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Sample records for high binding constant

  1. Simultaneous Determination of Binding Constants for Multiple Carbohydrate Hosts in Complex Mixtures

    DEFF Research Database (Denmark)

    Meier, Sebastian; Beeren, Sophie

    2014-01-01

    to determine binding constants for all other detectable and resolvable hosts. With the use of high-resolution 1H−13C HSQC experiments, complexes of amphiphiles with more than 10 different maltooligosaccharides can be resolved. Hereby, the binding capabilities of a set of structurally related hosts can...

  2. Kinetic evaluation of highly active supported gold catalysts prepared from monolayer-protected clusters: an experimental Michaelis-Menten approach for determining the oxygen binding constant during CO oxidation catalysis.

    Science.gov (United States)

    Long, Cormac G; Gilbertson, John D; Vijayaraghavan, Ganesh; Stevenson, Keith J; Pursell, Christopher J; Chandler, Bert D

    2008-08-06

    Thiol monolayer-protected Au clusters (MPCs) were prepared using dendrimer templates, deposited onto a high-surface-area titania, and then the thiol stabilizers were removed under H2/N2. The resulting Au catalysts were characterized with transmission electron microscopy, X-ray photoelectron spectroscopy, and infrared spectroscopy of adsorbed CO. The Au catalysts prepared via this route displayed minimal particle agglomeration during the deposition and activation steps. Structural data obtained from the physical characterization of the Au catalysts were comparable to features exhibited from a traditionally prepared standard Au catalyst obtained from the World Gold Council (WGC). A differential kinetic study of CO oxidation catalysis by the MPC-prepared Au and the standard WGC catalyst showed that these two catalyst systems have essentially the same reaction order and Arrhenius apparent activation energies (28 kJ/mol). However, the MPC-prepared Au catalyst shows 50% greater activity for CO oxidation. Using a Michaelis-Menten approach, the oxygen binding constants for the two catalyst systems were determined and found to be essentially the same within experimental error. To our knowledge, this kinetic evaluation is the first experimental determination of oxygen binding by supported Au nanoparticle catalysts under working conditions. The values for the oxygen binding equilibrium constant obtained from the Michaelis-Menten treatment (ca. 29-39) are consistent with ultra-high-vacuum measurements on model catalyst systems and support density functional theory calculations for oxygen binding at corner or edge atoms on Au nanoparticles and clusters.

  3. Palmitate and stearate binding to human serum albumin. Determination of relative binding constants

    DEFF Research Database (Denmark)

    Vorum, H; Fisker, K; Honoré, B

    1997-01-01

    . The experimental data were analysed by a computerised curve fitting procedure using equilibrium equations for multiple binding of ligands, containing relative binding constants, valid whether the ligands are truly insoluble or are slightly soluble and irrespective of aggregation in aqueous solution. A best-fit set...... of relative binding constants was found, and subsequently 30 sets of acceptable constants for each set of data in order to evaluate the variation. The data were first fitted by the relative Scatchard's equation, then by the relative, stoichiometric equation. Scatchard's equation is deduced on the presumption...... that cooperativity is absent while the stoichiometric equation is valid even when cooperativity is present. It was found with palmitate as well as with stearate that the two equations fitted the data equally well, and it was concluded that the observations were compatible with absence of cooperativity. The relative...

  4. Determining the binding mode and binding affinity constant of tyrosine kinase inhibitor PD153035 to DNA using optical tweezers

    Energy Technology Data Exchange (ETDEWEB)

    Cheng, Chih-Ming [School of Dental Technology, Taipei Medical University, Taipei 110, Taiwan (China); Research Center for Biomedical Implants and Microsurgery Devices, Taipei Medical University, Taipei 110, Taiwan (China); Department of Biomedical Engineering and Environmental Sciences, National Tsing Hua University, Hsinchu 30043, Taiwan (China); Lee, Yuarn-Jang [Section of Infectious Diseases, Department of Internal Medicine, Taipei Medical University Hospital, Taipei 110, Taiwan (China); Wang, Wei-Ting [School of Dental Technology, Taipei Medical University, Taipei 110, Taiwan (China); Institute of Biomedical Materials and Engineering, Taipei Medical University, Taipei 110, Taiwan (China); Research Center for Biomedical Implants and Microsurgery Devices, Taipei Medical University, Taipei 110, Taiwan (China); Hsu, Chien-Ting [Institute of Biomedical Materials and Engineering, Taipei Medical University, Taipei 110, Taiwan (China); Research Center for Biomedical Implants and Microsurgery Devices, Taipei Medical University, Taipei 110, Taiwan (China); Tsai, Jing-Shin [School of Dental Technology, Taipei Medical University, Taipei 110, Taiwan (China); Institute of Biomedical Materials and Engineering, Taipei Medical University, Taipei 110, Taiwan (China); Research Center for Biomedical Implants and Microsurgery Devices, Taipei Medical University, Taipei 110, Taiwan (China); Wu, Chien-Ming [Department of Biomedical Engineering and Environmental Sciences, National Tsing Hua University, Hsinchu 30043, Taiwan (China); Ou, Keng-Liang [Institute of Biomedical Materials and Engineering, Taipei Medical University, Taipei 110, Taiwan (China); Research Center for Biomedical Implants and Microsurgery Devices, Taipei Medical University, Taipei 110, Taiwan (China); and others

    2011-01-07

    Research highlights: {yields} PD153035 is a DNA intercalator and intercalation occurs only under very low salt concentration. {yields} The minimum distance between adjacent bound PD153035 {approx} 11 bp. {yields} Binding affinity constant for PD153035 is 1.18({+-}0.09) x 10{sup 4} (1/M). {yields} The change of binding free energy of PD153035-DNA interaction is -5.49 kcal mol{sup -1} at 23 {+-} 0.5 {sup o}C. -- Abstract: Accurately predicting binding affinity constant (K{sub A}) is highly required to determine the binding energetics of the driving forces in drug-DNA interactions. Recently, PD153035, brominated anilinoquinazoline, has been reported to be not only a highly selective inhibitor of epidermal growth factor receptor but also a DNA intercalator. Here, we use a dual-trap optical tweezers to determining K{sub A} for PD153035, where K{sub A} is determined from the changes in B-form contour length (L) of PD153035-DNA complex. Here, L is fitted using a modified wormlike chain model. We found that a noticeable increment in L in 1 mM sodium cacodylate was exhibited. Furthermore, our results showed that K{sub A} = 1.18({+-}0.09) x 10{sup 4} (1/M) at 23 {+-} 0.5 {sup o}C and the minimum distance between adjacent bound PD153035 {approx} 11 bp. We anticipate that by using this approach we can determine the complete thermodynamic profiles due to the presence of DNA intercalators.

  5. Binding constants of phenylalanine for the four mononucleotides

    Science.gov (United States)

    Khaled, M. A.; Mullins, D. W., Jr.; Lacey, J. C., Jr.

    1984-01-01

    Earlier work has shown that several properties of amino acids correlate directly with properties of their anticodonic nucleotides. Furthermore, in precipitation studies with thermal proteinoids and homopolyribonucleotides, an anticodonic preference was displayed between Lys-rich, Pro-rich and Gly-rich thermal proteinoids and their anticodonic polyribonucleotides. However, Phe-rich thermal proteinoid displayed a preference for its codonic nucleotide, poly U. This inconsistency seemed to be explained by a folding in of the hydrophobic residues of Phe causing the proteinoid to appear more hydrophilic. The present work used nuclear magnetic resonance techniques to resolve a limited question: to which of the four nucleotides does Phe bind most strongly? The results show quite clearly that Phe binds most strongly to its anticodonic nucleotide, AMP.

  6. Colossal dielectric constant in high entropy oxides

    Energy Technology Data Exchange (ETDEWEB)

    Berardan, David; Franger, Sylvain; Dragoe, Diana; Meena, Arun Kumar; Dragoe, Nita [ICMMO (UMR 8182 CNRS), Universite Paris-Sud, Universite Paris-Saclay, 91405, Orsay (France)

    2016-04-15

    Entropic contributions to the stability of solids are very well understood and the mixing entropy has been used for forming various solids, for instance such as inverse spinels, see Nawrotsky et al., J. Inorg. Nucl. Chem. 29, 2701 (1967) [1]. A particular development was related to high entropy alloys by Yeh et al., Adv. Eng. Mater. 6, 299 (2004) [2] and Cantor et al., Mater. Sci. Eng. A 375-377, 213 (2004) [3] (for recent reviews see Zhang et al., Prog. Mater. Sci. 61, 1 (2014) [4] and Tsai et al., Mater. Res. Lett. 2, 107 (2014) [5]) in which the configurational disorder is responsible for forming simple solid solutions and which are thoroughly studied for various applications especially due to their mechanical properties, e.g. Gludovatz et al., Science 345, 1153 (2014) [6] and Lu et al., Sci. Rep. 4, 6200 (2014) [7], but also electrical properties, Kozelj et al., Phys. Rev. Lett. 113, 107001 (2014) [8], hydrogen storage, Kao et al., Int. J. Hydrogen Energy 35, 9046 (2010) [9], magnetic properties, Zhang et al., Sci. Rep. 3, 1455 (2013) [10]. Many unexplored compositions and properties still remain for this class of materials due to their large phase space. In a recent report it has been shown that the configurational disorder can be used for stabilizing simple solid solutions of oxides, which should normally not form solid solutions, see Rost et al., Nature Commun. 6, 8485 (2015) [11] these new materials were called ''entropy-stabilized oxides''. In this pioneering report, it was shown that mixing five equimolar binary oxides yielded, after heating at high temperature and quenching, an unexpected rock salt structure compound with statistical distribution of the cations in a face centered cubic lattice. Following this seminal study, we show here that these high entropy oxides (named HEOx hereafter) can be substituted by aliovalent elements with a charge compensation mechanism. This possibility largely increases the potential development of new

  7. The constant region affects antigen binding of antibodies to DNA by altering secondary structure.

    Science.gov (United States)

    Xia, Yumin; Janda, Alena; Eryilmaz, Ertan; Casadevall, Arturo; Putterman, Chaim

    2013-11-01

    We previously demonstrated an important role of the constant region in the pathogenicity of anti-DNA antibodies. To determine the mechanisms by which the constant region affects autoantibody binding, a panel of isotype-switch variants (IgG1, IgG2a, IgG2b) was generated from the murine PL9-11 IgG3 autoantibody. The affinity of the PL9-11 antibody panel for histone was measured by surface plasmon resonance (SPR). Tryptophan fluorescence was used to determine wavelength shifts of the antibody panel upon binding to DNA and histone. Finally, circular dichroism spectroscopy was used to measure changes in secondary structure. SPR analysis revealed significant differences in histone binding affinity between members of the PL9-11 panel. The wavelength shifts of tryptophan fluorescence emission were found to be dependent on the antibody isotype, while circular dichroism analysis determined that changes in antibody secondary structure content differed between isotypes upon antigen binding. Thus, the antigen binding affinity is dependent on the particular constant region expressed. Moreover, the effects of antibody binding to antigen were also constant region dependent. Alteration of secondary structures influenced by constant regions may explain differences in fine specificity of anti-DNA antibodies between antibodies with similar variable regions, as well as cross-reactivity of anti-DNA antibodies with non-DNA antigens.

  8. Binding constants of membrane-anchored receptors and ligands depend strongly on the nanoscale roughness of membranes.

    Science.gov (United States)

    Hu, Jinglei; Lipowsky, Reinhard; Weikl, Thomas R

    2013-09-17

    Cell adhesion and the adhesion of vesicles to the membranes of cells or organelles are pivotal for immune responses, tissue formation, and cell signaling. The adhesion processes depend sensitively on the binding constant of the membrane-anchored receptor and ligand proteins that mediate adhesion, but this constant is difficult to measure in experiments. We have investigated the binding of membrane-anchored receptor and ligand proteins with molecular dynamics simulations. We find that the binding constant of the anchored proteins strongly decreases with the membrane roughness caused by thermally excited membrane shape fluctuations on nanoscales. We present a theory that explains the roughness dependence of the binding constant for the anchored proteins from membrane confinement and that relates this constant to the binding constant of soluble proteins without membrane anchors. Because the binding constant of soluble proteins is readily accessible in experiments, our results provide a useful route to compute the binding constant of membrane-anchored receptor and ligand proteins.

  9. Modeling Nonlinear Adsorption with a Single Chemical Parameter: Predicting Chemical Median Langmuir Binding Constants.

    Science.gov (United States)

    Davis, Craig Warren; Di Toro, Dominic M

    2015-07-07

    Procedures for accurately predicting linear partition coefficients onto various sorbents (e.g., organic carbon, soils, clay) are reliable and well established. However, similar procedures for the prediction of sorption parameters of nonlinear isotherm models are not. The purpose of this paper is to present a procedure for predicting nonlinear isotherm parameters, specifically the median Langmuir binding constants, K̃L, obtained utilizing the single-chemical parameter log-normal Langmuir isotherm developed in the accompanying work. A reduced poly parameter linear free energy relationship (pp-LFER) is able to predict median Langmuir binding constants for graphite, charcoal, and Darco granular activated carbon (GAC) adsorption data. For the larger F400 GAC data set, a single pp-LFER model was insufficient, as a plateau is observed for the median Langmuir binding constants of larger molecular volume sorbates. This volumetric cutoff occurs in proximity to the median pore diameter for F400 GAC. A log-linear relationship exists between the aqueous solubility of these large compounds and their median Langmuir binding constants. Using this relationship for the chemicals above the volumetric cutoff and the pp-LFER below the cutoff, the median Langmuir binding constants can be predicted with a root-mean square error for graphite (n = 13), charcoal (n = 11), Darco GAC (n = 14), and F400 GAC (n = 44) of 0.129, 0.307, 0.407, and 0.424, respectively.

  10. THE USE OF HAMMETT CONSTANTS TO UNDERSTAND THE NON-COVALENT BINDING OF AROMATICS

    Directory of Open Access Journals (Sweden)

    Michael Lewis

    2012-04-01

    Full Text Available Non-covalent interactions of aromatics are important in a wide range of chemical and biological applications. The past two decades have seen numerous reports of arene-arene binding being understood in terms Hammett substituent constants, and similar analyses have recently been extended to cation-arene and anion-arene binding. It is not immediately clear why electrostatic Hammett parameters should work so well in predicting the binding for all three interactions, given that different intermolecular forces dominate each interaction. This review explores such anomalies, and summarizes how Hammett substituent constants have been employed to understand the non-covalent binding in arene-arene, cation-arene and anion-arene interactions.

  11. Prediction and dissection of widely-varying association rate constants of actin-binding proteins.

    Directory of Open Access Journals (Sweden)

    Xiaodong Pang

    Full Text Available Actin is an abundant protein that constitutes a main component of the eukaryotic cytoskeleton. Its polymerization and depolymerization are regulated by a variety of actin-binding proteins. Their functions range from nucleation of actin polymerization to sequestering G-actin in 1∶1 complexes. The kinetics of forming these complexes, with rate constants varying at least three orders of magnitude, is critical to the distinct regulatory functions. Previously we have developed a transient-complex theory for computing protein association mechanisms and association rate constants. The transient complex refers to an intermediate in which the two associating proteins have near-native separation and relative orientation but have yet to form short-range specific interactions of the native complex. The association rate constant is predicted as k(a = k(a0 e(-ΔG(el*/k(BT, where k(a0 is the basal rate constant for reaching the transient complex by free diffusion, and the Boltzmann factor captures the bias of long-range electrostatic interactions. Here we applied the transient-complex theory to study the association kinetics of seven actin-binding proteins with G-actin. These proteins exhibit three classes of association mechanisms, due to their different molecular shapes and flexibility. The 1000-fold k(a variations among them can mostly be attributed to disparate electrostatic contributions. The basal rate constants also showed variations, resulting from the different shapes and sizes of the interfaces formed by the seven actin-binding proteins with G-actin. This study demonstrates the various ways that actin-binding proteins use physical properties to tune their association mechanisms and rate constants to suit distinct regulatory functions.

