Search

In this study, different amounts of boric acid (BA, 1.25, 2.5, 3.75 and 5.0 wt%) were used to enhance the effectiveness of an intumescent flame retardant (IFR) system composed of ammonium polyphosphate (APP) and pentaerythritol (PER) in polypropylene (PP) including 2 wt% montmorillonite nanoclay (MMT). Meanwhile, metaboric acid and boron oxide which were generated by the decomposition of BA appeared in the melt compounding and the burning processes, respectively. Extensive experimental studies were performed to investigate the effects of BA/boron oxide and MMT combinations on the properties of PP/IFR. The fire resistances of the composites were studied by UL 94, limiting oxygen index (LOI) and cone calorimetry tests. The thermal properties were determined by using thermogravimetric analysis, differential scanning calorimetry and thermal conductivity measurements. In addition, the mechanical properties of the composites were examined. The experimental results revealed that although the additions of 1.25 and 2.5 wt% BA with 2 wt% MMT significantly enhanced thermal and flame resistances of PP composites, 3.75 and 5.0 wt% BA additions generated antagonistic effects and deteriorated the fire resistance of the composites. The sample including 2.5 wt% BA addition achieved the best flame retardancy. The LOI value was increased from 18 to 31% with UL 94 V-0 rating. In addition, the peak heat release rate was reduced from 668.6 to 150.0 kW/m2 and the total heat release value was decreased from 247.9 to 98.4 MJ/m2. In the meantime, the thermal conductivity was increased from 0.22 up to 0.28 W/mK. Furthermore, CO, CO2 and the smoke productions were significantly decreased with respect to PP. NO generation was decreased with BA replacements. At the same time, although there was a slight decrease in the tensile strength, the flexural strength significantly increased with BA and MMT additions.

Aggregation kinetics of proteins and peptides have been studied extensively due to their significance in many human diseases, including neurodegenerative disorders, and the roles they play in some key physiological processes. However, most of these studies have been performed as bulk measurements using Thioflavin T or other fluorescence turn-on reagents as indicators of fibrillization. Such techniques are highly successful in making inferences about the nucleation and growth mechanism of fibrils, yet cannot directly measure assembly reactions at low protein concentrations which is the case for amyloid-β (Aβ) peptide under physiological conditions. In particular, the evolution from monomer to low-order oligomer in early stages of aggregation cannot be detected. Single-molecule methods allow direct access to such fundamental information. We developed a high-throughput protocol for single-molecule photobleaching experiments using an automated fluorescence microscope. Stepwise photobleaching analysis of the time profiles of individual foci allowed us to determine stoichiometry of protein oligomers and probe protein aggregation kinetics. Furthermore, we investigated the potential application of supervised machine learning with support vector machines (SVMs) as well as multilayer perceptron (MLP) artificial neural networks to classify bleaching traces into stoichiometric categories based on an ensemble of measurable quantities derivable from individual traces. Both SVM and MLP models achieved a comparable accuracy of more than 80% against simulated traces up to 19-mer, although MLP offered considerable speed advantages, thus making it suitable for application to high-throughput experimental data. We used our high-throughput method to study the aggregation of Aβ40 in the presence of metal ions and the aggregation of α-synuclein in the presence of gold nanoparticles.

Accurate disease monitoring is essential following transarterial chemoembolization (TACE) in hepatocellular carcinoma (HCC) due to potential for profound adverse event and large variation in survival outcome. Post-treatment changes on conventional imaging can confound determination of residual/recurrent disease, magnifying the clinical challenge. Based on increased expression of thymidylate synthase (TYMS), thymidine kinase-1 (TK-1) and SLC29A1 (Equilibrative nucleoside transporter 1, ENT1) in HCC compared with liver tissue, we conducted a proof of concept study evaluating the efficacy of 18F-fluorothymidine (18F-FLT)-PET to assess response to TACE. As previous PET studies in HCC have been hampered by high background liver signal, we investigated if a temporal-intensity voxel-clustering (“Kinetic Spatial Filtering”) (KSF) improved lesion detection. Methods: A tissue microarray (TMA) was built from 36 HCC samples and matched surrounding cirrhotic tissue and was stained for thymidine kinase-1 (TK-1). A prospective study was conducted; eighteen patients with a diagnosis of HCC by American Association for the Study of Liver Diseases criteria (AALSD) who were eligible to treatment with TACE were enrolled. Patients underwent baseline conventional imaging and dynamic 18F-FLT-PET/KSF followed by TACE. Repeat imaging was performed 6-8 weeks post TACE. PET parameters were compared with modified-Response Evaluation in Solid Tumours (mRECIST) enhancement-based criteria. Results: Cancer Genome Atlas analysis revealed increased RNA expression of TYMS, TK-1 and SLC29A1 in HCC. TK-1 protein expression was significantly higher in HCC (p

Xylan, a prominent component of cellulosic biomass, has a high potential for degradation into reducing sugars, and subsequent conversion into bioethanol. This process requires a range of xylanolytic enzymes. Among them, β-xylosidases are crucial, because they hydrolyze more glycosidic bonds than any of the other xylanolytic enzymes. They also enhance the efficiency of the process by degrading xylooligosaccharides, which are potent inhibitors of other hemicellulose-/xylan-converting enzymes. On the other hand, the β-xylosidase itself is also inhibited by monosaccharides that may be generated in high concentrations during the saccharification process. Structurally, β-xylosidases are diverse enzymes with different substrate specificities and enzyme mechanisms. Here, we review the structural diversity and catalytic mechanisms of β-xylosidases, and discuss their inhibition by monosaccharides.

