was used and the indenter with a dwell time of 30 seconds. The degree of conversion and hardness mean values were analyzed separately by ANOVA and Tukey's test, with a significance level set at 5%. Results: The mean values of degree of conversion for the polymer and fiber optic light guide tip were statistically different (P<.001). The hardness mean values were statistically different among the light guide tips (P<.001), but also there was difference between top and bottom surfaces (P<.001). Conclusions: The results showed that the resins photo-activated with the fiber optic light guide tip promoted higher values for degree of conversion and hardness.

The structure of planktonic communities under variable coastal upwelling conditions off Cape Ghir (31ºN), in the Canary Current System (NW Africa)

Universidad de Las Palmas de Gran Canaria. Facultad de Ciencias del Mar. Programa de doctorado en Oceanografía. Diploma de Estudios Avanzados

Role of slowly settling particles in the ocean carbon cycle

[EN] Here we present results from sediment traps that separate particles as a function of their settling velocity, which were moored in the Canary Current region over a 1.5-year period. This study represents the longest time series using “in situ” particle settling velocity traps to date and are unique in providing year-round estimates. We find that, at least during half of the year in subtropical waters (the largest ocean domain), more than 60% of total particulate organic carbon is contained in slowly settling particles (0.7–11 m d−1). Analyses of organic biomarkers reveal that these particles have the same degradation state, or are even fresher than rapidly sinking particles. Thus, if slowly settling particles dominate the exportable carbon pool, most organic matter would be respired in surface waters, acting as a biological source of CO2 susceptible to exchange with the atmosphere. In the context of climate change, if the predicted changes in phytoplankton community structure occur, slowly settling particles would be favored, affecting the strength of the biological pump in the ocean.

p300/CBP tyrosine phosphorylation in response to dna damage activated by c-Abl tyrosine kinase

The nuclear signaling that is triggered in response to DNA damage entails the recruitment and assembly of repair proteins and the induction of genes involved in the activation of cell cycle checkpoint, apoptosis or senescence. The extensive changes in chromatin structure underlying these processes suggest that chromatin-modifying enzymes could be relevant targets of DNA damage-activated signaling. The acetyltransferases p300 and CBP participate in DNA damage-activated responses, including local histone hyperacetylation, cell cycle regulation, and co-activation of DNA damage activated proteins, such as p53, p73 and BRCA1. However, the link between DNA damage and p300/CBP activation has not been identified.We have detected p300 tyrosine phosphorylation in response to DNA damage. We show that the DNA damage-activated cAbl tyrosine kinase enters the nuclei of cells exposed to genotoxic agents and phosphorylates p300 on a tyrosine residue within the bromodomain that is conserved in p300, CBP and many other bromodomain-containing proteins. Antibodies against tyrosine phosphorylated p300/CBP show a DNA damage-inducible nuclear staining, suggesting that p300 tyrosine phosphorylation is an event linking DNA damage and chromatin modifications.

Reactivity of activated electrophiles and nucleophiles: labile intermediates and properties of the reaction products

The main topic of my Ph.D. thesis is the study of nucleophilic and electrophilic aromatic substitution reaction, in particular from a mechanistic point of view. The research was mainly focused on the reactivity of superactivated aromatic systems. In spite of their high reactivity (hence the high reaction’s rate), we were able to identify and in some case to isolate -complexes until now only hypothesized. For example, interesting results comes from the study of the protonation of the supernucleophiles tris(dialkylamino)benzenes. However, the best result obtained in this field was the isolation and structural characterization of the first stables zwitterionic Wheland-Meisenheimer complexes by using 2,4-dipyrrolidine-1,3-thiazole as supernucleophile and 4,6-dinitrobenzofuroxan or 4,6-dinitrotetrazolepyridine as superelectrophile. These reactions were also studied by means of computational chemistry, which allowed us to better investigate on the energetic and properties of the reactions and reactants studied. We also discovered, in some case fortuitously, some relevant properties and application of the compounds we synthesized, such as fluorescence in solid state and nanoparticles, or textile dyeing. We decided to investigate all these findings also by collaborating with other research groups. During a period in the “Laboratoire de Structure et Réactivité des Systèmes Moléculaires Complexes-SRSMC, Université de Lorraine et CNRS, France, I carried out computational studies on new iron complexes for the use as dyes in Dye Sensitized Solar Cells (DSSC). Furthermore, thanks to this new expertise, I was involved in a collaboration for the study of the ligands’ interaction in biological systems. A collaboration with University of Urbino allowed us to investigate on the reactivity of 1,2-diaza-1,3-dienes toward nucleophiles such as amino and phosphine derivatives, which led to the synthesis of new products some of which are 6 or 7 member heterocycles containing both phosphorus and nitrogen atoms.

Structure-property relations in polymers from renewable resources

The present research project focuses its attention on the study of structure-property relations in polymers from renewable sources (bio-based polymers) such as polymers microbially produced, i.e. polyhydrohyalkanoates (PHAs) or chemically synthesized using monomers from renewable sources, i.e. polyammide 11 (PA11). By means of a broad spectrum of experimental techniques, the influence of different modifications on bio-based polymers such as blending with other components, copolymerization with different co-monomers and introduction of branching to yield complex architectures have been investigated. The present work on PHAs focused on the study of the dependence of polymer properties on both the fermentation process conditions (e.g. bacterial strain and carbon substrate used) and the method adopted to recover PHAs from cells. Furthermore, a solvent-free method using an enzyme and chemicals in an aqueous medium, was developed in order to recover PHAs from cells. Such a method allowed to recover PHA granules in their amorphous state, i.e. in native form useful for specific applications (e.g. paper coating). In addition, a commercial PHA was used as polymeric matrix to develop biodegradable and bio-based composites for food packaging applications. Biodegradable, non-toxic, food contact plasticizers and low cost, widely available lignocellulosic fibers (wheat straw fibers) were incorporated in such a polymeric matrix, in order to decrease PHA brittleness and the polymer cost, respectively. As concerns the study of polyamide 11, both the rheological and the solid-state behavior of PA11 star samples with different arm number and length was studied. Introduction of arms in a polymer molecule allows to modulate melt viscosity behavior which is advantageous for industrial applications. Also, several important solid-state properties, in particular mechanical properties, are affected by the presence of branching. Given the importance of using ‘green’ synthetic strategies in polymer chemistry, novel poly(-amino esters), synthesized via enzymatic-catalyzed polymerization, have also been investigated in this work.