  12. Absorption spectroscopy and binding constants for first-row transition metal complexes of a DOPA-containing peptide.

    Science.gov (United States)

    Sever, Mary J; Wilker, Jonathan J

    2006-02-14

    A diverse array of biological systems incorporate 3,4-dihydroxyphenlyalanine (DOPA) into proteins and small molecules for cross-linking and material generation. Marine worm eggshells, sea squirt wound plugs, and marine mussel adhesives may all be formed by combining DOPA-containing molecules with high levels of metals. In order to provide model systems for characterizing these biomaterials, we carried out a study on metal binding to a DOPA-containing peptide. Ultraviolet-visible absorption spectra are presented for the AdopaTP peptide binding to Fe3+, V3+, VO2+, Mn3+, Ti4+, Cu2+, Co2+, and Ni2+ in mono, bis, and where applicable, tris coordination modes. Association constants were determined for selected metal ions binding to the peptide. In general, the spectroscopic and binding properties of this DOPA-containing peptide were found to be similar to those of catechol.

  13. Constants for mercury binding by organic matter isolates from the Florida Everglades

    Science.gov (United States)

    Benoit, J.M.; Mason, R.P.; Gilmour, C.C.; Aiken, G.R.

    2001-01-01

    Dissolved organic matter (DOM) has been implicated as an important complexing agent for Hg that can affect its mobility and bioavailability in aquatic ecosystems. However, binding constants for natural Hg-DOM complexes are not well known. We employed a competitive ligand approach to estimate conditional stability constants for Hg complexes with DOM isolates collected from Florida Everglades surface waters. The isolates examined were the hydrophobic fraction of DOM from a eutrophic, sulfidic site (F1-HPoA) and the hydrophilic fraction from an oligotrophic, low-sulfide site (2BS-HPiA). Our experimental determinations utilized overall octanol-water partitioning coefficients (Dow) for 203Hg at 0.01 M chloride and across pH and DOM concentration gradients. Use of this radioisotope allowed rapid determinations of Hg concentrations in both water and octanol phases without problems of matrix interference. Conditional stability constants (1 = 0.06, 23??C) were log K??? = 11.8 for F1-HPoA and log K' = 10.6 for 2BS-HPiA. These are similar to previously published stability constants for Hg binding to low-molecular-weight thiols. Further, F1-HPoA showed a pH-dependent decline in Dow that was consistent with models of Hg complexation with thiol groups as the dominant Hg binding sites in DOM. These experiments demonstrate that the DOM isolates are stronger ligands for Hg than chloride ion or ethylenediamine-tetraacetic acid. Speciation calculations indicate that at the DOM concentrations frequently measured in Everglades, 20 to 40 ??M, significant complexation of Hg by DOM would be expected in aerobic (sulfide-free) surface waters. Copyright ?? 2001 Elsevier Science Ltd.

  14. Surface tension method for determining binding constants for cyclodextrin inclusion complexes of ionic surfactants

    Energy Technology Data Exchange (ETDEWEB)

    Dharmawardana, U.R.; Christian, S.D.; Tucker, E.E.; Taylor, R.W.; Scamehorn, J.F. (Univ. of Oklahoma, Norman, OK (United States))

    1993-09-01

    A new method has been developed for determining binding constants of complexes of cyclodextrins with surface-active compounds, including water-soluble ionic surfactants. The technique requires measuring the change in surface tension caused by addition of a cyclodextrin (CD) to aqueous solutions of the surfactant; the experimental results lead directly to inferred values of the thermodynamic activity of the surfactant. Surface tension results are reported for three different surfactants sodium dodecyl sulfate (SDS), cetylpyridinium chloride (CPC), and cetyltrimethylammonium bromide (CTAB) in the presence and in the absence of added [beta]-CD. Data for CPC have been obtained at surfactant concentrations below and above the critical micelle concentration. Correlations between surface tension and surfactant activity are expressed by the Szyszkowski equation, which subsumes the Langmuir adsorption model and the Gibbs equation. It is observed that the surface tension increases monotonically as [beta]-cyclodextrin is added to ionic surfactant solutions. At concentrations of CD well in excess of the surfactant concentration, the surface tension approaches that of pure water, indicating that neither the surfactant-CD complexes nor CD itself are surface active. Binding constants are inferred from a model that incorporates the parameters of the Szyszkowski equation and mass action constants relating to the formation of micelles from monomers of the surfactant and the counterion. Evidence is given that two molecules of CD can complex the C-16 hydrocarbon chain of the cetyl surfactants. 30 refs., 5 figs., 1 tab.

  15. High precision fundamental constants at the TeV scale

    CERN Document Server

    Moch, S; Alekhin, S; Blumlein, J; de la Cruz, L; Dittmaier, S; Dowling, M; Erler, J; Espinosa, J R; Fuster, J; Tormo, X Garcia i; Hoang, A H; Huss, A; Kluth, S; Mulders, M; Papanastasiou, A S; Piclum, J; Rabbertz, K; Schwinn, C; Schulze, M; Shintani, E; Uwer, P; Zerf, N

    2014-01-01

    This report summarizes the proceedings of the 2014 Mainz Institute for Theoretical Physics (MITP) scientific program on "High precision fundamental constants at the TeV scale". The two outstanding parameters in the Standard Model dealt with during the MITP scientific program are the strong coupling constant $\\alpha_s$ and the top-quark mass $m_t$. Lacking knowledge on the value of those fundamental constants is often the limiting factor in the accuracy of theoretical predictions. The current status on $\\alpha_s$ and $m_t$ has been reviewed and directions for future research have been identified.

  16. Kinetic mechanism of phenylalanine hydroxylase: intrinsic binding and rate constants from single-turnover experiments.

    Science.gov (United States)

    Roberts, Kenneth M; Pavon, Jorge Alex; Fitzpatrick, Paul F

    2013-02-12

    Phenylalanine hydroxylase (PheH) catalyzes the key step in the catabolism of dietary phenylalanine, its hydroxylation to tyrosine using tetrahydrobiopterin (BH(4)) and O(2). A complete kinetic mechanism for PheH was determined by global analysis of single-turnover data in the reaction of PheHΔ117, a truncated form of the enzyme lacking the N-terminal regulatory domain. Formation of the productive PheHΔ117-BH(4)-phenylalanine complex begins with the rapid binding of BH(4) (K(d) = 65 μM). Subsequent addition of phenylalanine to the binary complex to form the productive ternary complex (K(d) = 130 μM) is approximately 10-fold slower. Both substrates can also bind to the free enzyme to form inhibitory binary complexes. O(2) rapidly binds to the productive ternary complex; this is followed by formation of an unidentified intermediate, which can be detected as a decrease in absorbance at 340 nm, with a rate constant of 140 s(-1). Formation of the 4a-hydroxypterin and Fe(IV)O intermediates is 10-fold slower and is followed by the rapid hydroxylation of the amino acid. Product release is the rate-determining step and largely determines k(cat). Similar reactions using 6-methyltetrahydropterin indicate a preference for the physiological pterin during hydroxylation.

  17. Partition coefficient vs. binding constant: How best to assess molecular lipophilicity.

    Science.gov (United States)

    Cevc, Gregor

    2015-05-01

    Partition coefficient, P, is the preferred descriptor of molecular lipo- or hydrophilicity, and thus of relationships between a solute (S, e.g., a drug), a polar medium (W, e.g., an aqueous buffer), and an essentially apolar, organic, medium or a drug carrier (O). The coefficient is commonly identified with the linear ratio of solute quantities in the two media, P=nSO/nSW, even to characterise solute association with or binding to a surface (e.g., of a HPLC column or a drug carrier). To check the latter practice correctness-and credibility of the prevailing P definition-this paper compares an ideal solute distribution between two separate homogeneous fluid media (i.e., partitioning) to solute association with a uniform surface immersed in one such medium (i.e., binding). This reveals that solute partitioning and binding fundamentally differ and can only exceptionally be described, or analysed, with similar equations. Nonlinearised formulae that describe partitioning (Eq. (9)) and binding (Eq. (11)) can yield similar lipophilicity descriptor values only if solute preparation is relatively dilute; employing a large organic medium fraction is helpful in this respect. Additional prerequisites for partitioning and binding models match are: 1:1 stoichiometry at the association maximum and identical interfacial area of solute and organic medium molecules. If these requirements are not met, binding model yields different, potentially somewhat higher, but more often up to >10 times lower results than partitioning model. The main reason is saturation of organic medium with solute molecules. Partitioning model excludes this phenomenon, which cannot always be prevented by focussing on dilute solute preparations. The current practice of using a linear model and approximate equations to study partitioning or binding can cause large errors (>10(3)×), and is one possible reason for the notoriously high experimental logP values scattering. The latter makes logP predictions more

  18. Synthetic Strategies for High Dielectric Constant Silicone Elastomers

    DEFF Research Database (Denmark)

    Madsen, Frederikke Bahrt

    Dielectric electroactive polymers (DEAPs) are a new and promising transducer technology and are often referred to as ‘artificial muscles’, due to their ability to undergo large deformations when stimulated by electric fields. DEAPs consist of a soft and thin elastomeric film (an elastomer...... synthetic strategies were developed in this Ph.D. thesis, in order to create silicone elastomers with high dielectric constants and thereby higher energy densities. The work focused on maintaining important properties such as dielectric loss, electrical breakdown strength and elastic modulus....... The methodology therefore involved chemically grafting high dielectric constant chemical groups onto the elastomer network, as this would potentially provide a stable elastomer system upon continued activation of the material. The first synthetic strategy involved the synthesis of a new type of cross...

  19. Ferroelectric metal-organic framework with a high dielectric constant.

    Science.gov (United States)

    Ye, Qiong; Song, Yu-Mei; Wang, Guo-Xi; Chen, Kai; Fu, Da-Wei; Chan, Philip Wai Hong; Zhu, Jin-Song; Huang, Songping D; Xiong, Ren-Gen

    2006-05-24

    Hydrothermal reaction of (l)-N-(4'-cyanobenzy)-(S)-proline with CdCl2 as a Lewis acid catalyst and NaN3 gives colorless block compound 1, in which 1 displays a complicated 3D framework. Ferroelectric and dielectric property measurements reveal that 1 exhibits physical properties comparable to that of a typical ferroelectric compound with a dipole relaxation process and a dielectric constant of ca. 38.6 that makes it, by definition, a high dielectric material.

  20. Optical constants of refractory oxides at high temperatures

    CERN Document Server

    Zeidler, Simon; Mutschke, Harald; 10.1051/0004-6361/201220459

    2013-01-01

    Many cosmic dust species, among them refractory oxides, form at temperatures higher than 300 K. Nevertheless, most astrophysical studies are based on the room-temperature optical constants of solids, such as corundum and spinel. A more realistic approach is needed for these materials, especially in the context of modeling late-type stars. We aimed at deriving sets of optical constants of selected, astrophysically relevant oxide dust species with high melting points. A high-temperature-high-pressure-cell and a Fourier-transform spectrometer were used to measure reflectance spectra of polished samples. For corundum (alpha-Al$_2$O$_3$), spinel (MgAl$_2$O$_4$), and alpha-quartz (SiO$_2$), temperature-dependent optical constants were measured from 300 K up to more than 900 K. Small particle spectra were also calculated from these data. All three examined oxides show a significant temperature dependence of their mid-IR bands. For the case of corundum, we find that the 13$\\mu$m emission feature - seen in the IR spec...

  1. PMN-PT nanowires with a very high piezoelectric constant.

    Science.gov (United States)

    Xu, Shiyou; Poirier, Gerald; Yao, Nan

    2012-05-09

    A profound way to increase the output voltage (or power) of the piezoelectric nanogenerators is to utilize a material with higher piezoelectric constants. Here we report the synthesis of novel piezoelectric 0.72Pb(Mg(1/3)Nb(2/3))O(3)-0.28PbTiO(3) (PMN-PT) nanowires using a hydrothermal process. The unpoled single-crystal PMN-PT nanowires show a piezoelectric constant (d(33)) up to 381 pm/V, with an average value of 373 ± 5 pm/V. This is about 15 times higher than the maximum reported value of 1-D ZnO nanostructures and 3 times higher than the largest reported value of 1-D PZT nanostructures. These PMN-PT nanostructures are of good potential being used as the fundamental building block for higher power nanogenerators, high sensitivity nanosensors, and large strain nanoactuators.

  2. Comparative study of binding constants from Love wave surface acoustic wave and surface plasmon resonance biosensors using kinetic analysis.

    Science.gov (United States)

    Lee, Sangdae; Kim, Yong-Il; Kim, Ki-Bok

    2013-11-01

    Biosensors are used in a variety of fields for early diagnosis of diseases, measurement of toxic contaminants, quick detection of pathogens, and separation of specific proteins or DNA. In this study, we fabricated and evaluated the capability of a high sensitivity Love wave surface acoustic wave (SAW) biosensor. The experimental setup was composed of the fabricated 155-MHz Love wave SAW biosensor, a signal measurement system, a liquid flow system, and a temperature-control system. Subsequently, we measured the lower limit of detection (LOD) of the 155-MHz Love wave SAW biosensor, and calculated the association and dissociation constants between protein G and anti-mouse IgG using kinetic analysis. We compared these results with those obtained using a commercial surface plasmon resonance (SPR) biosensor. We found that the LOD of the SAW biosensor for anti-mouse IgG and mouse IgG was 0.5 and 1 microg/ml, respectively, and the resultant equilibrium association and dissociation constants were similar to the corresponding values obtaining using the commercial SPR biosensor. Thus, we conclude that the fabricated 155-MHz Love wave SAW biosensor exhibited the high sensitivity of the commercial SPR biosensor and was able to analyze the binding properties of the ligand and receptor by kinetic analysis similarly to the commercial SPR biosensor.

  3. High-temperature rate constant measurements for OH+xylenes

    KAUST Repository

    Elwardani, Ahmed Elsaid

    2015-06-01

    The overall rate constants for the reactions of hydroxyl (OH) radicals with o-xylene (k 1), m-xylene (k 2), and p-xylene (k 3) were measured behind reflected shock waves over 890-1406K at pressures of 1.3-1.8atm using OH laser absorption near 306.7nm. Measurements were performed under pseudo-first-order conditions. The measured rate constants, inferred using a mechanism-fitting approach, can be expressed in Arrhenius form as:k1=2.93×1013exp(-1350.3/T)cm3mol-1s-1(890-1406K)k2=3.49×1013exp(-1449.3/T)cm3mol-1s-1(906-1391K)k3=3.5×1013exp(-1407.5/T)cm3mol-1s-1(908-1383K)This paper presents, to our knowledge, first high-temperature measurements of the rate constants of the reactions of xylene isomers with OH radicals. Low-temperature rate-constant measurements by Nicovich et al. (1981) were combined with the measurements in this study to obtain the following Arrhenius expressions, which are applicable over a wider temperature range:k1=2.64×1013exp(-1181.5/T)cm3mol-1s-1(508-1406K)k2=3.05×109exp(-400/T)cm3mol-1s-1(508-1391K)k3=3.0×109exp(-440/T)cm3mol-1s-1(526-1383K) © 2015 The Combustion Institute.

  4. Exchanging murine and human immunoglobulin constant chains affects the kinetics and thermodynamics of antigen binding and chimeric antibody autoreactivity.

    Directory of Open Access Journals (Sweden)

    Marcela Torres

    Full Text Available Mouse-human chimeric antibodies composed of murine variable (V and human (C chains are useful therapeutic reagents. Consequently, we investigated whether heterologous C-regions from mice and humans affected specificity and affinity, and determined the contribution of C(H glycosylation to antigen binding. The interaction of a 12-mer peptide mimetic with monoclonal antibody (mAb 18B7 to Cryptococcus neoformans glucuronoxylomannan, and its chimeric (ch and deglycosylated forms were studied by surface plasmon resonance. The equilibrium and rate association constants for the chAb were higher than for mAb 18B7. V region affinity was not affected by C(H region glycosylation whereas heterologous C region of the same isotype altered the Ab binding affinity and the specificity for self-antigens. Structural models displayed local differences that implied changes on the connectivity of residues. These findings suggest that V region conformational changes can be dictated by the C(H domains through an allosteric effect involving networks of highly connected amino acids.