Isobutene is an important intermediate in the pyrolysis and oxidation of higher-order branched alkanes, and it is also a component of commercial gasolines. To better understand its combustion characteristics, a series of ignition delay time (IDT) and laminar flame speed (LFS) measurements have been performed. In addition, flow reactor speciation data recorded for the pyrolysis and oxidation of isobutene is also reported. Predictions of an updated kinetic model described herein are compared with each of these data sets, as well as with existing jet-stirred reactor (JSR) species measurements.IDTs of isobutene oxidation were measured in four different shock tubes and in two rapid compression machines (RCMs) under conditions of relevance to practical combustors. The combination of shock tube and RCM data greatly expands the range of available validation data for isobutene oxidation models to pressures of 50 atm and temperatures in the range 666-1715 K. Isobutene flame speeds were measured experimentally at 1 atm and at unburned gas temperatures of 298-398 K over a wide range of equivalence ratios. For the flame speed results, there was good agreement between different facilities and the current model in the fuel-rich region. Ab initio chemical kinetics calculations were carried out to calculate rate constants for important reactions such as H-atom abstraction by hydroxyl and hydroperoxyl radicals and the decomposition of 2-methylallyl radicals.A comprehensive chemical kinetic mechanism has been developed to describe the combustion of isobutene and is validated by comparison to the presently considered experimental measurements. Important reactions, highlighted via flux and sensitivity analyses, include: (a) hydrogen atom abstraction from isobutene by hydroxyl and hydroperoxyl radicals, and molecular oxygen; (b) radical-radical recombination reactions, including 2-methylallyl radical self-recombination, the recombination of 2-methylallyl radicals with hydroperoxyl radicals; and the recombination of 2-methylallyl radicals with methyl radicals; (c) addition reactions, including hydrogen atom and hydroxyl radical addition to isobutene; and (d) 2-methylallyl radical decomposition reactions. The current mechanism accurately predicts the IDT and LFS measurements presented in this study, as well as the JSR and flow reactor speciation data already available in the literature.The differences in low-temperature chemistry between alkanes and alkenes are also highlighted. in this work. In normal alkanes, the fuel radical (R) over dot adds to molecular oxygen forming alkylperoxyl (R(O) over dot(2)) radicals followed by isomerization and chain branching reactions which promote low-temperature fuel reactivity. However, in alkenes, because of the relatively shallow well (similar to 20 kcal mol(-1)) for R(O) over dot(2) formation compared to similar to 35 kcal mol(-1) in alkanes, the (R) over dot+O-2 (sic) R(O) over dot(2) equilibrium lies more to the left favoring (R) over dot+O-2 rather than R(O) over dot(2) radical stabilization. Based on this work, and related studies of allylic systems, it is apparent that reactivity for alkene components at very low temperatures (1300 K), the reactivity is mainly governed by the competition between hydrogen abstractions by molecular oxygen and OH radicals. (C) 2016 The Combustion Institute. Published by Elsevier Inc. All rights reserved. The work at NUI Galway was supported by Saudi Aramco under the FUELCOM program. Collaboration between NUI Galway and LRGP enters in the frame the COST Action CM1404. peer-reviewed 2017-03-17

© 2019, Springer Science+Business Media, LLC, part of Springer Nature. The growth behaviour of Cu 6 Sn 5 intermetallic compound (IMC) at the solder height controlled post-spread Sn/Cu interface is investigated for different initial solder ball volumes, reflow temperatures and cooling rates. Because of the limited solder thickness at the periphery, the IMC retains a preferential scalloped morphology, even after cooling. For solder balls of initial diameters of 500, 1000 and 1700 μm, with the maximum solder height at the peripheral regime not exceeding 150 μm, reflowed at 250 °C and undergoing air cooling, it has been revealed that IMC characterized with larger layer thickness and grain diameter, correspond to the sample of smaller ball size. The increase in reflow temperature for a solder of initial size of 500 μm, is characterized by the increase in IMC thickness, developments of few but quite large faceted planes over the original scalloped morphology and non-uniformity in the grain diameter. In contrary the to air cooling (cooling rate ≥ 4.0 K/s), the IMCs obtained at the thin film zone, for furnace cooling (cooling rate = 0.037 K/s), are very larger, both in grain size and layer thickness. Moreover, the scalloped Cu 6 Sn 5 surface in furnace cooled specimens, bear many but tiny facets. Finite element analysis is utilized to numerically estimate the diffusion of Cu into the geometrical volumes of solder. Ostwald ripening and film height are the important parameters defining the growth behavior of the compound in miniaturized solder joints. ispartof: JOURNAL OF MATERIALS SCIENCE-MATERIALS IN ELECTRONICS vol:30 issue:3 pages:2872-2887 status: published

PrimPol is a human primase/polymerase specialized in re-starting stalled forks by repriming beyond lesions such as pyrimidine dimers, and replication-perturbing structures including G-quadruplexes and R-loops. Unlike most conventional primases, PrimPol proficiently discriminates against ribonucleotides (NTPs), being able to start synthesis using deoxynucleotides (dNTPs), yet the structural basis and physiological implications for this discrimination are not understood. In silico analyses based on the three-dimensional structure of human PrimPol and related enzymes enabled us to predict a single residue, Tyr, as the main effector of sugar discrimination in human PrimPol and a change of Tyr to histidine to boost the efficiency of NTP incorporation. We show here that the Y100H mutation profoundly stimulates NTP incorporation by human PrimPol, with an efficiency similar to that for dNTP incorporation during both primase and polymerase reactions in vitro. As expected from the higher cellular concentration of NTPs relative to dNTPs, Y100H expression in mouse embryonic fibroblasts and U2OS osteosarcoma cells caused enhanced resistance to hydroxyurea, which decreases the dNTP pool levels in S-phase. Remarkably, the Y100H PrimPol mutation has been identified in cancer, suggesting that this mutation could be selected to promote survival at early stages of tumorigenesis, which is characterized by depleted dNTP pools., Spanish Ministry of Economy and Competitiveness (MINECO; BFU2012–3769, BFU2014–51672-REDC and BFU2015–65880-P (co-funded with European Union FEDER funds) to L.B.; BFU2013–49153-P and BFU2016–80402-R (co-funded with European Union FEDER funds) to J.M.). A.D.T., P.A.C. and D.G.A. are recipients of MINECO FPI-predoctoral fellowships. M.D. was the recipient of a FPI-predoctoral fellowship from Programa de Excelencia “Severo Ochoa” (CNIO-MINECO)