  5. Relaxor behavior and electrical properties of high dielectric constant materials

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    Several typical high dielectric constant materials are reviewed to study the electrical properties and relaxation mechanism. It is found that a Lorenz-type law can be used to describe the dielectric permit- tivity of either the normal ferroelectrics with or without diffuse phase transitions (DPT) or the typical ferroelectric relaxors. The ferroelectric DPT can be well described by just one fitting process using the Lorenz-type law, while the relaxor ferroelectric transition needs two independent fitting processes. The Lorenz-type law fails at the low temperature side of the dielectric maximum of a first-order ferroelectric phase transition. Above the transition temperature, the dielectric curves of all the studied materials can be well described by a Lorenz-type law.

  6. Relaxor behavior and electrical properties of high dielectric constant materials

    Institute of Scientific and Technical Information of China (English)

    FAN HuiQing; KE ShanMing

    2009-01-01

    Several typical high dielectric constant materials are reviewed to study the electrical properties and relaxation mechanism.It is found that a Lorenz-type law can be used to describe the dielectric permitUvity of either the normal ferroelectrics with or without diffuse phase transitions(DPT)or the typical ferroelectric relaxors.The ferroelectric DPT can be well described by just one fitting process using the Lorenz-type law,while the relaxor ferroelectric transition needs two independent fitting processes.The Lorenz-type law fails at the low temperature side of the dielectric maximum of a first-order ferroelectric phase transition.Above the transition temperature,the dielectric curves of all the studied materials can be well described by a Lorenz-type law.

  7. Thermodynamics of Calcium binding to the Calmodulin N-terminal domain to evaluate site-specific affinity constants and cooperativity.

    Science.gov (United States)

    Beccia, Maria Rosa; Sauge-Merle, Sandrine; Lemaire, David; Brémond, Nicolas; Pardoux, Romain; Blangy, Stéphanie; Guilbaud, Philippe; Berthomieu, Catherine

    2015-07-01

    Calmodulin (CaM) is an essential Ca(II)-dependent regulator of cell physiology. To understand its interaction with Ca(II) at a molecular level, it is essential to examine Ca(II) binding at each site of the protein, even if it is challenging to estimate the site-specific binding properties of the interdependent CaM-binding sites. In this study, we evaluated the site-specific Ca(II)-binding affinity of sites I and II of the N-terminal domain by combining site-directed mutagenesis and spectrofluorimetry. The mutations had very low impact on the protein structure and stability. We used these binding constants to evaluate the inter-site cooperativity energy and compared it with its lower limit value usually reported in the literature. We found that site I affinity for Ca(II) was 1.5 times that of site II and that cooperativity induced an approximately tenfold higher affinity for the second Ca(II)-binding event, as compared to the first one. We further showed that insertion of a tryptophan at position 7 of site II binding loop significantly increased site II affinity for Ca(II) and the intra-domain cooperativity. ΔH and ΔS parameters were studied by isothermal titration calorimetry for Ca(II) binding to site I, site II and to the entire N-terminal domain. They showed that calcium binding is mainly entropy driven for the first and second binding events. These findings provide molecular information on the structure-affinity relationship of the individual sites of the CaM N-terminal domain and new perspectives for the optimization of metal ion binding by mutating the EF-hand loops sequences.

  8. Estimation of apparent binding constant of complexes of selected acyclic nucleoside phosphonates with β-cyclodextrin by affinity capillary electrophoresis.

    Science.gov (United States)

    Šolínová, Veronika; Mikysková, Hana; Kaiser, Martin Maxmilián; Janeba, Zlatko; Holý, Antonín; Kašička, Václav

    2016-01-01

    Affinity capillary electrophoresis (ACE) has been applied to estimation of apparent binding constant of complexes of (R,S)-enantiomers of selected acyclic nucleoside phosphonates (ANPs) with chiral selector β-cyclodextrin (βCD) in aqueous alkaline medium. The noncovalent interactions of five pairs of (R,S)-enantiomers of ANPs-based antiviral drugs and their derivatives with βCD were investigated in the background electrolyte (BGE) composed of 35 or 50 mM sodium tetraborate, pH 10.0, and containing variable concentration (0-25 mM) of βCD. The apparent binding constants of the complexes of (R,S)-enantiomers of ANPs with βCD were estimated from the dependence of effective electrophoretic mobilities of (R,S)-enantiomers of ANPs (measured simultaneously by ACE at constant reference temperature 25°C inside the capillary) on the concentration of βCD in the BGE using different nonlinear and linear calculation methodologies. Nonlinear regression analysis provided more precise and accurate values of the binding constants and a higher correlation coefficient as compared to the regression analysis of the three linearized plots of the effective mobility dependence on βCD concentration in the BGE. The complexes of (R,S)-enantiomers of ANPs with βCD have been found to be relatively weak - their apparent binding constants determined by the nonlinear regression analysis were in the range 13.3-46.4 L/mol whereas the values from the linearized plots spanned the interval 12.3-55.2 L/mol.

  9. Measurement of Nanomolar Dissociation Constants by Titration Calorimetry and Thermal Shift Assay – Radicicol Binding to Hsp90 and Ethoxzolamide Binding to CAII

    Science.gov (United States)

    Zubrienė, Asta; Matulienė, Jurgita; Baranauskienė, Lina; Jachno, Jelena; Torresan, Jolanta; Michailovienė, Vilma; Cimmperman, Piotras; Matulis, Daumantas

    2009-01-01

    The analysis of tight protein-ligand binding reactions by isothermal titration calorimetry (ITC) and thermal shift assay (TSA) is presented. The binding of radicicol to the N-terminal domain of human heat shock protein 90 (Hsp90αN) and the binding of ethoxzolamide to human carbonic anhydrase (hCAII) were too strong to be measured accurately by direct ITC titration and therefore were measured by displacement ITC and by observing the temperature-denaturation transitions of ligand-free and ligand-bound protein. Stabilization of both proteins by their ligands was profound, increasing the melting temperature by more than 10 ºC, depending on ligand concentration. Analysis of the melting temperature dependence on the protein and ligand concentrations yielded dissociation constants equal to 1 nM and 2 nM for Hsp90αN-radicicol and hCAII-ethoxzolamide, respectively. The ligand-free and ligand-bound protein fractions melt separately, and two melting transitions are observed. This phenomenon is especially pronounced when the ligand concentration is equal to about half the protein concentration. The analysis compares ITC and TSA data, accounts for two transitions and yields the ligand binding constant and the parameters of protein stability, including the Gibbs free energy and the enthalpy of unfolding. PMID:19582223

  10. Measurement of Nanomolar Dissociation Constants by Titration Calorimetry and Thermal Shift Assay – Radicicol Binding to Hsp90 and Ethoxzolamide Binding to CAII

    Directory of Open Access Journals (Sweden)

    Vilma Michailovienė

    2009-06-01

    Full Text Available The analysis of tight protein-ligand binding reactions by isothermal titration calorimetry (ITC and thermal shift assay (TSA is presented. The binding of radicicol to the N-terminal domain of human heat shock protein 90 (Hsp90aN and the binding of ethoxzolamide to human carbonic anhydrase (hCAII were too strong to be measured accurately by direct ITC titration and therefore were measured by displacement ITC and by observing the temperature-denaturation transitions of ligand-free and ligand-bound protein. Stabilization of both proteins by their ligands was profound, increasing the melting temperature by more than 10 ºC, depending on ligand concentration. Analysis of the melting temperature dependence on the protein and ligand concentrations yielded dissociation constants equal to 1 nM and 2 nM for Hsp90aN-radicicol and hCAII-ethoxzolamide, respectively. The ligand-free and ligand-bound protein fractions melt separately, and two melting transitions are observed. This phenomenon is especially pronounced when the ligand concentration is equal to about half the protein concentration. The analysis compares ITC and TSA data, accounts for two transitions and yields the ligand binding constant and the parameters of protein stability, including the Gibbs free energy and the enthalpy of unfolding.

  11. High dielectric constant, low loss and high photocatalytic activity in Gd doped ZnO systems

    Science.gov (United States)

    Divya, N. K.; Pradyumnan, P. P.

    2017-01-01

    Enhanced photocatalytic activity and high dielectric constant values are achieved by gadolinium (Gd) doping in ZnO. The changes that happened to the wurtzite structure of ZnO on doping are depicted in detail by using x-ray diffraction spectroscopy. The chemical composition is confirmed using energy dispersive x-ray spectroscopy (EDAX). The influence of Gd incorporation in the emission spectra of ZnO is analysed from photoluminescence studies. The photocatalytic activity enhancement occurred in ZnO system on Gd doping was explored by kinetic rate analysis. The optimum incorporation of Gd has enhanced the dielectric constant value and decreased the loss of pristine. The high dielectric constant value and low loss make the system suitable for large scale of applications in microelectronics. The work also proposes large scale synthesis of highly efficient fluorescent Gd doped ZnO photocatalysts.

  12. Dielectric constant of graphene-on-polarized substrate: A tight-binding model study

    Indian Academy of Sciences (India)

    SIVABRATA SAHU; S K S PARASHAR; G C ROUT

    2017-07-01

    We report here a microscopic tight-binding theoretical study of the dynamic dielectric response of graphene-on-polarizable substrate with impurity. The Hamiltonian consists of first, second and third nearest neighbour electron hopping interactions besides doping and substrate-induced effects on graphene. We have introduced electron–electron correlation effect at A and B sublattices of graphene which is considered within Hartree–Fock mean-field approximation. The electron occupancies at both sublattices are calculated and solvedself-consistently and numerically for both up- and down-spin orientations. The polarization function appearing in the dielectric function is a two-particle Green’s function which is calculated by using Zubarev’s Green’s function technique. The temperature and optical frequency-dependent dielectric function is evaluated and compared with experimental data by varying Coulomb correlation energy, substrate-induced gap and impurity concentrations.

  13. Predicting the Strength of Anion-π Interactions of Substituted Benzenes: the Development of Anion-π Binding Substituent Constants.

    Science.gov (United States)

    Bagwill, Christina; Anderson, Christa; Sullivan, Elizabeth; Manohara, Varun; Murthy, Prithvi; Kirkpatrick, Charles C; Stalcup, Apryll; Lewis, Michael

    2016-11-23

    A computational study aimed at accurately predicting the strength of the anion-π binding of substituted benzenes is presented. The anion-π binding energies (Ebind) of 37 substituted benzenes and the parent benzene, with chloride or bromide were investigated at the MP2(full)/6-311++G** level of theory. In addition, energy decomposition analysis was performed on 27 selected chloride-arene complexes via symmetry adapted perturbation theory (SAPT), using the SAPT2+ approach. Initial efforts aimed to correlate the anion-π Ebind values with the sum of the Hammett constants σp (Σσp) or σm (Σσm), as done by others. This proved a decent approach for predicting the binding strength of aromatics with electron-withdrawing substituents. For the Cl(-)-substituted benzene Ebind values, the correlation with the Σσp and Σσm values of aromatics with electron-withdrawing groups had r(2) values of 0.89 and 0.87 respectively. For the Br(-)-substituted benzene Ebind values, the correlation with the Σσp and Σσm values of aromatics with electron-withdrawing groups had r(2) values of 0.90 and 0.87. However, adding aromatics with electron-donating substituents to the investigation caused the correlation to deteriorate. For the Cl(-)-substituted benzene complexes the correlation between Ebind values and the Hammett constants had r(2) = 0.81 for Σσp and r(2) = 0.84 for Σσm. For the Br(-)-substituted benzene complexes, the respective r(2) values were 0.71 for Σσp and 0.79 for Σσm. The deterioration in correlation upon consideration of substituted benzenes with electron-donating substituents is due to the anion-π binding energies becoming more attractive regardless of what type of substituent is added to the aromatic. A similar trend has been reported for parallel face-to-face substituted benzene-benzene binding. This is certainly counter to what electrostatic arguments would predict for trends in anion-π binding energies, and this discrepancy is further highlighted

  14. Is the electron radiation length constant at high energies?

    Science.gov (United States)

    Hansen, H D; Uggerhøj, U I; Biino, C; Ballestrero, S; Mangiarotti, A; Sona, P; Ketel, T J; Vilakazi, Z Z

    2003-07-04

    Experimental results for the radiative energy loss of 149, 207, and 287 GeV electrons in a thin Ir target are presented. From the data we conclude that at high energies the radiation length increases in accordance with the Landau-Pomeranchuk-Migdal (LPM) theory and thus electrons become more penetrating the higher the energy. The increase of the radiation length as a result of the LPM effect has a significant impact on the behavior of high-energy electromagnetic showers.

  15. High dielectric constant ceramics for ion-electron sources

    CERN Document Server

    Boscolo, I

    2002-01-01

    Ferroelectric disks, coated with proper electrodes, can easily produce a dense plasma cloud when excited with a high-voltage pulse. This plasma can be a source of either electrons or ions depending on the sign of the extracting field set in front of the disk. We present the behavior of the disks operating at high frequency as emitters of both electrons and ions in two experimental configurations: (a) without and (b) with two screening grids. These two screening grids are inserted when the plasma must be confined within the cathode region. The system is capable of providing ion pulses of a few hundred milliamperes, whose length can range from a hundred nanoseconds to dozen microseconds. The electron pulses of energetic electrons have typically an amplitude higher than a couple of amperes. Tests at MHz repetition rate were positive as for stable operation.

  16. Novel high dielectric constant hybrid elastomers based on glycerol-insilicone emulsions

    DEFF Research Database (Denmark)

    Mazurek, Piotr Stanislaw; Skov, Anne Ladegaard

    2016-01-01

    distributed within PDMS in shape of discrete droplets thus acting as a high dielectric constant filler efficiently enhancing the dielectric constant of the composites. Low- and high-voltage dielectric spectroscopy measurements were conducted in order to verify applicability of the composites as dielectric...

  17. Behavior of concrete and concrete-filled circular steel tubular stub columns at constant high temperatures

    Institute of Scientific and Technical Information of China (English)

    DING Fa-xing; YU Zhi-wu

    2006-01-01

    Based on reanalyzing test results of uniaxial compressive behavior of concrete at constant high temperatures in China,with the compressive cube strength of concrete from 20 to 80 Mpa, unified formulas for uniaxial compressive strength, elastic modulus, strain at peak uniaxial compression and mathematical expression for unaxial compressive stress-strain relations for the concrete at constant high temperatures were studied. Furthermore, the axial stress-axial strain relations between laterally confined concrete under axial compression and multiaxial stress-strain relations for steel at constant high temperatures were studied. Finally,based on continuum mechanics, the mechanics model for concentric cylinders of circular steel tube with concrete core of entire section loaded at constant high temperatures was established. Applying elasto-plastic analysis method, a FORTRAN program was developed, and the concrete-filled circular steel tubular (CFST) stub columns at constant high temperatures were analyzed. The analysis results are in agreement with the experiment ones from references.

  18. Evaluation of three neutral capillary coatings for the determination of analyte-cyclodextrin binding constants by affinity capillary electrophoresis. Application to N,N'-disubstituted piperazine derivatives.

    Science.gov (United States)

    Danel, Cécile; Melnyk, Patricia; Azaroual, Nathalie; Larchanché, Paul-Emmanuel; Goossens, Jean-François; Vaccher, Claude

    2016-07-15

    The performances of three neutral static coatings (hydroxypropyl cellulose, polyethylene oxide and poly(N,N-dimethylacrylamide) have been evaluated in order to determine the binding constants of the complexes formed between four polycationic compounds (piperazine derivatives) and four cyclodextrins of pharmaceutical interest (β-CD, HP-β-CD, Me-β-CD and sulfobutyl ether-β-CD) by affinity capillary electrophoresis. The physically-adsorbed poly(N,N-dimethylacrylamide) coating proves to be the more efficient to mask the silanol groups of the capillary wall since the lowest electroosmotic flow was measured for this coating. Moreover, it drastically reduces the adsorption of the compounds since it allows a correct repeatability of their migration time, higher efficiencies of the peaks and no baseline shift. Then, it was verified for four complexes that this coating allows a correct determination of the binding constants avoiding the CD adsorption which is responsible of an undervaluation of binding constants. The highest binding constants are obtained using the anionic sulfobutyl ether-β-CD (SBE-β-CD). The structure of the complex formed between the tacrine derivative and the SBE-β-CD was further investigated through 2D ROESY NMR experiments and structure-binding constant relationships. Results suggest that the inclusion in the SBE-β-CD cavity occurs through the aliphatic ring portion of the tacrine moiety. Copyright © 2016 Elsevier B.V. All rights reserved.

  19. Competitive reactions in solutions of poly-L-histidine, calf thymus DNA, and synthetic polyanions: determining the binding constants of polyelectrolytes.

    Science.gov (United States)

    Zelikin, Alexander N; Trukhanova, Elizabeth S; Putnam, David; Izumrudov, Vladimir A; Litmanovich, Andrey A

    2003-11-12

    to the developed procedure to yield the effective binding constants of PLH with DNA and polyanion-competitor, calculated both per mole of interacting units K(1) and mole of interacting chains K(n). In all cases, K(1) had similar numerical values reflecting common type of interaction stabilizing the complexes, i.e., electrostatics. Slight variation of K(1) yielded in drastic changes in K(n) and alteration of dominance of PLH interaction with DNA or synthetic polyanion. The results of the study can have a high impact in deriving the correlation between the binding constant of a polycation to DNA and its ability to serve as gene delivery vehicle.

  20. High-gain, high-bandwidth, rail-to-rail, constant-gm CMOS operational amplifier

    Science.gov (United States)

    Huang, Hong-Yi; Wang, Bo-Ruei

    2013-01-01

    This study presents a high-gain, high-bandwidth, constant-gm , rail-to-rail operational amplifier (op-amp). The constant transconductance is improved with a source-to-bulk bias control of an input pair. A source degeneration scheme is also adapted to the output stage for receiving wide input range without degradation of the gain. Additionally, several compensation schemes are employed to enhance the stability. A test chip is fabricated in a 0.18 µm complementary metal-oxide semiconductor process. The active area of the op-amp is 181 × 173 µm2 and it consumes a power of 2.41 mW at a supply voltage of 1.8 V. The op-amp achieves a dc gain of 94.3 dB and a bandwidth of 45 MHz when the output capacitive load is connected to an effective load of 42.5 pF. A class-AB output stage combining a slew rate (SR) boost circuit provides a sinking current of 6 mA and an SR of 17 V/µs.

  1. The effects of varied versus constant high-, medium-, and low-preference stimuli on performance

    National Research Council Canada - National Science Library

    Wine, Byron; Wilder, David A

    2009-01-01

    The purpose of the current study was to compare the delivery of varied versus constant high-, medium-, and low-preference stimuli on performance of 2 adults on a computer-based task in an analogue employment setting...

  2. Biodegradation testing of chemicals with high Henry’s constants – separating mass and effective concentration reveals higher rate constants

    DEFF Research Database (Denmark)

    Birch, Heidi; Andersen, Henrik Rasmus; Comber, Mike

    During simulation-type biodegradation tests, volatile chemicals will continuously partition between water phase and headspace. This study addressed how (1) this partitioning affects biodegradation test results and (2) it can be accounted for by combining mass balance and dynamic biodegradation...... Microextraction (HS-SPME) was applied directly on the test systems to measure substrate depletion by biodegradation relative to abiotic controls. HS-SPME was also applied to determine air to water partitioning ratios. Water phase biodegradation rate constants, kwater, were up to 72 times higher than test system...... biodegradation rate constants, ksystem. True water phase degradation rate constants facilitate extrapolation to other air-water systems and are more suitable input parameters for aquatic exposure and fate models. As such, they should be considered more appropriate for risk assessments than test system rate...

  3. Flexible Acyclic Polyol-Chloride Anion Complexes and Their Characterization by Photoelectron Spectroscopy and Variable Temperature Binding Constant Determinations

    Energy Technology Data Exchange (ETDEWEB)

    Shokri, Alireza; Wang, Xue B.; Wang, Yangping; O' Doherty, George A.; Kass, Steven R.

    2016-03-17

    Flexible acyclic alcohols with 1–5 hydroxyl groups were bound to chloride anion and these complexes were interrogated by negative ion photoelectron spectroscopy and companion density functional theory computations. The resulting vertical detachment energies are reproduced on average to 0.10 eV by M06-2X/aug-cc-pVTZ predictions and range from 4.45 – 5.96 eV. These values are 0.84 – 2.35 eV larger than the adiabatic detachment energy of Cl– as a result of the larger hydrogen bond networks in the bigger polyols. Adiabatic detachment energies of the alcohol–Cl– clusters are more difficult to determine both experimentally and computationally. This is due to the large geometry changes that occur upon photodetachment and the large bond dissociation energy of H–Cl which enables the resulting chlorine atom to abstract a hydrogen from any of the methylene (CH2) or methine (CH) positions. Both ionic and non-ionic hydrogen bonds (i.e., OH•••Cl– and OH•••OH•••Cl–) form in the larger polyols complexes, and are found to be energetically comparable. Subtle structural differences, consequently can lead to the formation of different types of hydrogen bonds and maximizing the ionic ones is not always preferred. Solution equilibrium binding constants between the alcohols and tetrrabuylammonium chloride (TBACl) in acetonitrile at -24.2, 22.0, and 53.6 °C were also determined. The free energies of association are nearly identical for all of the substrates (i.e., ΔG° = -2.8 ± 0.7 kcal mol–1). Compensating enthalpy and entropy values reveal, contrary to expectation and the intrinsic gas-phase preferences, that the bigger systems with more hydroxyl groups are entropically favored and enthalpically disfavored relative to the smaller species. This suggests that more solvent molecules are released upon binding TBACl to alcohols with more hydroxyl groups and is consistent with the measured negative heat capacities. These quantities increase with

  4. High Affinity Binding of Indium and Ruthenium Ions by Gastrins.

    Directory of Open Access Journals (Sweden)

    Graham S Baldwin

    Full Text Available The peptide hormone gastrin binds two ferric ions with high affinity, and iron binding is essential for the biological activity of non-amidated forms of the hormone. Since gastrins act as growth factors in gastrointestinal cancers, and as peptides labelled with Ga and In isotopes are increasingly used for cancer diagnosis, the ability of gastrins to bind other metal ions was investigated systematically by absorption spectroscopy. The coordination structures of the complexes were characterized by extended X-ray absorption fine structure (EXAFS spectroscopy. Changes in the absorption of gastrin in the presence of increasing concentrations of Ga3+ were fitted by a 2 site model with dissociation constants (Kd of 3.3 x 10-7 and 1.1 x 10-6 M. Although the absorption of gastrin did not change upon the addition of In3+ ions, the changes in absorbance on Fe3+ ion binding in the presence of indium ions were fitted by a 2 site model with Kd values for In3+ of 6.5 x 10-15 and 1.7 x 10-7 M. Similar results were obtained with Ru3+ ions, although the Kd values for Ru3+ of 2.6 x 10-13 and 1.2 x 10-5 M were slightly larger than observed for In3+. The structures determined by EXAFS all had metal:gastrin stoichiometries of 2:1 but, while the metal ions in the Fe, Ga and In complexes were bridged by a carboxylate and an oxygen with a metal-metal separation of 3.0-3.3 Å, the Ru complex clearly demonstrated a short range Ru-Ru separation, which was significantly shorter, at 2.4 Å, indicative of a metal-metal bond. We conclude that gastrin selectively binds two In3+ or Ru3+ ions, and that the affinity of the first site for In3+ or Ru3+ ions is higher than for ferric ions. Some of the metal ion-gastrin complexes may be useful for cancer diagnosis and therapy.

  5. Biodegradation testing of chemicals with high Henry's constants - Separating mass and effective concentration reveals higher rate constants.

    Science.gov (United States)

    Birch, Heidi; Andersen, Henrik R; Comber, Mike; Mayer, Philipp

    2017-05-01

    During simulation-type biodegradation tests, volatile chemicals will continuously partition between water phase and headspace. This study addressed how (1) this partitioning affects test results and (2) can be accounted for by combining equilibrium partition and dynamic biodegradation models. An aqueous mixture of 9 (semi)volatile chemicals was first generated using passive dosing and then diluted with environmental surface water producing concentrations in the ng/L to μg/L range. After incubation for 2 h to 4 weeks, automated Headspace Solid Phase Microextraction (HS-SPME) was applied directly on the test systems to measure substrate depletion by biodegradation relatively to abiotic controls. HS-SPME was also applied to determine air to water partitioning ratios. Biodegradation rate constants relating to the chemical in the water phase, kwater, were generally a factor 1 to 11 times higher than biodegradation rate constants relating to the total mass of chemical in the test system, ksystem, with one exceptional factor of 72 times for a long chain alkane. True water phase degradation rate constants were found (i) more appropriate for risk assessment than test system rate constants, (ii) to facilitate extrapolation to other air-water systems and (iii) to be better defined input parameters for aquatic exposure and fate models.

  6. Isothermal titration calorimetry for drug design: Precision of the enthalpy and binding constant measurements and comparison of the instruments.

    Science.gov (United States)

    Linkuvienė, Vaida; Krainer, Georg; Chen, Wen-Yih; Matulis, Daumantas

    2016-12-15

    Isothermal titration calorimetry (ITC) is one of the most robust label- and immobilization-free techniques used to measure protein - small molecule interactions in drug design for the simultaneous determination of the binding affinity (ΔG) and the enthalpy (ΔH), both of which are important parameters for structure-thermodynamics correlations. It is important to evaluate the precision of the method and of various ITC instrument models by performing a single well-characterized reaction. The binding between carbonic anhydrase II and acetazolamide was measured by four ITC instruments - PEAQ-ITC, iTC200, VP-ITC, and MCS-ITC and the standard deviation of ΔG and ΔH was determined. Furthermore, the limit of an approach to reduce the protein concentration was studied for a high-affinity reaction (Kd = 0.3 nM), too tight to be measured by direct (non-displacement) ITC. Chemical validation of the enthalpy measurements is discussed. Copyright © 2016 Elsevier Inc. All rights reserved.

  7. High dielectric constant materials and their application to IC gate stack systems

    Institute of Scientific and Technical Information of China (English)

    TU; Hailing

    2005-01-01

    High dielectric constant (high-k) materials are vital tothe nanoelectronic devices.The paper reviews research development of high-k materials, describes a variety of manufacture technologies and discusses the application of the gate stack systems to non-classical device structures.

  8. Charge regulation of weak polyelectrolytes at low- and high-dielectric-constant substrates

    CERN Document Server

    Netz, R R

    2003-01-01

    As is well known, the effective charge of weak polyelectrolytes (PEs) decreases with decreasing salt concentration due to the electrostatic repulsion between dissociated charges. Close to dielectric boundaries, image-charge effects influence the dissociation equilibrium. At low-dielectric-constant substrates, one finds a further charge decrease and repulsion from the interface, while at high-dielectric-constant (e.g. metallic) substrates, the effective charge increases and the PE is attracted to the interface.

  9. Determination of the kinetic rate constant of cyclodextrin supramolecular systems by high performance affinity chromatography.

    Science.gov (United States)

    Li, Haiyan; Ge, Jingwen; Guo, Tao; Yang, Shuo; He, Zhonggui; York, Peter; Sun, Lixin; Xu, Xu; Zhang, Jiwen

    2013-08-30

    It is challenging and extremely difficult to measure the kinetics of supramolecular systems with extensive, weak binding (Kahigh performance affinity chromatography (HPAC) was established to determine the dissociation rate constant of cyclodextrin supramolecular systems. The interactions of β-cyclodextrin with acetaminophen and sertraline were used to exemplify the method. The retention times, variances and the plate heights of the peaks for acetaminophen or sertraline, conventional non-retained substance (H2O) on the β-cyclodextrin bonded column and a control column were determined at four flow rates under linear elution conditions. Then, plate heights for the theoretical non-retained substance were estimated by the modified HPAC method, in consideration of the diffusion and stagnant mobile phase mass transfer. As a result, apparent dissociation rate constants of 1.82 (±0.01)s(-1) and 3.55 (±0.37)s(-1) were estimated for acetaminophen and sertraline respectively at pH 6.8 and 25°C with multiple flow rates. Following subtraction of the non-specific binding with the support, dissociation rate constants were estimated as 1.78 (±0.00) and 1.91 (±0.02)s(-1) for acetaminophen and sertraline, respectively. These results for acetaminophen and sertraline were in good agreement with the magnitude of the rate constants for other drugs determined by capillary electrophoresis reported in the literature and the peak fitting method we performed. The method described in this work is thought to be suitable for other supramolecules, with relatively weak, fast and extensive interactions.

  10. Molecular statics calculations of proton binding to goethite surfaces: A new approach to estimation of stability constants for multisite surface complexation models

    Science.gov (United States)

    Rustad, James R.; Felmy, Andrew R.; Hay, Benjamin P.

    1996-05-01

    A new approach to estimating stability constants for proton binding in multisite surface complexation models is presented. The method is based on molecular statics computation of energies for the formation of proton vacancies and interstitials in ideal periodic slabs representing the (100), (110), (010), (001), and (021) surfaces of goethite. Gas-phase energies of clusters representing the hydrolysis products of ferric iron are calculated using the same potential energy functions used for the surface. These energies are linearly related to the hydrolysis constants for ferric iron in aqueous solution. Stability constants for proton binding at goethite surfaces are estimated by assuming the same log K- Δ E relationship for goethite surface protonation reactions. These stability constants predict a pH of zero charge of 8.9, in adequate agreement with measurements on CO 2-free goethite. The estimated stability constants differ significantly from previous estimations based on Pauling bond strength. We find that nearly all the surface oxide ions are reactive; nineteen of the twenty-six surface sites investigated have log Kint between 7.7 and 9.4. This implies a site density between fifteen and sixteen reactive sites/nm for crystals dominated by (110) and (021) crystal faces.

  11. A model of mitochondrial creatine kinase binding to membranes: adsorption constants, essential amino acids and the effect of ionic strength

    DEFF Research Database (Denmark)

    Fedosov, Sergey; Belousova, Lubov; Plesner, Igor

    1993-01-01

    The quantitative aspects of mitochondrial creatinekinase (mitCK) binding to mitochondrial membranes were investigated. A simple adsorption and binding model was used for data fitting, taking into account the influence of protein concentration, pH, ionic strength and substrate concentration on the...

  12. Presence of a highly efficient binding to bacterial contamination can distort data from binding studies

    Energy Technology Data Exchange (ETDEWEB)

    Balcar, V.J. (Department of Anatomy, University of Sydney, N.S.W. (Australia))

    1990-12-01

    {sup 3}HGABA at low concentrations (5-10 nM) was bound by what appeared to be a GABA receptor binding site in bacterial contamination originating from a batch of distilled water. Under experimental conditions similar to those usually employed in {sup 3}HGABA binding studies, the apparent binding displayed a very high specific component and a high efficiency in terms of {sup 3}HGABA bound per mg of protein. The binding was blocked by muscimol but not by isoguvacine, SR95531 and nipecotic acid. These characteristics suggest that the presence of such spurious binding in the experiments using 3H-labeled ligands in brain homogenates may not always be very obvious and, moreover, it can result in subtle, but serious, distortions of data from such studies, which may not be immediately recognized.