Summary Alternative battery technologies are required to meet growing energy demands and address the limitations of present technologies. As such, it is necessary to look beyond lithium-ion batteries. Zinc batteries enable high power density while being sourced from ubiquitous and cost-effective materials. This paper presents, for the first time known to the authors, multi-length scale tomography studies of failure mechanisms in zinc batteries with and without commercial microporous separators. In both cases, dendrites were grown, dissolved, and regrown, critically resulting in different morphology of dendritic layer formed on both the electrode and the separator. The growth of dendrites and their volume-specific areas were quantified using tomography and radiography data in unprecedented resolution. High-resolution ex situ analysis was employed to characterize single dendrites and dendritic deposits inside the separator. The findings provide unique insights into mechanisms of metal-battery failure effected by growing dendrites.

A new dual-curing system based on sequential thiol-epoxy click polycondensation and epoxy anionic homopolymerization was studied. Formulations of diglycidyl ether of bisphenol A and trimethylolpropane tris(3-mercaptopropionate) with 1-methylimidazole as a base catalyst and excess of epoxy groups were prepared and characterized. The curing process is sequential: fast thiol-epoxy polycondensation takes place first, followed by slower homopolymerization of excess epoxy groups. This makes it possible to define curing sequences with easy time-temperature control for both curing stages. The network buildup process during the first curing stage can be easily modelled assuming ideal polycondensation, which allows tailoring the structure and properties of the intermediate materials. The homopolymerization of the excess epoxy groups in the second curing stage results in a higher glass transition temperature (T-g) in comparison with the stoichiometric thiol-epoxy material, thus extending the application of thiol-epoxy thermosets to wider temperature ranges.

The author acknowledges with gratitude financial assistance from Rhodes University and the National Research Foundation of South Africa (Grant UID74438). The kinetic analysis of thermoluminescence of beta-irradiated SrAl2O4:Eu2+,Dy3+ is reported. The glow-curve is dominated by an apparently-single peak. It has been demonstrated using a number of tests including partial dynamic-heating, isothermal heating, phosphorescence and, the effect of fading, that the peak and the glow-curve consists of a set of closely-spaced peaks. In view of the peak being complex, its first few components were abstracted and analysed and for comparison, the peak was also analysed assuming it is genuinely single. In the latter, the order of kinetics is calculated to be intermediate between first and second-order and not first-order as predicted by qualitative tests such as the Tm − Tstop or Tm − dose procedures. A model based on density of energy states has been used to account for and reconcile the qualitative and quantitative results. The activation energy is found as ∼1 eV, consistent with the value expected of Dy2+, the presumed electron trapping state of the Dy3+ electron trap. The thermoluminescence is subject to thermal quenching with an activation energy of 0.520 ± 0.002 eV. The luminescence is ascribed to 5d→4f7 Eu2+ transitions whereas the thermal quenching is presumed to occur from an alternative level of the degenerate 5d energy level of the Eu2+ cation. Postprint

l-galactono-1,4-lactone dehydrogenase (GALDH) catalyzes the terminal step of vitamin C biosynthesis in plant mitochondria. Here we investigated the communication between Arabidopsis thaliana GALDH and its natural electron acceptor cytochrome c (Cc). Using laser-generated radicals we observed the formation and stabilization of the GALDH semiquinone anionic species (GALDHSQ ). GALDHSQ oxidation by Cc exhibited a nonlinear dependence on Cc concentration consistent with a kinetic mechanism involving protein-partner association to form a transient bimolecular complex prior to the electron transfer step. Oxidation of GALDHSQ by Cc was significantly impaired at high ionic strength, revealing the existence of attractive charge-charge interactions between the two reactants. Isothermal titration calorimetry showed that GALDH weakly interacts with both oxidized and reduced Cc. Chemical shift perturbations for (1) H and (15) N nuclei of Cc, arising from the interactions with unlabeled GALDH, were used to map the interacting surface of Cc. For Arabidopsis Cc and yeast Cc, similar residues are involved in the interaction with GALDH. These residues are confined to a single surface surrounding the heme edge. The range of chemical shift perturbations for the physiological Arabidopsis Cc-GALDH complex is larger than that of the non-physiological yeast Cc-GALDH complex, indicating that the former complex is more specific. In summary, the results point to a relatively low affinity GALDH-Cc interaction, similar for all partner redox states, involving protein-protein dynamic motions. Evidence is also provided that Cc utilizes a conserved surface surrounding the heme edge for the interaction with GALDH and other redox partners.