  13. Binding constants of inclusion complexes of nitroimidazoles with {beta}-cyclodextrins in the absence and presence of PVP

    Energy Technology Data Exchange (ETDEWEB)

    Chadha, Renu [University Institute of Pharmaceutical Sciences, Panjab University, Chandigarh 160014 (India)]. E-mail: renukchadha2004@yahoo.co.in; Jain, D.V.S. [Depatment of Chemistry, Panjab University, Chandigarh 160014 (India); Aggarwal, Amit [University Institute of Pharmaceutical Sciences, Panjab University, Chandigarh 160014 (India); Singh, Surjit [GND University, Amritsar, Panjab (India); Thakur, Deepika [University Institute of Pharmaceutical Sciences, Panjab University, Chandigarh 160014 (India)

    2007-07-01

    Thermodynamics of complexation of 5-nitroimidazoles with {beta}-cyclodextrin and its methylated and hydroxypropyl derivatives in water and in 0.25% polyvinylpyrrolidone are determined by solution calorimetry. A 1:1 stoichiometry was established. The equilibrium constant (K) for all the nitroimidazoles fall in the range 1000-1900 M{sup -1} suitable for use of cyclodextrins as drug carriers. The complexation ability is significantly enhanced by methylation of the {beta}-cyclodextrin. The stability constant increased in the order metronidazole < ornidazole < tinidazole < secnidazole. The presence of polyvinylpyrrolidone enhances the stability constants.

  14. Specific high-affinity binding of fatty acids to epidermal cytosolic proteins

    Energy Technology Data Exchange (ETDEWEB)

    Raza, H.; Chung, W.L.; Mukhtar, H. (Department of Dermatology, University Hospitals of Cleveland, Case Western Reserve University, OH (USA))

    1991-08-01

    Cytosol from rat, mouse, and human skin or rat epidermis was incubated with (3H)arachidonic acid, (14C)retinoic acid, (14C)oleic acid, (3H)leukotriene A4, (3H)prostaglandin E2 (PGE2) or (3H) 15-hydroxyeicosatetraenoic acid (15-HETE), and protein-bound ligands were separated using Lipidex-1000 at 4C to assess the binding specificity. The binding of oleic acid and arachidonic acid with rat epidermal cytosol was rapid, saturable, and reversible. Binding of oleic acid was competed out with the simultaneous addition of other ligands and found to be in the following order: arachidonic acid greater than oleic acid greater than linoleic acid greater than lauric acid greater than leukotriene A4 greater than 15-HETE = PGE1 greater than PGE2 = PGF2. Scatchard analysis of the binding with arachidonic acid, oleic acid, and retinoic acid revealed high-affinity binding sites with the dissociation constant in the nM range. SDS-PAGE analysis of the oleic acid-bound epidermal cytosolic protein(s) revealed maximum binding at the 14.5 kDa region. The presence of the fatty acid-binding protein in epidermal cytosol and its binding to fatty acids and retinoic acid may be of significance both in the trafficking and the metabolism of fatty acids and retinoids across the skin.

  15. Binding of hairpin pyrrole and imidazole polyamides to DNA: relationship between torsion angle and association rate constants.

    Science.gov (United States)

    Han, Yong-Woon; Matsumoto, Tomoko; Yokota, Hiroaki; Kashiwazaki, Gengo; Morinaga, Hironobu; Hashiya, Kaori; Bando, Toshikazu; Harada, Yoshie; Sugiyama, Hiroshi

    2012-12-01

    N-methylpyrrole (Py)-N-methylimidazole (Im) polyamides are small organic molecules that bind to DNA with sequence specificity and can be used as synthetic DNA-binding ligands. In this study, five hairpin eight-ring Py-Im polyamides 1-5 with different number of Im rings were synthesized, and their binding behaviour was investigated with surface plasmon resonance assay. It was found that association rate (k(a)) of the Py-Im polyamides with their target DNA decreased with the number of Im in the Py-Im polyamides. The structures of four-ring Py-Im polyamides derived from density functional theory revealed that the dihedral angle of the Py amide carbonyl is 14∼18°, whereas that of the Im is significantly smaller. As the minor groove of DNA has a helical structure, planar Py-Im polyamides need to change their conformation to fit it upon binding to the minor groove. The data explain that an increase in planarity of Py-Im polyamide induced by the incorporation of Im reduces the association rate of Py-Im polyamides. This fundamental knowledge of the binding of Py-Im polyamides to DNA will facilitate the design of hairpin Py-Im polyamides as synthetic DNA-binding modules.

  16. A comparative study of capillary electrophoresis and isothermal titration calorimetry for the determination of binding constant of human serum albumin to monoclonal antibody.

    Science.gov (United States)

    Andrási, Melinda; Lehoczki, Gábor; Nagy, Zoltán; Gyémánt, Gyöngyi; Pungor, András; Gáspár, Attila

    2015-06-01

    This paper focuses on the investigation of the interactions between the anti-HSA-mAb and its protein antigen using CZE, ACE, and isothermal titration calorimetry. The CZE revealed the formation of the anti-HSA-mAb·HSA and anti-HSA-mAb·(HSA)2 complexes and the binding constants determined by plotting the amount of the bound anti-HSA-mAb as a function of the concentration of HSA. The ACE provided information on the binding strength from the change in effective electrophoretic mobility of the anti-HSA-mAb. These two separation techniques estimated the presence of two binding sites. The equilibrium dissociation constant values obtained by CZE and ACE were found to be 2.26 × 10(-6) M for anti-HSA-mAb·HSA, 1.22 × 10(-6) M for anti-HSA-mAb·(HSA)2 and 4.45 × 10(-8) M for anti-HSA-mAb·HSA, 1.08 × 10(-7) M for anti-HSA-mAb·(HSA)2 , respectively. The dissociation constant data obtained by ACE were in congruence with the values obtained by isothermal titration calorimetry (2.74 × 10(-8) M, 1.04 × 10(-7) M). © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  17. Determination of the binding constant between alprostadil and alpha-cyclodextrin by capillary electrophoresis: implications for a freeze-dried formulation.

    Science.gov (United States)

    Schipper, Benjamin R; Ramstad, Tore

    2005-07-01

    The binding constant between alprostadil (PGE1) and alpha-cyclodextrin (alpha-CD) was determined at three temperatures by capillary electrophoresis. alpha-CD is an excipient material in Caverject Dual Chamber Syringe (DCS), added to enhance stability. The binding constant was used to calculate the amount of PGE1 free upon reconstitution and injection, the latter of which is critical to product performance. Measurement was made in a pH 7.2 separation buffer to ensure a negative charge on PGE1. The concentration of PGE1 was fixed while the concentration of alpha-CD was varied over a suitable range. As the amount of PGE1 bound to alpha-CD increases, the weighted average of the resultant mobility decreases, thereby allowing a binding isotherm to be generated from which the stability constant was extracted via nonlinear regression analysis. A value of 708 +/- 64 M(-1) was obtained at 27 degrees C, while at physiological temperature (37 degrees C) the value was 537 +/- 27 M(-1). These results compare favorably to values previously obtained by NMR. Calculation of the percent PGE1 free upon reconstitution and injection show it to be near the desired outcome of 100%. Hence, we were able to conclude that the amount of free drug delivered by Caverject DCS is nominally the same as for Caverject S. Po., an earlier-developed product that contains no alpha-CD.

  18. Determination by NMR of the binding constant for the molecular complex between alprostadil and alpha-cyclodextrin. Implications for a freeze-dried formulation.

    Science.gov (United States)

    Ramstad, Tore; Hadden, Chad E; Martin, Gary E; Speaker, Stanley M; Teagarden, Dirk L; Thamann, Thomas J

    2005-05-30

    A binding constant was determined for the complexation reaction between alprostadil (PGE1) and alpha-cyclodextrin (alpha-CD). This constant was used to calculate the fraction PGE1 free upon reconstitution of Caverject dual chamber syringe, indicated for the treatment of erectile dysfunction. The determination was based on the measurement of the chemical shift of the C20 methyl protons of PGE1. The observed chemical shift varies as a linear function of the amount of PGE1 bound. The binding constant was obtained from the binding isotherm, a curve of the observed chemical shift versus free ligand (alpha-CD) concentration, through the application of non-linear regression analysis. A value K11 = 966 M(-1) +/- 130 M(-1) (2s), measured at 27 degrees C, was obtained. This value is in good agreement with those reported in the literature. The percent PGE1 free was subsequently calculated for the reconstituted solution and in the corpora cavernosum after injection. The latter showed PGE1 to be delivered essentially quantitatively to the targeted site.

  19. High Precision Calculations of the Lennard-Jones Lattice Constants for Five Lattices

    Science.gov (United States)

    Stein, Matthew

    2017-01-01

    The total potential energy of a crystal as described by the Lennard-Jones (L-J) potential depends in part upon the calculation of lattice constants. Knowing these constants to high precision is useful for prediction of the lattice type and simulation of crystals such as rare-gas solids or germanium detectors, but reaching higher precision is computationally costly and challenging. Presented here is the extension of the precision of the lattice constants, Lp, up to 32 decimal digits, and in some cases corrections from previous publication. The Lp terms are given for 4 cubic, face-centered cubic, body-centered cubic, hexagonal-close-pack, and diamond lattices. This precision was obtained through the use of careful parallelization technique, exploitation of the symmetries of each lattice, and the ``onionization'' of the simulated crystal. The results of this computation, along with the tools and algorithm strategies to make this computation possible, are explained in detail graphically.

  20. Applications of a tight-binding total energy method for transition and noble metals Elastic Constants, Vacancies, and Surfaces of Monatomic Metals

    CERN Document Server

    Mehl, M J; Mehl, Michael J.; Papaconstantopoulos, Dimitrios A.

    1996-01-01

    A recent tight-binding scheme provides a method for extending the results of first principles calculations to regimes involving $10^2 - 10^3$ atoms in a unit cell. The method uses an analytic set of two-center, non-orthogonal tight-binding parameters, on-site terms which change with the local environment, and no pair potential. The free parameters in this method are chosen to simultaneously fit band structures and total energies from a set of first-principles calculations for monatomic fcc and bcc crystals. To check the accuracy of this method we evaluate structural energy differences, elastic constants, vacancy formation energies, and surface energies, comparing to first-principles calculations and experiment. In most cases there is good agreement between this theory and experiment. We present a detailed account of the method, a complete set of tight-binding parameters, and results for twenty-nine of the alkaline earth, transition and noble metals.

  1. High resolution genome wide binding event finding and motif discovery reveals transcription factor spatial binding constraints.

    Directory of Open Access Journals (Sweden)

    Yuchun Guo

    Full Text Available An essential component of genome function is the syntax of genomic regulatory elements that determine how diverse transcription factors interact to orchestrate a program of regulatory control. A precise characterization of in vivo spacing constraints between key transcription factors would reveal key aspects of this genomic regulatory language. To discover novel transcription factor spatial binding constraints in vivo, we developed a new integrative computational method, genome wide event finding and motif discovery (GEM. GEM resolves ChIP data into explanatory motifs and binding events at high spatial resolution by linking binding event discovery and motif discovery with positional priors in the context of a generative probabilistic model of ChIP data and genome sequence. GEM analysis of 63 transcription factors in 214 ENCODE human ChIP-Seq experiments recovers more known factor motifs than other contemporary methods, and discovers six new motifs for factors with unknown binding specificity. GEM's adaptive learning of binding-event read distributions allows it to further improve upon previous methods for processing ChIP-Seq and ChIP-exo data to yield unsurpassed spatial resolution and discovery of closely spaced binding events of the same factor. In a systematic analysis of in vivo sequence-specific transcription factor binding using GEM, we have found hundreds of spatial binding constraints between factors. GEM found 37 examples of factor binding constraints in mouse ES cells, including strong distance-specific constraints between Klf4 and other key regulatory factors. In human ENCODE data, GEM found 390 examples of spatially constrained pair-wise binding, including such novel pairs as c-Fos:c-Jun/USF1, CTCF/Egr1, and HNF4A/FOXA1. The discovery of new factor-factor spatial constraints in ChIP data is significant because it proposes testable models for regulatory factor interactions that will help elucidate genome function and the

  2. High resolution genome wide binding event finding and motif discovery reveals transcription factor spatial binding constraints.

    Science.gov (United States)

    Guo, Yuchun; Mahony, Shaun; Gifford, David K

    2012-01-01

    An essential component of genome function is the syntax of genomic regulatory elements that determine how diverse transcription factors interact to orchestrate a program of regulatory control. A precise characterization of in vivo spacing constraints between key transcription factors would reveal key aspects of this genomic regulatory language. To discover novel transcription factor spatial binding constraints in vivo, we developed a new integrative computational method, genome wide event finding and motif discovery (GEM). GEM resolves ChIP data into explanatory motifs and binding events at high spatial resolution by linking binding event discovery and motif discovery with positional priors in the context of a generative probabilistic model of ChIP data and genome sequence. GEM analysis of 63 transcription factors in 214 ENCODE human ChIP-Seq experiments recovers more known factor motifs than other contemporary methods, and discovers six new motifs for factors with unknown binding specificity. GEM's adaptive learning of binding-event read distributions allows it to further improve upon previous methods for processing ChIP-Seq and ChIP-exo data to yield unsurpassed spatial resolution and discovery of closely spaced binding events of the same factor. In a systematic analysis of in vivo sequence-specific transcription factor binding using GEM, we have found hundreds of spatial binding constraints between factors. GEM found 37 examples of factor binding constraints in mouse ES cells, including strong distance-specific constraints between Klf4 and other key regulatory factors. In human ENCODE data, GEM found 390 examples of spatially constrained pair-wise binding, including such novel pairs as c-Fos:c-Jun/USF1, CTCF/Egr1, and HNF4A/FOXA1. The discovery of new factor-factor spatial constraints in ChIP data is significant because it proposes testable models for regulatory factor interactions that will help elucidate genome function and the implementation of combinatorial

  3. Theoretical calculation of the NMR spin-spin coupling constants and the NMR shifts allow distinguishability between the specific direct and the water-mediated binding of a divalent metal cation to guanine.

    Science.gov (United States)

    Sychrovský, Vladimír; Sponer, Jirí; Hobza, Pavel

    2004-01-21

    The calculated intermolecular and intramolecular indirect NMR spin-spin coupling constants and NMR shifts were used for the discrimination between the inner-shell and the outer-shell binding motif of hydrated divalent cations Mg(2+) or Zn(2+) with a guanine base. The intermolecular coupling constants (1)J(X,O6) and (1)J(X,N7) (X = Mg(2+), Zn(2+)) can be unambiguously assigned to the specific inner-shell binding motif of the hydrated cation either with oxygen O6 or with nitrogen N7 of guanine. The calculated coupling constants (1)J(Mg,O6) and (1)J(Zn,O6) were 6.2 and -17.5 Hz, respectively, for the inner-shell complex of cation directly interacting with oxygen O6 of guanine. For the inner-shell coordination of the cation at nitrogen N7, the calculated coupling constants (1)J(Mg,N7) and (1)J(Zn,N7) were 5.6 and -36.5 Hz, respectively. When the binding of the cation is water-mediated, the coupling constant is zero. To obtain reliable shifts in NMR parameters, hydrated guanine was utilized as the reference state. The calculated change of NMR spin-spin coupling constants due to the hydration and coordination of the cation with guanine is caused mainly by the variation of Fermi-contact coupling contribution while the variation of diamagnetic spin-orbit, paramagnetic spin-orbit, and spin-dipolar coupling contributions is small. The change of s-character of guanine sigma bonding, sigma antibonding, and lone pair orbitals upon the hydration and cation coordination (calculated using the Natural Bond Orbital analysis) correlates with the variation of the Fermi-contact term. The calculated NMR shifts delta(N7) of -15.3 and -12.2 ppm upon the coordination of Mg(2+) and Zn(2+) ion are similar to the NMR shift of 19.6 ppm toward the high field measured by Tanaka for N7 of guanine upon the coordination of the Cd(2+) cation (Tanaka, Y.; Kojima, C.; Morita, E. H.; Kasai. Y.; Yamasaki, K.; Ono, A.; Kainosho, M.; Taira, K. J. Am. Chem. Soc. 2002, 124, 4595-4601). The present data

  4. Using Electrophoretic Mobility Shift Assays to Measure Equilibrium Dissociation Constants: GAL4-p53 Binding DNA as a Model System

    Science.gov (United States)

    Heffler, Michael A.; Walters, Ryan D.; Kugel, Jennifer F.

    2012-01-01

    An undergraduate biochemistry laboratory experiment is described that will teach students the practical and theoretical considerations for measuring the equilibrium dissociation constant (K[subscript D]) for a protein/DNA interaction using electrophoretic mobility shift assays (EMSAs). An EMSA monitors the migration of DNA through a native gel;…

  5. Highly selective ligand binding by Methylophilus methylotrophus cytochrome c''.

    Science.gov (United States)

    Quintas, Pedro O; Catarino, Teresa; Todorovic, Smilja; Turner, David L

    2011-06-28

    Cytochrome c'' (cyt c'') from Methylophilus methylotrophus is unusual insofar as the heme has two axial histidine ligands in the oxidized form but one is detached when the protein is reduced. Despite cyt c'' having an axial site available for binding small ligands, we show here that only NO binds readily to the ferrous cyt c''. Binding of CO, as well as CN(-), on the other hand requires considerable structural reorganization, or reduction of the disulfide bridge close to the heme. Standard free energies for the binding of NO and CO reveal high selectivity of the ferrous cyt c'' for NO, indicating its putative physiological role. In this work, we characterize in detail the kinetics of NO binding and the structural features of the Fe(2+)-NO adduct by stopped-flow and resonance Raman spectroscopy, respectively.

  6. Raman spectroscopic determination of the molecular constants of the hydrogen isotopologues with high accuracy

    Energy Technology Data Exchange (ETDEWEB)

    Krasch, Bennet; Mirz, Sebastian; Groessle, Robin [Karlsruhe Institute of Technology KIT (Germany). Institute for Technical Physics (ITEP), Tritium Laboratory Karlsruhe (TLK); Collaboration: KATRIN-Collaboration

    2016-07-01

    The interest in the thermodynamic properties of gases as the chemical equilibrium is faced by the challenge of time-consuming and technical extensive experimental setups. One possible solution is the derivation of these properties from the molecular constants. The rotational and vibrational movement of diatomic molecules, as the hydrogen isotopologues, is described by the concept of the rotational anharmonic oscillator. The molecular constants are the free parameters of this concept. Molecular constants themselves can be determined by measuring the line position of rotational and/or rotational transitions e.g. with Raman spectroscopy for hydrogen as it has been done since several years. In this contribution a Raman method was development to measure the molecular constant of the hydrogen isotopologues with high accuracy to obtain reliable results. But not only the method was development but also a complete measurement uncertainty budget was set up. The uncertainty budget contains all possible sources for uncertainties from the measurement period or the analysis process as well the contribution of each single uncertainty. The method and the uncertainty budget were exemplary tested on Deuterium.

  7. Enhanced laboratory sensitivity to variation of the fine-structure constant using highly charged ions.

    Science.gov (United States)

    Berengut, J C; Dzuba, V A; Flambaum, V V

    2010-09-17

    We study atomic systems that are in the frequency range of optical atomic clocks and have enhanced sensitivity to potential time variation of the fine-structure constant α. The high sensitivity is due to coherent contributions from three factors: high nuclear charge Z, high ionization degree, and significant differences in the configuration composition of the states involved. Configuration crossing keeps the frequencies in the optical range despite the large ionization energies. We discuss a few promising examples that have the largest α sensitivities seen in atomic systems.

  8. Shock Tube Measurement of the High-Temperature Rate Constant for OH + CH3 → Products.

    Science.gov (United States)

    Wang, Shengkai; Li, Sijie; Davidson, David F; Hanson, Ronald K

    2015-08-20

    The reaction between hydroxyl (OH) and methyl radicals (CH3) is critical to hydrocarbon oxidation. Motivated by the sparseness of its high-temperature rate constant data and the large uncertainties in the existing literature values, the current study has remeasured the overall rate constant of the OH + CH3 reaction and extended the measurement temperature range to 1214-1933 K, using simultaneous laser absorption diagnostics for OH and CH3 radicals behind incident and reflected shock waves. tert-Butyl hydroperoxide and azomethane were used as pyrolytic sources for the OH and CH3 radicals, respectively. The current study bridged the temperature ranges of existing experimental data, and good agreement is seen between the current measurement and some previous experimental and theoretical high-temperature studies. A recommendation for the rate constant expression of the title reaction, based on the weighted average of the high-temperature data from selected studies, is given by k1 = 4.19 × 10(1)(T/K)(3.15) exp(5270 K/T) cm(3) mol(-1) s(-1) ±30%, which is valid over 1000-2500 K.

  9. Controllable reduction of graphene oxide and its application during the fabrication of high dielectric constant composites

    Science.gov (United States)

    Liu, Hui; Xu, Peng; Yao, Haibo; Chen, Wenhui; Zhao, Jianying; Kang, Chuanqing; Bian, Zheng; Gao, Lianxun; Guo, Haiquan

    2017-10-01

    The synthesis of reduced graphene oxide (RGO) with various reduction extents was carried out in organic solvent using 1,4-diiodobutane as the reducing agent at moderate temperatures. Results showed that the C/O ratio of RGO nanosheet surface could be tailored by adjusting the ratio of graphene oxide (GO) and reducing agent. The controllable reduction strategy was applied to the fabrication of high dielectric constant graphene/polyimide composites via the in situ reduction of GO. The reduction extents of RGO in polymer matrix can be readily manipulated just through altering the addition of the reducing agent. The dielectric constants of gaphene/polyimide composites were significantly enhanced with the increasing of the reduction extent of RGO. Moreover, the mechanical properties of the composites were also affected by the reduction extent of RGO due to the decreases of the oxygen functional groups of RGO surface. Hence, the in situ controllable reduction of GO should be quite an ideal method for the fabrication of high dielectric constant composites with the tunable combination properties.

  10. An investigation of constant-pressure gas well testing influenced by high-velocity flow

    Energy Technology Data Exchange (ETDEWEB)

    Berumen, Sergio; Samaniego, Fernando; Cinco, Heber [PEMEX E and P and UNAM, Ciudad Universitaria, Coyoacan, Mexico (Mexico)

    1997-11-05

    This paper presents the results of a study of transient pressure analysis of gas flow under either constant bottom-hole pressure conditions or the constant wellhead pressure conditions. The effects of formation damage, wellbore storage and high-velocity flow are included in the model. The analysis of simulated well tests showed that the interpretation methods used for liquid flow are generally accurate when the m(p) is used. For these conditions, a graph of 1/q{sub D} vs. logt{sub D} presents gradually lower values of 1.1513 as the value of p{sub wf} decreases: for pressure buildup conditions, a graph of m{sub D}(1, {Delta}t{sub a{sub D}})/q{sub D}({Delta}t{sub a{sub D}}=0) vs. (t{sub a{sub D}}+{Delta}t{sub a{sub D}})/{Delta}t{sub a{sub D}} shows values of this slope within 1% of the 1.1513 value. However, when high-velocity flow influences constant pressure production tests, the slope can yield errors up to 13%. This upper limit occurs when the formation has a relatively `high` permeability (around 1 mD) and the rate performance test is affected by high-velocity flow. It was found that pressure buildup tests are superior to rate performance tests because high-velocity flow does not affect the slope of the straight line portion of the buildup curve. Derivative analysis of simulated buildup tests showed that the skin factor is considerably miscalculated when the high-velocity flow effect is significant. This problem could lead to errors in the calculation of the skin factor, s, up to 300%

  11. High Throughput Constant Envelope Pre-coder for Massive MIMO Systems

    OpenAIRE

    Prabhu, Hemanth; Rusek, Fredrik; Rodrigues, Joachim; Edfors, Ove

    2015-01-01

    This study describes a high throughput constant envelope (CE) pre-coder for Massive MIMO systems. A large number of antennas (M), in the order of 100s, serve a relatively small number of users (K) simultaneously. The stringent amplitude constraint (only phase changes) in the CE scheme is motivated by the use of highly power-efficient non-linear RF power amplifiers. We propose a scheme that computes the CE signals to be transmitted based on box-constrained regression (coordinatedescent),with a...

  12. Mannosylerythritol lipid, a yeast extracellular glycolipid, shows high binding affinity towards human immunoglobulin G

    Directory of Open Access Journals (Sweden)

    Ikegami Toru

    2001-09-01

    Full Text Available Abstract Background There have been many attempts to develop new materials with stability and high affinity towards immunoglobulins. Some of glycolipids such as gangliosides exhibit a high affinity toward immunoglobulins. However, it is considerably difficult to develop these glycolipids into the practical separation ligand due to their limited amounts. We thus focused our attention on the feasible use of "mannosylerythritol lipid A", a yeast glycolipid biosurfactant, as an alternative ligand for immunoglobulins, and undertook the investigation on the binding between mannosylerythritol lipid A (MEL-A and human immunoglobulin G (HIgG. Results In ELISA assay, MEL-A showed nearly the same binding affinity towards HIgG as that of bovine ganglioside GM1. Fab of human IgG was considered to play a more important role than Fc in the binding of HIgG by MEL-A. The bound amount of HIgG increased depending on the attached amount of MEL-A onto poly (2-hydroxyethyl methacrylate (polyHEMA beads, whereas the amount of human serum albumin slightly decreased. Binding-amount and -selectivity of HIgG towards MEL-A were influenced by salt species, salt concentration and pH in the buffer solution. The composite of MEL-A and polyHEMA, exhibited a significant binding constant of 1.43 × 106 (M-1 for HIgG, which is approximately 4-fold greater than that of protein A reported. Conclusions MEL-A shows high binding-affinity towards HIgG, and this is considered to be due to "multivalent effect" based on the binding molar ratio. This is the first report on the binding of a natural human antibody towards a yeast glycolipid.

  13. HypCal, a general-purpose computer program for the determination of standard reaction enthalpy and binding constant values by means of calorimetry.

    Science.gov (United States)

    Arena, Giuseppe; Gans, Peter; Sgarlata, Carmelo

    2016-09-01

    The program HypCal has been developed to provide a means for the simultaneous determination, from data obtained by isothermal titration calorimetry, of both standard enthalpy of reaction and binding constant values. The chemical system is defined in terms of species of given stoichiometry rather than in terms of binding models (e.g., independent or cooperative). The program does not impose any limits on the complexity of the chemical systems that can be treated, including competing ligand systems. Many titration curves may be treated simultaneously. HypCal can also be used as a simulation program when designing experiments. The use of the program is illustrated with data obtained with nicotinic acid (niacin, pyridine-3 carboxylic acid). Preliminary experiments were used to establish the rather different titration conditions for the two sets of titration curves that are needed to determine the parameters for protonation of the carboxylate and amine groups.

  14. Human immunodeficiency virus drug development assisted with AlGaN/GaN high electron mobility transistors and binding-site models

    Science.gov (United States)

    Kang, Yen-Wen; Lee, Geng-Yen; Chyi, Jen-Inn; Hsu, Chen-Pin; Hsu, You-Ren; Hsu, Chia-Hsien; Huang, Yu-Fen; Sun, Yuh-Chang; Chen, Chih-Chen; Chun Hung, Sheng; Ren, Fan; Andrew Yeh, J.; Wang, Yu-Lin

    2013-04-01

    Human immunodeficiency virus (HIV) Reverse Transcriptase (RT)-immobilized AlGaN/GaN high electron mobility transistors (HEMTs) and binding-site models were used to find out the dissociation constants of the HIV RT-inhibitor complex and the number of the binding sites on RT for the inhibitor, Efavirenz. One binding site on the RT for the inhibitor is predicted and the dissociation constant extracted from the binding-site model is 0.212 nM. The AlGaN/GaN HEMTs and the binding-site-models are demonstrated to be good tools to assist drug developments by elucidating the dissociation constants and the number of binding sites, which can largely reduce the cost and time for drug developments.

  15. Atomic spectroscopy and highly accurate measurement: determination of fundamental constants; Spectroscopie atomique et mesures de grande precision: determination de constantes fonfamentales

    Energy Technology Data Exchange (ETDEWEB)

    Schwob, C

    2006-12-15

    This document reviews the theoretical and experimental achievements of the author concerning highly accurate atomic spectroscopy applied for the determination of fundamental constants. A pure optical frequency measurement of the 2S-12D 2-photon transitions in atomic hydrogen and deuterium has been performed. The experimental setting-up is described as well as the data analysis. Optimized values for the Rydberg constant and Lamb shifts have been deduced (R = 109737.31568516 (84) cm{sup -1}). An experiment devoted to the determination of the fine structure constant with an aimed relative uncertainty of 10{sup -9} began in 1999. This experiment is based on the fact that Bloch oscillations in a frequency chirped optical lattice are a powerful tool to transfer coherently many photon momenta to the atoms. We have used this method to measure accurately the ratio h/m(Rb). The measured value of the fine structure constant is {alpha}{sub -1} = 137.03599884 (91) with a relative uncertainty of 6.7*10{sup -9}. The future and perspectives of this experiment are presented. This document presented before an academic board will allow his author to manage research work and particularly to tutor thesis students. (A.C.)

  16. Mangiferin binding to serum albumin is non-saturable and induces conformational changes at high concentrations

    Energy Technology Data Exchange (ETDEWEB)

    Freitas, P.G.; Barbosa, A.F. [Biochemistry Laboratory, Institute of Exact Sciences, Federal University of Alfenas, Unifal-MG, R. Gabriel Monteiro da Silva, 700, 37130-000 Alfenas, MG (Brazil); Saraiva, L.A. [Phytochemistry and Medicinal Chemistry Laboratory, Institute of Exact Sciences, Unifal-MG (Brazil); Camps, I. [Physics Laboratory, Institute of Exact Sciences, Unifal-MG (Brazil); Silveira, N.J.F. da [Bioinformatics Laboratory, Institute of Exact Sciences, Unifal-MG (Brazil); Veloso, M.P. [Phytochemistry and Medicinal Chemistry Laboratory, Institute of Exact Sciences, Unifal-MG (Brazil); Santos, M.H., E-mail: poliany.santos@gmail.com [Phytochemistry and Medicinal Chemistry Laboratory, Institute of Exact Sciences, Unifal-MG (Brazil); Schneedorf, J.M., E-mail: zemasfs@gmail.com [Biochemistry Laboratory, Institute of Exact Sciences, Federal University of Alfenas, Unifal-MG, R. Gabriel Monteiro da Silva, 700, 37130-000 Alfenas, MG (Brazil)

    2012-11-15

    The binding interaction between mangiferin (MGF), which a natural xanthone isolated from mangoes, and bovine serum albumin (BSA) was studied with absorbance and fluorescence spectroscopy, cyclic voltammetry and molecular modeling. The data were analyzed to assess the binding mechanism, effect of pH and ionic strength, conformational changes in the protein and electrical charge transfer involved. The MGF-BSA complex exhibited positive cooperativity with a 1:1 stoichiometry (K{sub d}=0.38 mmol L{sup -1}) for the first binding site and a non-saturable binding at high ligand concentrations. Furthermore, the data also suggest an increase in drug bioavailability in the acidic region and relatively low ionic strength values, which are close to physiological levels. The data suggest a specific electrostatic interaction together with hydrophobic effects and H-bonding displayed in MGF binding to the BSA IIA subdomain. Synchronous fluorescence spectra indicate that there are conformational changes in the polypeptide backbone upon ligand binding. Cyclic voltammetry indicates that there is an irreversible charge transfer between MGF and BSA that is modulated by diffusion on the electrode surface, where two electrons are transferred. These results can help the knowledge of the pharmacokinetic activities of natural or chemical xanthone-based drugs. - Highlights: Black-Right-Pointing-Pointer The MGF-BSA complex exhibited positive cooperativity beyond 1:1 stoichiometry. Black-Right-Pointing-Pointer The interaction of MGF with BSA is non-saturable at higher ligand concentration. Black-Right-Pointing-Pointer The binding was accomplished by H-bonding, hydrophobic and electrostatic forces. Black-Right-Pointing-Pointer The apparent binding constant for MGF-BSA was 0.38 mmol L{sup -1}. Black-Right-Pointing-Pointer MGF binds electrostatically to BSA, different from a hydrophobic interaction to HSA.