Nisin is a posttranslationally modified antimicrobial peptide containing the cyclic thioether amino acids lanthionine and methyllanthionine. Although much is known about its antimicrobial activity and mode of action, knowledge about the nisin modification process is still rather limited. The dehydratase NisB is believed to be the initial interaction partner in modification. NisB dehydrates specific serine and threonine residues in prenisin, whereas the cyclase NisC catalyzes the (methyl) lanthionine formation. The fully modified prenisin is exported and the leader peptide is cleaved off by the extracellular protease NisP. Light scattering analysis demonstrated that purified NisB is a dimer in solution. Using size exclusion chromatography and surface plasmon resonance, the interaction of NisB and prenisin, including several of its modified derivatives, was studied. Unmodified prenisin binds to NisB with an affinity of 1.05 +/- 0.25 mu M, whereas the dehydrated and the fully modified derivatives bind with respective affinities of 0.31 +/- 0.07 and 10.5 +/- 1.7 mu M. The much lower affinity for the fully modified prenisin was related to a > 20-fold higher off-rate. For all three peptides the stoichiometry of binding was 1:1. Active nisin, which is the equivalent of fully modified prenisin lacking the leader peptide did not bind to NisB, nor did prenisin in which the highly conserved FNLD box within the leader peptide was mutated to AAAA. Taken together our data indicate that the leader peptide is essential for initial recognition and binding of prenisin to NisB.

Lung surfactant (LS) is a lipid-rich material lining the inside of the lungs. It reduces surface tension at the liquid/air interface and thus, it confers protection of the alveoli from collapsing. The surface-active component of LS is dipalmitoyl-phosphatidylcholine, while anionic phospholipids such as phosphatidylinositol (PtdIns) and primarily phosphatidylglycerol are involved in the stabilization of the LS monolayer. The exact role of PtdIns in this system is not well-understood; however, PtdIns levels change dramatically during the acute respiratory distress syndrome (ARDS) evolution. In this report we present evidence of a phosphoinositide-specific phospholipase C (PI-PLC) activity in bronchoalveolar lavage (BAL) fluid, which may regulate PtdIns levels. Characterization of this extracellular activity showed specificity for PtdIns and phosphatidylinositol 4,5-bisphosphate, sharing the typical substrate concentration-, pH-, and calcium-dependencies with mammalian PI-PLCs. Fractionation of BAL fluid showed that PI-PLC did not co-fractionate with large surfactant aggregates, but it was found mainly in the soluble fraction. Importantly, analysis of BAL samples from control subjects and from patients with ARDS showed that the PI-PLC specific activity was decreased by 4-fold in ARDS samples concurrently with the increase in BAL PtdIns levels. Thus, we have identified for the first time an extracellular PI-PLC enzyme activity that may be acutely involved in the regulation of PtdIns levels in LS. Am J Respir Cell Mol Biol

Formation of liquid-ordered domains in model membranes can be linked to raft formation in cellular membranes. The lipid stoichiometry has a governing influence on domain formation and consequently, biochemical hydrolysis of specific lipids has the potential to remodel domain features. Activation of phospholipase A(2) (PLA(2)) by ternary model membranes with three components (DOPC/DPPC/Cholesterol) can potentially change the domain structure by preferential hydrolysis of the phospholipids. Using fluorescence microscopy, this work investigates the changes in domain features that occur upon PLA(2) activation by such ternary membranes. Double-supported membranes are used, which have minimal interactions with the solid support. For membranes prepared in the coexistence region, PLA(2) induces a decrease of the liquid-disordered (L(d)) phase and an increase of the liquid-ordered (L(o)) phase. A striking observation is that activation by a uniform membrane in the L(d) phase leads to nucleation and growth of L(o)-like domains. This phenomenon relies on the initial presence of cholesterol and no PLA(2) activation is observed by membranes purely in the L(o) phase. The observations can be rationalized by mapping partially hydrolyzed islands onto trajectories in the phase diagram. It is proposed that DPPC is protected from hydrolysis through interactions with cholesterol, and possibly the formation of condensed complexes. This leads to specific trajectories which can account for the observed trends. The results demonstrate that PLA(2) activation by ternary membrane islands may change the global lipid composition and remodel domain features while preserving the overall membrane integrity.

Understanding the relationship between the location of nanoparticles (NPs) in an organic matrix and their catalytic performances is essential for catalyst design. Here we show that catalytic activities of Au, Ag and CuNPs stabilized by dendrimers using coordination to intradendritic triazoles, galvanic replacement or stabilization outside dendrimers strongly depends on their location. AgNPs are found at the inner click dendrimer periphery, whereas CuNPs and AuNPs are encapsulated in click dendrimer nanosnakes. AuNPs and AgNPs formed by galvanic replacement are larger than precursors and only partly encapsulated. AuNPs are all the better 4-nitrophenol reduction catalysts as they are less sterically inhibited by the dendrimer interior, whereas on the contrary CuNPs are all the better alkyne azide cycloaddition catalysts as they are better protected from aerobic oxidation inside dendrimers. This work highlights the role of the location in macromolecules on the catalytic efficiency of metal nanoparticles and rationalizes optimization in catalyst engineering., Organic supports are often employed for encapsulation of metal nanoparticles. Here the authors report a method for the precise localisation of gold, silver and copper nanoparticles within a dendrimer support and show how this localisation influences catalytic performance.

In order to understand the mechanism by which a polypeptide chain folds into its functionally active native state it is necessary to characterize in detail all the species accumulated along the pathway. The elusive nature of protein folding intermediates poses their identification and characterization as an extremely difficult task in the protein folding field. In the case of small single domain proteins, the direct measurement of the thermodynamics and structural parameters of protein folding intermediates has provided new insights on the nature of the forces involved in the stabilization of nascent protein structures. Here we summarize some of the experimental approaches aimed at the detection and characterization of folding intermediates along with a discussion of some general structural features emerging from these studies.

Amyloids are cross-β-sheet fibrillar aggregates, associated with various human diseases and native functions such as protein/peptide hormone storage inside secretory granules of neuroendocrine cells. In the current study, using amyloid detecting agents, we show that growth hormone (GH) could be stored as amyloid in the pituitary of rat. Moreover, to demonstrate the formation of GH amyloid in vitro, we studied various conditions (solvents, glycosaminoglycans, salts and metal ions) and found that in presence of zinc metal ions (Zn(II)), GH formed short curvy fibrils. The amyloidogenic nature of these fibrils was examined by Thioflavin T binding, Congo Red binding, transmission electron microscopy and X-ray diffraction. Our biophysical studies also suggest that Zn(II) initiates the early oligomerization of GH that eventually facilitates the fibrillation process. Furthermore, using immunofluorescence study of pituitary tissue, we show that GH in pituitary significantly co-localizes with Zn(II), suggesting the probable role of zinc in GH aggregation within secretory granules. We also found that GH amyloid formed in vitro is capable of releasing monomers. The study will help to understand the possible mechanism of GH storage, its regulation and monomer release from the somatotrophs of anterior pituitary.

Perfusion systems have the possibility to be operated continuously for several months. It is important that the performance of the cell retention device does not limit the operation time of a perfusion process used in the production of active pharmaceutical ingredients. Therefore, the aim of this study was to investigate the reliability and long-term stability of an acoustic perfusion process using the 200 L/d BioSep. As the BioSep is an external device, it is possible that dependent on the recirculation rate nutrient gradients occur in the external loop, which could affect the cell metabolism. Therefore, the effect of possible nutrient gradients on cell metabolism, viability and productivity was studied by varying the recirculation rate. In this study, it is shown that a perfusion process using a pilot-scale acoustic cell-retention device (200 L/d) is reliable and simple to operate, resulting in a stable 75-day cultivation of a hybridoma cell line producing a monoclonal antibody. The recirculation rate had a significant effect on the oxygen concentration in the external loop, with oxygen being depleted within the cell-retention device at recirculation rates below 6 m3/m3reactor · d (=600 L/d). The oxygen depletion at low circulation rates correlated with a slightly increased lactate production rate. For all other parameters no effect of the recirculation rate was observed, including cell death measured through the release of lactate dehydrogenase and specific productivity. A maximum specific productivity of 12 pg/cell · d was reached.

Increased glucose metabolism of inflammatory tissues is the main source of false-positive F-18-FDG PET findings in oncology. It has been suggested that radiolabeled nucleosides might be more tumor specific. Methods: To test this hypothesis, we compared the biodistribution of 3'-deoxy-3'-F-18-fluorothymidine (FLT) and F-18-FDG in Wistar rats that bore tumors (C6 rat glioma in the right shoulder) and also had sterile inflammation in the left calf muscle (induced by injection of 0.1 mL of turpentine). Twenty-four hours after turpentine injection, the rats received an intravenous bolus (30 MBq) of either F-18-FLT (n = 5) or F-18-FDG (n = 5). Pretreatment of the animals with thymidine phosphorylase (>1,000 U/kg, intravenously) before injection of F-18-FLT proved to be necessary to reduce the serum levels of endogenous thymidine and achieve satisfactory tumor uptake of radioactivity. Results: Tumor-to-muscle ratios of F-18-FDG at 2 h after injection (13.2 +/- 3.0) were higher than those of F-18-FLT (3.8 +/- 1.3). F-18-FDG showed high physiologic uptake in brain and heart, whereas F-18-FLT was avidly taken up by bone marrow. F-18-FDG accumulated in the inflamed muscle, with 4.8 +/- 1.2 times higher uptake in the affected thigh than in the contralateral healthy thigh, in contrast to F-18-FLT, for which this ratio was not significantly different from unity (1.3 +/- 0.4). Conclusion; In F-18-FDG PET images, both tumor and inflammation were visible, but F-18-FLT PET showed only the tumor. Thus, the hypothesis that F-18-FLT has a higher tumor specificity was confirmed in our animal model.

Haloalkane dehalogenases are bacterial enzymes capable of carbon-halogen bond cleavage in halogenated compounds. To obtain insights into the mechanism of the haloalkane dehalogenase from Sphingomonas paucimobilis UT26 (LinB), we studied the steady-state and presteady-state kinetics of the conversion of the substrates 1-chlorohexane, chlorocyclohexane, and bromocyclohexane. The results lead to a proposal of a minimal kinetic mechanism consisting of three main steps: (i) substrate binding, (ii) cleavage of the carbon-halogen bond with simultaneous formation of an alkyl-enzyme intermediate, and (iii) hydrolysis of the alkyl-enzyme intermediate. Release of both products, halide and alcohol, is a fast process that was not included in the reaction mechanism as a distinct step. Comparison of the kinetic mechanism of LinB with that of haloalkane dehalogenase DhlA from Xantobacter autotrophicus GJ10 and the haloalkane dehalogenase DhaA from Rhodococcus rhodochrous NCIMB 13064 shows that the overall mechanisms are similar. The main difference is in the rate-limiting step, which is hydrolysis of the alkylenzyme intermediate in LinB, halide release in DhlA, and liberation of an alcohol in DhaA. The occurrence of different rate-limiting steps for three enzymes that belong to the same protein family indicates that extrapolation of this important catalytic property from one enzyme to another can be misleading even for evolutionary closely related proteins. The differences in the rate-limiting step were related to: (i) number and size of the entrance tunnels, (ii) protein flexibility, and (iii) composition of the halide-stabilizing active site residues based on comparison of protein structures.