  17. High pressure phase transition and variation of elastic constants of diluted magnetic semiconductors

    Energy Technology Data Exchange (ETDEWEB)

    Varshney, Dinesh; Sharma, P.; Kaurav, N. [School of Physics, Vigyan Bhawan, Devi Ahilya University, Takshila Campus, Indore 452017 (India); Shah, S. [Department of Physics, P. M. B. Gujarati Science College, Indore-452001 (India); Singh, R.K. [M. P. Bhoj (Open) University, Shivaji Nagar, Bhopal-462016 (India)

    2004-11-01

    A theoretical study of the high-pressure phase transition and elastic behavior in diluted magnetic semiconductors Zn{sub 0.83}Mn{sub 0.17}Se, using a three-body interaction (TBI) potential caused by the electron-shell deformation of the overlapping ions is carried out. The estimated values of phase transition pressure and the vast volume discontinuity in pressure-volume (PV) phase diagram indicate the structural phase transition from zincblende (B3) to rock salt (B1). The variation of second-order elastic constants with pressure resembles that observed in some binary semiconductors. The inconsistency in the deduced value of pressure derivative of second order elastic constant with the available data is attributed to the fact that we derive expressions neglecting thermal effects and assuming the overlap repulsion significant only up to nearest neighbors. The vdW interaction is effective in obtaining the thermodynamical parameters such as Debye temperature, Gruneisen parameter, thermal expansion coefficient, compressibility as well phase stability in diluted magnetic semiconductors. It is revealed that TBI model has a promise to predict the phase transition pressure and the pressure variation of elastic constants of other semiconductors as well. (copyright 2004 WILEY-VCH Verlag GmbH and Co. KGaA, Weinheim) (orig.)

  18. Varying Constants

    CERN Document Server

    Damour, Thibault Marie Alban Guillaume

    2003-01-01

    We review some string-inspired theoretical models which incorporate a correlated spacetime variation of coupling constants while remaining naturally compatible both with phenomenological constraints coming from geochemical data (Oklo; Rhenium decay) and with present equivalence principle tests. Barring unnatural fine-tunings of parameters, a variation of the fine-structure constant as large as that recently ``observed'' by Webb et al. in quasar absorption spectra appears to be incompatible with these phenomenological constraints. Independently of any model, it is emphasized that the best experimental probe of varying constants are high-precision tests of the universality of free fall, such as MICROSCOPE and STEP. Recent claims by Bekenstein that fine-structure-constant variability does not imply detectable violations of the equivalence principle are shown to be untenable.

  19. High-Performance Constant Power Generation in Grid-Connected PV Systems

    DEFF Research Database (Denmark)

    Sangwongwanich, Ariya; Yang, Yongheng; Blaabjerg, Frede

    2016-01-01

    An advanced power control strategy by limiting the maximum feed-in power of PV systems has been proposed, which can ensure a fast and smooth transition between maximum power point tracking and Constant Power Generation (CPG). Regardless of the solar irradiance levels, high-performance and stable...... operation are always achieved by the proposed control strategy. It can regulate the PV output power according to any set-point, and force the PV systems to operate at the left side of the maximum power point without stability problems. Experimental results have verified the effectiveness of the proposed CPG...

  20. Rate constants for chemical reactions in high-temperature nonequilibrium air

    Science.gov (United States)

    Jaffe, R. L.

    1986-01-01

    In the nonequilibrium atmospheric chemistry regime that will be encountered by the proposed Aeroassisted Orbital Transfer Vehicle in the upper atmosphere, where air density is too low for thermal and chemical equilibrium to be maintained, the detailed high temperature air chemistry plays a critical role in defining radiative and convective heating loads. Although vibrational and electronic temperatures remain low (less than 15,000 K), rotational and translational temperatures may reach 50,000 K. Attention is presently given to the effects of multiple temperatures on the magnitudes of various chemical reaction rate constants, for the cases of both bimolecular exchange reactions and collisional excitation and dissociation reactions.

  1. Determination of binding constants and stoichiometries of short-range, hydrogen-bonding solvation by use of a proton-transfer indicator reaction

    Science.gov (United States)

    Schullery, Stephen E.; Wojdyla, Stephen M.; Ostroski, Robert A.; Scott, Ronald M.

    1997-10-01

    A recent method for determination of stoichiometries and binding constants for short-range, hydrogen bonding solvation is reviewed and new results are presented. The method exploits the sensitivity of a proton-transfer equilibrium, KPT to changes in solvent composition. Solvation numbers and binding constants for primary and secondary stages of solvation of an aminephenol proton-transfer adduct and the phenol and amine are determined as adjustable parameters when model isotherms are fitted to KPT versus [ S] data, where [ S] is the concentration of a hydrogen-bonding minor component of a mixed solvent, Results for most of the twenty-three aprotic solvents investigated are modeled by bifurcation-type hydrogen bonding of two or more electron pairs to a single polar hydrogen. Results for the seven protic solvents studied, including new data for ethanol, 2-choloroethanol, and 2,2-dichloroethanol, are modeled by two, or possibly three, successive stages of solvation, assumed to involve hydrogen-bonded chains. Preliminary results indicate that solvation by water is amenable to this analysis.

  2. Sulfone-Containing Dipolar Glass Polymers with High Dielectric Constant and Low Loss Property

    Science.gov (United States)

    Zhu, Yufeng; Zhang, Zhongbo; Litt, Morton; Zhu, Lei

    Sulfone-containing polyoxetanes are designed and synthesized for high dielectric constant and low loss dipolar glasses. The precursor polymer, poly(3,3-bis(chloromethyl)oxetane) (PBCMO) is synthesized by bulk cationic polymerization with boron trifluoride diethyl etherate as initiator. The number-average molecular weight of PBCMO is 73 kDa, with a polydispersity of 1.53 as obtained from size-exclusion chromatography results. Post-modification of PBCMO yields the dipolar glass polymer, poly(3,3-bis(methylsulfonylmethyl)oxetane) (MST). Nuclear magnetic resonance result shows 100% conversion. Differential scanning calorimetry result indicates that MST has a glass transition temperature of ca. 120 °C. Due to the large dipole moment (4.25 D) and small size of the side-chain sulfone groups, MST exhibits a high dielectric constant of 8.7 and a low dissipation factor of 0.01 at 25 °C and 1 Hz. This study suggests that dipolar glass polymers with large dipole moments and small-sized dipoles in the side chains are promising candidates for high energy density and low loss dielectric applications. This work is supported by NSF Polymers Program (DMR-1402733).

  3. Nanocomposites of TiO₂/cyanoethylated cellulose with ultra high dielectric constants.

    Science.gov (United States)

    Madusanka, Nadeesh; Shivareddy, Sai G; Hiralal, Pritesh; Eddleston, Mark D; Choi, Youngjin; Oliver, Rachel A; Amaratunga, Gehan A J

    2016-05-13

    A novel dielectric nanocomposite containing a high permittivity polymer, cyanoethylated cellulose (CRS) and TiO2 nanoparticles was successfully prepared with different weight percentages (10%, 20% and 30%) of TiO2. The intermolecular interactions and morphology within the polymer nanocomposites were analysed. TiO2/CRS nanofilms on SiO2/Si wafers were used to form metal-insulator-metal type capacitors. Capacitances and loss factors in the frequency range of 1 kHz-1 MHz were measured. At 1 kHz CRS-TiO2 nanocomposites exhibited ultra high dielectric constants of 118, 176 and 207 for nanocomposites with 10%, 20% and 30% weight of TiO2 respectively, significantly higher than reported values of pure CRS (21), TiO2 (41) and other dielectric polymer-TiO2 nanocomposite films. Furthermore, all three CRS-TiO2 nanocomposites show a loss factor dielectric constant hybrids using nanoscale inorganic dielectrics dispersed in a high permittivity polymer suitable for energy management applications is reported.

  4. Nanocomposites of TiO2/cyanoethylated cellulose with ultra high dielectric constants

    Science.gov (United States)

    Madusanka, Nadeesh; Shivareddy, Sai G.; Hiralal, Pritesh; Eddleston, Mark D.; Choi, Youngjin; Oliver, Rachel A.; Amaratunga, Gehan A. J.

    2016-05-01

    A novel dielectric nanocomposite containing a high permittivity polymer, cyanoethylated cellulose (CRS) and TiO2 nanoparticles was successfully prepared with different weight percentages (10%, 20% and 30%) of TiO2. The intermolecular interactions and morphology within the polymer nanocomposites were analysed. TiO2/CRS nanofilms on SiO2/Si wafers were used to form metal-insulator-metal type capacitors. Capacitances and loss factors in the frequency range of 1 kHz-1 MHz were measured. At 1 kHz CRS-TiO2 nanocomposites exhibited ultra high dielectric constants of 118, 176 and 207 for nanocomposites with 10%, 20% and 30% weight of TiO2 respectively, significantly higher than reported values of pure CRS (21), TiO2 (41) and other dielectric polymer-TiO2 nanocomposite films. Furthermore, all three CRS-TiO2 nanocomposites show a loss factor dielectric constant hybrids using nanoscale inorganic dielectrics dispersed in a high permittivity polymer suitable for energy management applications is reported.

  5. The effect of dielectric constant on binding energy and impurity self-polarization in a GaAs-Ga1- x Al x As spherical quantum dot

    Science.gov (United States)

    Mese, A. I.; Cicek, E.; Erdogan, I.; Akankan, O.; Akbas, H.

    2017-03-01

    The ground state, 1s, and the excited state, 2p, energies of a hydrogenic impurity in a GaAs-Ga1- x Al x As spherical quantum dot, are computed as a function of the donor positions. We study how the impurity self-polarization depends on the location of the impurity and the dielectric constant. The excited state anomalous impurity self-polarization in the quantum dot is found to be present in the absence of any external influence and strongly depends on the impurity position and the radius of the dot. Therefore, the excited state anomalous impurity self-polarization can give information about the impurity position in the system. Also, the variation of E_{b1s} and E_{b2p} with the dielectric constant can be utilized as a tool for finding out the correct dielectric constant of the dot material by measuring the 1s or 2p state binding energy for a fixed dot radius and a fixed impurity position.

  6. Liquid Crystalline Epoxies with Lateral Substituents Showing a Low Dielectric Constant and High Thermal Conductivity

    Science.gov (United States)

    Guo, Huilong; Lu, Mangeng; Liang, Liyan; Wu, Kun; Ma, Dong; Xue, Wei

    2017-02-01

    In this work, liquid crystalline epoxies with lateral substituents were synthesized and cured with aromatic amines or anhydride. The liquid crystalline phase structure of liquid crystalline epoxies with lateral substituents was determined by polarized optical microscopy. The relationship between thermal conductivity and dielectric properties and liquid crystalline domain structure was discussed in the paper. The samples show high thermal conductivity up to 0.29 W/(m × K), due to the orientation of mesogenic units in epoxies. The sample's low dielectric constant of 2.29 is associated with the oriented mesogenic units and long nonpolar lateral substituents. This indicates a new way to obtain materials with high thermal conductivity and a low dielectric constant by introducing oriented mesogenic units into cross-linked epoxy systems. The water repellency is reflected in the contact angles of 92-98°, which are apparently higher than that of conventional epoxy systems. It was also found that the better toughness of liquid crystalline epoxies with lateral substituents was attributed to the existence of long flexible alkyl lateral substituents.

  7. Beta 1 integrin binding plays a role in the constant traction force generation in response to varying stiffness for cells grown on mature cardiac extracellular matrix.

    Science.gov (United States)

    Gershlak, Joshua R; Black, Lauren D

    2015-01-15

    We have previously reported a unique response of traction force generation for cells grown on mature cardiac ECM, where traction force was constant over a range of stiffnesses. In this study we sought to further investigate the role of the complex mixture of ECM on this response and assess the potential mechanism behind it. Using traction force microscopy, we measured cellular traction forces and stresses for mesenchymal stem cells (MSCs) grown on polyacrylamide gels at a range of stiffnesses (9, 25, or 48 kPa) containing either adult rat heart ECM, different singular ECM proteins including collagen I, fibronectin, and laminin, or ECM mimics comprised of varying amounts of collagen I, fibronectin, and laminin. We also measured the expression of integrins on these different substrates as well as probed for β1 integrin binding. There was no significant change in traction force generation for cells grown on the adult ECM, as previously reported, whereas cells grown on singular ECM protein substrates had increased traction force generation with an increase in substrate stiffness. Cells grown on ECM mimics containing collagen I, fibronectin and laminin were found to be reminiscent of the traction forces generated by cells grown on native ECM. Integrin expression generally increased with increasing stiffness except for the β1 integrin, potentially implicating it as playing a role in the response to adult cardiac ECM. We inhibited binding through the β1 integrin on cells grown on the adult ECM and found that the inhibition of β1 binding led to a return to the typical response of increasing traction force generation with increasing stiffness. Our data demonstrates that cells grown on the mature cardiac ECM are able to circumvent typical stiffness related cellular behaviors, likely through β1 integrin binding to the complex composition. Copyright © 2014 Elsevier Inc. All rights reserved.

  8. Evidence for a Constant `Edge' in Proton-Proton Scattering at Very High Energies

    CERN Document Server

    Block, Martin M; Halzen, Francis; Stodolsky, Leo; Weiler, Thomas J

    2014-01-01

    Accurate fits to $pp$ and $\\bar pp$ cross section data up to Tevatron energies, incorporating the constraints imposed by analyticity and unitarity, successfully predict the results of recent LHC and cosmic ray measurements, and suggest that the cross sections approach a black disc limit asymptotically. The approach to the limit is, however, very slow. We present a simple geometric picture which explains these features in a natural way. A black disc of logarithmically growing radius is supplemented by a soft `edge' whose properties are invariant with energy. The constancy of the edge results in the prediction that the quantity $(\\sigma^{TOT}-2\\sigma^{El})/\\surd\\sigma^{TOT}$ approaches a constant at high energy. Using the existing fits, this prediction appears to be verified. The value of the limiting constant allows an estimate of the thickness of the edge, which turns out to be on the order of $1\\,{\\rm fm}$. One thus arrives at a picture where the proton-proton scattering at lower energies is dominated by wha...

  9. Single-experiment displacement assay for quantifying high-affinity binding by isothermal titration calorimetry.

    Science.gov (United States)

    Krainer, Georg; Keller, Sandro

    2015-04-01

    Isothermal titration calorimetry (ITC) is the gold standard for dissecting the thermodynamics of a biomolecular binding process within a single experiment. However, reliable determination of the dissociation constant (KD) from a single titration is typically limited to the range 100 μM>KD>1 nM. Interactions characterized by a lower KD can be assessed indirectly by so-called competition or displacement assays, provided that a suitable competitive ligand is available whose KD falls within the directly accessible window. However, this protocol is limited by the fact that it necessitates at least two titrations to characterize one high-affinity inhibitor, resulting in considerable consumption of both sample material and time. Here, we introduce a fast and efficient ITC displacement assay that allows for the simultaneous characterization of both a high-affinity ligand and a moderate-affinity ligand competing for the same binding site on a receptor within a single experiment. The protocol is based on a titration of the high-affinity ligand into a solution containing the moderate-affinity ligand bound to the receptor present in excess. The resulting biphasic binding isotherm enables accurate and precise determination of KD values and binding enthalpies (ΔH) of both ligands. We discuss the theoretical background underlying the approach, demonstrate its practical application to metal ion chelation, explore its potential and limitations with the aid of simulations and statistical analyses, and elaborate on potential applications to protein-inhibitor interactions.