Reference tissue models have gained significant traction over the last two decades as the methods of choice for the quantification of brain positron emission tomography data because they balance quantitative accuracy with less invasive procedures. The principal advantage is the elimination of the need to perform arterial cannulation of the subject to measure blood and metabolite concentrations for input function generation. In particular, the simplified reference tissue model (SRTM) has been widely adopted as it uses a simplified model configuration with only three parameters that typically produces good fits to the kinetic data and a stable parameter estimation process. However, the model's simplicity and its ability to generate good fits to the data, even when the model assumptions are not met, can lead to misplaced confidence in binding potential (BPND) estimates. Computer simulation were used to study the bias introduced in BPND estimates as a consequence of violating each of the four core SRTM model assumptions. Violation of each model assumption led to bias in BPND (both over and underestimation). Careful assessment of the bias in SRTM BPND should be performed for new tracers and applications so that an appropriate decision about its applicability can be made.

Protein glycosylation with O-linked N-acetylglucosamine (O-GlcNAc) is a post-translational modification of serine/ threonine residues in nucleocytoplasmic proteins. O-GlcNAc has been shown to play a role in many different cellular processes and O-GlcNAcylation is often found at sites that are also known to be phosphorylated. Unlike phosphorylation, O-GlcNAc levels are regulated by only two enzymes, O-GlcNAc transferase (OGT) and O-GlcNAc hydrolase (O-GlcNAcase or OGA). So far, no obvious consensus sequence has been found for sites of O-GlcNAcylation. Additionally, O-GlcNAcase recognizes and cleaves all O-GlcNAcylated proteins, independent of their sequence. In this work, we generate and analyze five models of O-GlcNAcylated peptides in complex with a bacterial OGA. Each of the five glycopeptides bind to OGA in a similar fashion, with OGA–peptide interactions primarily, but not exclusively, involving the peptide backbone atoms, thus explaining the lack of sensitivity to peptide sequence. Nonetheless, differences in peptide sequences, particularly at the � 1t o�4 positions, lead to variations in predicted affinity, consistent with observed experimental variations in enzyme kinetics. The potential exists, therefore, to employ the present analysis to guide the development glycopeptide-specific inhibitors, or conversely, the conversion of OGA into a reagent that could target specific O-GlcNAcylated peptide sequences.

Methods and models describing oxygen diffusion and desorption in oxides have been developed for slightly defective and well crystallised bulky materials. Does nanostructuring change the mechanism of oxygen mobility? In such a case, models should be properly checked and adapted to take into account new material properties. In order to do so, temperature programmed oxygen desorption and thermogravimetric analysis, either in isothermal or ramp mode, have been used to investigate some nanostructured La(1-x)A(x)MnO(3-delta) samples (A = Sr and Ce, 20-60 nm particle size) with perovskite-like structure. The experimental data have been elaborated by means of different models to define a set of kinetic parameters able to describe oxygen release properties and oxygen diffusion through the bulk. Different rate-determining steps have been identified, depending on the temperature range and oxygen depletion of the material. In particular, oxygen diffusion was shown to be rate-limiting at low temperature and at low defect concentration, whereas oxygen recombination at the surface seems to be the rate-controlling step at high temperature. However, the oxygen recombination step is characterised by an activation energy much lower than that for diffusion. In the present paper oxygen transport in nanosized materials is quantified by making use of widely diffused experimental techniques and by critically adapting to nanoparticles suitably chosen models developed for bulk materials

l-galactono-1,4-lactone dehydrogenase (GALDH) catalyzes the terminal step of vitamin C biosynthesis in plant mitochondria. Here we investigated the communication between Arabidopsis thaliana GALDH and its natural electron acceptor cytochrome c (Cc). Using laser-generated radicals we observed the formation and stabilization of the GALDH semiquinone anionic species (GALDHSQ ). GALDHSQ oxidation by Cc exhibited a nonlinear dependence on Cc concentration consistent with a kinetic mechanism involving protein-partner association to form a transient bimolecular complex prior to the electron transfer step. Oxidation of GALDHSQ by Cc was significantly impaired at high ionic strength, revealing the existence of attractive charge-charge interactions between the two reactants. Isothermal titration calorimetry showed that GALDH weakly interacts with both oxidized and reduced Cc. Chemical shift perturbations for (1) H and (15) N nuclei of Cc, arising from the interactions with unlabeled GALDH, were used to map the interacting surface of Cc. For Arabidopsis Cc and yeast Cc, similar residues are involved in the interaction with GALDH. These residues are confined to a single surface surrounding the heme edge. The range of chemical shift perturbations for the physiological Arabidopsis Cc-GALDH complex is larger than that of the non-physiological yeast Cc-GALDH complex, indicating that the former complex is more specific. In summary, the results point to a relatively low affinity GALDH-Cc interaction, similar for all partner redox states, involving protein-protein dynamic motions. Evidence is also provided that Cc utilizes a conserved surface surrounding the heme edge for the interaction with GALDH and other redox partners.

Determination of brain glucose transport kinetics in vivo at steady-state typically does not allow distinguishing apparent maximum transport rate (T(max)) from cerebral consumption rate. Using a four-state conformational model of glucose transport, we show that simultaneous dynamic measurement of brain and plasma glucose concentrations provide enough information for independent and reliable determination of the two rates. In addition, although dynamic glucose homeostasis can be described with a reversible Michaelis-Menten model, which is implicit to the large iso-inhibition constant (K(ii)) relative to physiological brain glucose content, we found that the apparent affinity constant (K(t)) was better determined with the four-state conformational model of glucose transport than with any of the other models tested. Furthermore, we confirmed the utility of the present method to determine glucose transport and consumption by analysing the modulation of both glucose transport and consumption by anaesthesia conditions that modify cerebral activity. In particular, deep thiopental anaesthesia caused a significant reduction of both T(max) and cerebral metabolic rate for glucose consumption. In conclusion, dynamic measurement of brain glucose in vivo in function of plasma glucose allows robust determination of both glucose uptake and consumption kinetics.

International audience; N-Heterocyclic carbenes (NHCs), namely, 1,3-bis-(diisopropyl)imidazol-2-ylidene (1) and 1, 3-bis(di-tert-butyl)imidazol-2-ylidene (2) were employed as neutral organocatalysts to bring about the group transfer polymerization (GTP) of both methacrylic and acrylic monomers, including methyl methacrylate (MMA), tert-butylacrylate (tBA), and n-butylacrylate (nBA). This could be achieved at room temperature using 1-methoxy-2-methyl-1-trimethylsiloxypropene (MTS) as initiator in polar or apolar medium. In this way, polymethacrylates and polyacrylates with molar masses in the range 10 000-300 000 g.mol(-1), corresponding to the initial [monomer]/[MTS] ratio and with polydispersities lower than 1.2, were obtained in quantitative yields. The kinetics of GTP of MMA catalyzed by 1 or 2 was further investigated. Though the first-order kinetic plot ln[M](0)/[M] versus time deviated from linearity at high monomer conversion, no inhibition period was noted at low monomer conversion. Moreover, the polymerization rate dramatically increased as the concentration of initiator increased, with first-order dependence in initiator. When mixed in 1/1 molar ratio, MTS and NHC I did not reveal the formation of enolate-type species by Si-29 or 13 C NMR spectroscopy. Based on these observations, we propose that NHCs activate the silyl ketene acetal initiator and further propagate GTP via an associative mechanism. The fact that In[M](0)/[M] does not evolve linearly with time in the terminal phase of the polymerization can be understood by a reduced diffusion of the catalyst to the trimethylsilyl end groups. The proposed associative mechanism can also account for the successful control of NHC-catalyzed GTP of acrylates during which termination reactions such as backbiting or internal isomerization could be drastically minimized. Next, was described the synthesis of all acrylic block copolymers based on polyacrylates and polymethacrylates (e.g., PMMA-b-PnBA-b-PMMA), utilizing the same NHC as catalyst in sequential GTP. It is again argued that such block copolymer formation is favored by an associative mechanism forming highly unstable activated silicon intermediates.

Mixed oxidic and perovskite-type materials based on the La, Sr, Ce and Fe elements were prepared using a mixture of nitrates salts and heating at 1000 degrees C. Three groups of solids were synthesized: (i) La1-yCeyFeO3 (y = 0.2, 0.3, 0.5), (ii) La1-xSrxFeO3 (x = 0.2, 0.3, 0.5) and (iii) La1-x-ySrxCeyFeO3 (x/y = 0.05/0.15, 0.15/0.05,.1/0.2, 0.2/0.1, 0.2/0.3 and 0.3/0.2). The structure of the solids was examined by XRD and the main crystal phases determined were LaFeO3, alpha-Fe2O3 and CeO2 in group (i), LaFeO3 and SrFeO3-x, in group (ii), and LaFeO3, alpha-Fe2O3 SrFeO3-x, and CeO2 in group (iii), while traces of La(OH)(3) and SrFe12O19 were also detected. The precise determination of the percentage amount of the iron-containing crystal phases in each solid composition was-determined by Mossbauer spectroscopy at 20 K. The ceramic materials had low surface areas and were tested for their catalytic activity for the NO + CO reaction in a flow reactor in the range of 280-560 degrees C. Conversions as high as 90% were achieved at 550 degrees C at a GHSV = 54000h(-1). The reaction rate of NO conversion is favored by the increased amount of CeO2 in groups (1) and (iii) of solids that contain cerium. In the case of solids without CeO2 (group ii), the NO conversion is favored by the existence of SrFeO3-x, phase at low temperatures (280-440 degrees C), while it decreases at high temperatures (440-560 degrees C). The double substituted solids La1-x-ySrxCeyFeO3 with x+y>0.3 and y>x were found to be the best catalysts for the NO+CO reaction as compared to the single substituted mixed oxides. Temperature programmed desorption (TPD) studies of NO and CO2 support the view that a synergistic effect takes place between the two phases of CeO2 and SrFeO3-x, whose co-existence results in the maximum enhancement of activity, via alternative oxidation-reduction cycles in the two phases. (C) 2000 Elsevier Science B.V. All rights reserved. Applied Catalysis B-Environmental

Solids of the nominal formula La1-xFeO3 where x=0.00, 0.05, 0.10, 0.15, 0.20, 0.25, 0.35, and containing only the perovskite (small x) and the perovskite plus Fe2O3 crystal phases (large X) were examined in the catalytic reaction of (NO+CO). The solids were prepared by heating at T=1000 degrees C under similar to 10(-5) Torr vacuum and showed appreciable catalytic activity at temperatures from 280 degrees C to 480 degrees C when SV is similar to 300 h(-1). XRD analysis and Rietveld refinement showed that for x=0.00 and 0.05 only the perovskite phase is apparent. For x greater than or equal to 0.10 an additional crystal phase of Fe2O3 appears which increases with x up to a maximum of 4.1% at x=0.35, Moessbauer examination indicated that iron exists in the perovskite structure at x=0.00 and 0.05 but for x greater than or equal to 0.10 the extra-perovskite Fe3+ increases proportional to the parameter x of the solids La1-xFeO3 and reaches 43% at x=0.35. These results are explained assuming that in the catalyst particles with x greater than or equal to 0.10 a Fe2O3 core of increasing size is covered by a LaFeO3 shell. The sample LaFeO3 showed lower catalytic activity for the NO+CO reaction than the rest of La1-xFeO3 solids. The Arrhenius plots showed two distinct regions of activity one at low temperature with high apparent activation energies and another at high temperature with lower apparent activation barriers. At low temperatures the low activity at x=0.00 is related to high apparent activation energies while for x greater than or equal to 0.05 the opposite is true. A detailed scrutinization of the apparent activation energies resulted in an estimation of the heats of adsorption of NO on LaFeO3 and La1-xFeO3. The reaction of NO+CO also resulted in small amounts of N2O which showed a maximum at similar to 320 degrees C. The dependence of N2O production and elimination on the temperature made it possible to determine the activation energies of its formation as well as the heat of desorption of nitrogen from the catalyst surface. The catalytic activity of the solids is destroyed if they are heated under atmospheric conditions. (C) 1999 Elsevier Science B.V. All rights reserved. Applied Catalysis a-General

Penning transfers, a group of processes by which excitation energy is used to ionise the gas, increase the gas gain in some detectors. Both the probability that such transfers occur and the mechanism by which the transfer takes place, vary with the gas composition and pressure. With a view to developing a microscopic electron transport model that takes Penning transfers into account, we use this dependence to identify the transfer mechanisms at play. We do this for a number of argon-based gas mixtures, using gain curves from the literature.

20 pages, 9 figures, 1 box, supporting information https://doi.org/10.1002/wcms.1623.-- Data availabitity statement: Data sharing is not applicable to this article as no new data were created or analyzed in this study., Adenylyl cyclases (ACs) play a key role in many signaling cascades. ACs catalyze the production of cyclic AMP from ATP and this function is stimulated or inhibited by the binding of their cognate stimulatory or inhibitory Gα subunits, respectively. Here we used simulation tools to uncover the molecular and subcellular mechanisms of AC function, with a focus on the AC5 isoform, extensively studied experimentally. First, quantum mechanical/molecular mechanical free energy simulations were used to investigate the enzymatic reaction and its changes upon point mutations. Next, molecular dynamics simulations were employed to assess the catalytic state in the presence or absence of Gα subunits. This led to the identification of an inactive state of the enzyme that is present whenever an inhibitory Gα is associated, independent of the presence of a stimulatory Gα. In addition, the use of coevolution-guided multiscale simulations revealed that the binding of Gα subunits reshapes the free-energy landscape of the AC5 enzyme by following the classical population-shift paradigm. Finally, Brownian dynamics simulations provided forward rate constants for the binding of Gα subunits to AC5, consistent with the ability of the protein to perform coincidence detection effectively. Our calculations also pointed to strong similarities between AC5 and other AC isoforms, including AC1 and AC6. Findings from the molecular simulations were used along with experimental data as constraints for systems biology modeling of a specific AC5-triggered neuronal cascade to investigate how the dynamics of downstream signaling depend on initial receptor activation, Jeanette Hellgren Kotaleski, Paolo Carloni, Rebecca C. Wade: Horizon 2020 Framework Programme (945539, HBP SGA3); Jeanette Hellgren Kotaleski: Swedish Research Council (VR-M-2017-02806, VR-M-2020-01652); Swedish e-science Research Center (SeRC). Francesco Colizzi is a Ramón y Cajal Fellow (RYC2019-026768-I). Rebecca C. Wade: Klaus Tschira Foundation. [...] The ICM-CSIC is recipient of the Severo Ochoa Centre of Excellence accreditation (CEX2019-000928-S) from the Spanish Ministry of Science and Innovation.

Increased glucose metabolism of inflammatory tissues is the main source of false-positive F-18-FDG PET findings in oncology. It has been suggested that radiolabeled nucleosides might be more tumor specific. Methods: To test this hypothesis, we compared the biodistribution of 3'-deoxy-3'-F-18-fluorothymidine (FLT) and F-18-FDG in Wistar rats that bore tumors (C6 rat glioma in the right shoulder) and also had sterile inflammation in the left calf muscle (induced by injection of 0.1 mL of turpentine). Twenty-four hours after turpentine injection, the rats received an intravenous bolus (30 MBq) of either F-18-FLT (n = 5) or F-18-FDG (n = 5). Pretreatment of the animals with thymidine phosphorylase (>1,000 U/kg, intravenously) before injection of F-18-FLT proved to be necessary to reduce the serum levels of endogenous thymidine and achieve satisfactory tumor uptake of radioactivity. Results: Tumor-to-muscle ratios of F-18-FDG at 2 h after injection (13.2 +/- 3.0) were higher than those of F-18-FLT (3.8 +/- 1.3). F-18-FDG showed high physiologic uptake in brain and heart, whereas F-18-FLT was avidly taken up by bone marrow. F-18-FDG accumulated in the inflamed muscle, with 4.8 +/- 1.2 times higher uptake in the affected thigh than in the contralateral healthy thigh, in contrast to F-18-FLT, for which this ratio was not significantly different from unity (1.3 +/- 0.4). Conclusion; In F-18-FDG PET images, both tumor and inflammation were visible, but F-18-FLT PET showed only the tumor. Thus, the hypothesis that F-18-FLT has a higher tumor specificity was confirmed in our animal model.

In this tutorial review, we compare and contrast the chemical mechanisms of electrophile/oxidant sensing, and the molecular mechanisms of signal propagation. We critically analyze biological systems in which these different pathways are believed to be manifest and what the data really mean. Finally, we discuss applications of this knowledge to disease treatment and drug development.