  10. Optical transitions in highly charged californium ions with high sensitivity to variation of the fine-structure constant.

    Science.gov (United States)

    Berengut, J C; Dzuba, V A; Flambaum, V V; Ong, A

    2012-08-17

    We study electronic transitions in highly charged Cf ions that are within the frequency range of optical lasers and have very high sensitivity to potential variations in the fine-structure constant, α. The transitions are in the optical range despite the large ionization energies because they lie on the level crossing of the 5f and 6p valence orbitals in the thallium isoelectronic sequence. Cf(16+) is a particularly rich ion, having several narrow lines with properties that minimize certain systematic effects. Cf(16+) has very large nuclear charge and large ionization energy, resulting in the largest α sensitivity seen in atomic systems. The lines include positive and negative shifters.

  11. Procedural Adaptations for Use of Constant Time Delay to Teach Highly Motivating Words to Beginning Braille Readers

    Science.gov (United States)

    Ivy, Sarah E.; Guerra, Jennifer A.; Hatton, Deborah D.

    2017-01-01

    Introduction: Constant time delay is an evidence-based practice to teach sight word recognition to students with a variety of disabilities. To date, two studies have documented its effectiveness for teaching braille. Methods: Using a multiple-baseline design, we evaluated the effectiveness of constant time delay to teach highly motivating words to…

  12. High-Dielectric-Constant Silver-Epoxy Composites as Embedded Dielectrics

    Energy Technology Data Exchange (ETDEWEB)

    Qi, Lai; Lee, Burtrand I.; Chen, Sihai; Samuels, William D.; Exarhos, Gregory J.

    2005-07-08

    We report the synthesis of an epoxy-based composite containing randomly distributed silver nanoparticles with a narrow size distribution centered around 40nm. The surface of the Ag nanoparticles is coated with a thin layer of mercaptosuccinic acid (MSA) in order to promote formation of the Ag-epoxy while retaining flexibility of the polymer matrix. This composite is the basis of a prototype capacitor formed by spin coating the synthesized Ag-epoxy (40nm Ag) mixture onto a gold-deposited aluminum substrate. A series of Ag-epoxy composites with various Ag volume fractions was investigated in order to determine the composition associated with optimum capacitor performance. The high dielectric constant (> 300) and relatively low dielectric loss (<0.05) render this material of promising potential for embedded capacitor applications.

  13. Polymer thin-film transistor based on a high dielectric constant gate insulator

    Institute of Scientific and Technical Information of China (English)

    Lü Wen; Peng Jun-Biao; Yang Kai-Xia; Lan Lin-Feng; Niu Qiao-Li; Cao Yong

    2007-01-01

    In this paper full polymer thin-film transistors (PTFTs) based on Poly (acrylonitrile) (PAN) as the gate dielectric and poly (2-methoxy-5-(2'-ethyl-hexyloxy)-1,4-phenylene-vinylene) (MEH-PPV) as the semiconductor layer were investigated by using different channel width/length ratios. Relatively high dielectric constant of the polymer dielectric layer (6.27) can remarkably reduce the threshold voltage of the transistors to below-3 V. Hole field-effect mobility of MEH-PPV of the PTFTs was about 4.8 × 10-4 cm2/Vs, and on/off current ratio was larger than 102, which was comparable with that of transistors with widely used Poly (4-vinyl phenol) (PVP) or SiO2 as gate dielectrics.

  14. Fabrication of europium-doped silica optical fiber with high Verdet constant.

    Science.gov (United States)

    Huang, Yi; Chen, Huangchao; Dong, Weilong; Pang, Fufei; Wen, Jianxiang; Chen, Zhenyi; Wang, Tingyun

    2016-08-08

    A europium-doped (Eu-doped) silica optical fiber is fabricated using modified chemical vapor deposition (MCVD) technology. Europium fluoride (EuF3) material is introduced into the fiber core with a high temperature vaporizing technique. Its concentration is approximately 0.11 at %. The outer and core diameters of doped fiber are approximately 122 and 9 μm, respectively. Refractive index difference (RID) between core and cladding is approximately 2%. A magneto-optical effect measurement system, which is based on the Stokes polarization parameters method, is set up to analyze its magneto-optical properties. The Verdet constant of the Eu-doped optical fiber is -4.563 rad T-1m-1, which is approximately double than that of single mode fiber (SMF) at 660 nm.

  15. Antigen-antibody selective recognition using LiTaO3 SH-SAW sensors: investigations on macromolecules effects on binding kinetic constants

    Science.gov (United States)

    Bergaoui, Y.; Zerrouki, C.; Fourati, N.; Fougnion, J. M.; Abdelghani, A.

    2011-10-01

    A gravimetric surface acoustic wave (SAW) biosensor, based on the biotin-streptavidin and antistreptavidin-streptavidin recognitions, has been carried out. A network analyser and a pulse excitation technique were used to monitor both amplitude and phase changes. The SAW biosensor presented a total selective recognition of streptavidin-antistreptavidin and HRPstreptavidin-antistreptavidin. The presence of HRP (Horseradish peroxidase) affects neither the selectivity nor the sensitivity (of order of 0.25°/nM) of the biosensor, nevertheless, it causes a reduction of binding kinetics by a factor ranging between 2 to 5, as well as a diminution of antistreptavidin saturation concentration (of 40%). Results showed that equilibrium constants can be different, depending on evaluation method (from saturation values or from linear part of the output signal variation according to solution concentration).

  16. Quantifying high-affinity binding of hydrophobic ligands by isothermal titration calorimetry.

    Science.gov (United States)

    Krainer, Georg; Broecker, Jana; Vargas, Carolyn; Fanghänel, Jörg; Keller, Sandro

    2012-12-18

    A fast and reliable quantification of the binding thermodynamics of hydrophobic high-affinity ligands employing a new calorimetric competition experiment is described. Although isothermal titration calorimetry is the method of choice for a quantitative characterization of intermolecular interactions in solution, a reliable determination of a dissociation constant (K(D)) is typically limited to the range 100 μM > K(D) > 1 nM. Interactions displaying higher or lower K(D) values can be assessed indirectly, provided that a suitable competing ligand is available whose K(D) falls within the directly accessible affinity window. This established displacement assay, however, requires the high-affinity ligand to be soluble at high concentrations in aqueous buffer and, consequently, poses serious problems in the study of protein binding involving small-molecule ligands dissolved in organic solvents--a familiar case in many drug-discovery projects relying on compound libraries. The calorimetric competition assay introduced here overcomes this limitation, thus allowing for a detailed thermodynamic description of high-affinity receptor-ligand interactions involving poorly water-soluble compounds. Based on a single titration of receptor into a dilute mixture of the two competing ligands, this competition assay provides accurate and precise values for the dissociation constants and binding enthalpies of both high- and moderate-affinity ligands. We discuss the theoretical background underlying the approach, demonstrate its practical application to metal ion chelation and high-affinity protein-inhibitor interactions, and explore its potential and limitations with the aid of simulations and statistical analyses.

  17. Estimation of the pH-independent binding constants of alanylphenylalanine and leucylphenylalanine stereoisomers with beta-cyclodextrin in the presence of urea.

    Science.gov (United States)

    Li, J; Waldron, K C

    1999-01-01

    The separation of stereoisomers, particularly enantiomers, is important when their physiological activity differs. We have resolved the four stereoisomers each of alanylphenylalanine (Ala-Phe) and of leucylphenylalanine (Leu-Phe) by capillary electrophoresis using beta-cyclodextrin as a buffer additive and urea to enhance its solubility. A study of the influence of pH and beta-cyclodextrin concentration on the separations showed that weak inclusion complexes were formed between the dipeptides and chiral selector. It was found that pH could alter the migration order of enantiomers L-Ala-L-Phe and D-Ala-D-Phe, as well as L-Leu-L-Phe and D-Leu-D-Phe; however, there was no change in order for the other pairs of optical isomers. Electrophoretic mobility data were used to estimate the acid dissociation constants of the dipeptide isomers at pH < 7 with no chiral selector present. By varying the concentration of beta-cyclodextrin, the chiral selector, the binding constants of Ala-Phe and Leu-Phe optical isomers in their fully protonated and zwitterionic forms were estimated. For the four Ala-Phe stereoisomers, K = 42-66 M(-1) and 4-41 M(-1) for the cationic and zwitterionic forms, respectively. For the four Leu-Phe stereoisomers, K = 43-94 M(-1) and 1-28 M(-1) for the cationic and zwitterionic forms, respectively.

  18. A high efficiency and power factor, segmented linear constant current LED driver

    Science.gov (United States)

    Yongyuan, Li; Wei, Guo; Zhangming, Zhu

    2015-04-01

    A high efficiency, high power factor, and linear constant current LED driver based on adaptive segmented linear architecture is presented. When the input voltage varied, the proposed LED driver automatically switched over LED strings according to the segmented LED voltage drop, which increased the LED lighting time. The efficiency and power factor are improved, while the system design is simplified by this control scheme. Without the usage of electrolytic capacitor and magnetic components, the proposed driver possesses advantages of smaller size, longer lifetime and lower cost over others. The proposed driver is implemented in 0.8 μm 5 V/40 V HVCMOS process, which occupies an active area of 820 × 920 μm2. The measured results show that the average value of the internal reference voltage is 500 ± 7 mV, with a standard deviation of only 4.629 mV, thus LED current can be set accurately. Under 220 V root mean square 50 Hz utility voltage and the number ratio of the three LED strings being 47 : 17 : 16, the system can realize a high power factor of 0.974 and power conversion efficiency of 93.4%. Project supported by the National Natural Science Foundation of China (Nos. 61234002, 61322405, 61306044, 61376033).

  19. Comprehensive Study of Lanthanum Aluminate High-Dielectric-Constant Gate Oxides for Advanced CMOS Devices

    Directory of Open Access Journals (Sweden)

    Masamichi Suzuki

    2012-03-01

    Full Text Available A comprehensive study of the electrical and physical characteristics of Lanthanum Aluminate (LaAlO3 high-dielectric-constant gate oxides for advanced CMOS devices was performed. The most distinctive feature of LaAlO3 as compared with Hf-based high-k materials is the thermal stability at the interface with Si, which suppresses the formation of a low-permittivity Si oxide interfacial layer. Careful selection of the film deposition conditions has enabled successful deposition of an LaAlO3 gate dielectric film with an equivalent oxide thickness (EOT of 0.31 nm. Direct contact with Si has been revealed to cause significant tensile strain to the Si in the interface region. The high stability of the effective work function with respect to the annealing conditions has been demonstrated through comparison with Hf-based dielectrics. It has also been shown that the effective work function can be tuned over a wide range by controlling the La/(La + Al atomic ratio. In addition, gate-first n-MOSFETs with ultrathin EOT that use sulfur-implanted Schottky source/drain technology have been fabricated using a low-temperature process.

  20. Organic/Inorganic Nano-hybrids with High Dielectric Constant for Organic Thin Film Transistor Applications

    Science.gov (United States)

    Yu, Yang-Yen; Jiang, Ai-Hua; Lee, Wen-Ya

    2016-11-01

    The organic material soluble polyimide (PI) and organic-inorganic hybrid PI-barium titanate (BaTiO3) nanoparticle dielectric materials (IBX, where X is the concentration of BaTiO3 nanoparticles in a PI matrix) were successfully synthesized through a sol-gel process. The effects of various BaTiO3 contents on the hybrid film performance and performance optimization were investigated. Furthermore, pentacene-based organic thin film transistors (OTFTs) with PI-BaTiO3/polymethylmethacrylate or cyclic olefin copolymer (COC)-modified gate dielectrics were fabricated and examined. The hybrid materials showed effective dispersion of BaTiO3 nanoparticles in the PI matrix and favorable thermal properties. X-ray diffraction patterns revealed that the BaTiO3 nanoparticles had a perovskite structure. The hybrid films exhibited high formability and planarity. The IBX hybrid dielectric films exhibited tunable insulating properties such as the dielectric constant value and capacitance in ranges of 4.0-8.6 and 9.2-17.5 nF cm-2, respectively. Adding the modified layer caused the decrease of dielectric constant values and capacitances. The modified dielectric layer without cross-linking displayed a hydrophobic surface. The electrical characteristics of the pentacene-based OTFTs were enhanced after the surface modification. The optimal condition for the dielectric layer was 10 wt% hybrid film with the COC-modified layer; moreover, the device exhibited a threshold voltage of 0.12 V, field-effect mobility of 4.32 × 10-1 cm2 V-1 s-1, and on/off current of 8.4 × 107.

  1. A Unitary Anesthetic Binding Site at High Resolution

    Energy Technology Data Exchange (ETDEWEB)

    Vedula, L. Sangeetha; Brannigan, Grace; Economou, Nicoleta J.; Xi, Jin; Hall, Michael A.; Liu, Renyu; Rossi, Matthew J.; Dailey, William P.; Grasty, Kimberly C.; Klein, Michael L.; Eckenhoff, Roderic G.; Loll, Patrick J.; (Drexel-MED); (UPENN)

    2009-10-21

    Propofol is the most widely used injectable general anesthetic. Its targets include ligand-gated ion channels such as the GABA{sub A} receptor, but such receptor-channel complexes remain challenging to study at atomic resolution. Until structural biology methods advance to the point of being able to deal with systems such as the GABA{sub A} receptor, it will be necessary to use more tractable surrogates to probe the molecular details of anesthetic recognition. We have previously shown that recognition of inhalational general anesthetics by the model protein apoferritin closely mirrors recognition by more complex and clinically relevant protein targets; here we show that apoferritin also binds propofol and related GABAergic anesthetics, and that the same binding site mediates recognition of both inhalational and injectable anesthetics. Apoferritin binding affinities for a series of propofol analogs were found to be strongly correlated with the ability to potentiate GABA responses at GABA{sub A} receptors, validating this model system for injectable anesthetics. High resolution x-ray crystal structures reveal that, despite the presence of hydrogen bond donors and acceptors, anesthetic recognition is mediated largely by van der Waals forces and the hydrophobic effect. Molecular dynamics simulations indicate that the ligands undergo considerable fluctuations about their equilibrium positions. Finally, apoferritin displays both structural and dynamic responses to anesthetic binding, which may mimic changes elicited by anesthetics in physiologic targets like ion channels.

  2. A Unitary Anesthetic Binding Site at High Resolution

    Energy Technology Data Exchange (ETDEWEB)

    L Vedula; G Brannigan; N Economou; J Xi; M Hall; R Liu; M Rossi; W Dailey; K Grasty; et. al.

    2011-12-31

    Propofol is the most widely used injectable general anesthetic. Its targets include ligand-gated ion channels such as the GABA{sub A} receptor, but such receptor-channel complexes remain challenging to study at atomic resolution. Until structural biology methods advance to the point of being able to deal with systems such as the GABA{sub A} receptor, it will be necessary to use more tractable surrogates to probe the molecular details of anesthetic recognition. We have previously shown that recognition of inhalational general anesthetics by the model protein apoferritin closely mirrors recognition by more complex and clinically relevant protein targets; here we show that apoferritin also binds propofol and related GABAergic anesthetics, and that the same binding site mediates recognition of both inhalational and injectable anesthetics. Apoferritin binding affinities for a series of propofol analogs were found to be strongly correlated with the ability to potentiate GABA responses at GABA{sub A} receptors, validating this model system for injectable anesthetics. High resolution x-ray crystal structures reveal that, despite the presence of hydrogen bond donors and acceptors, anesthetic recognition is mediated largely by van der Waals forces and the hydrophobic effect. Molecular dynamics simulations indicate that the ligands undergo considerable fluctuations about their equilibrium positions. Finally, apoferritin displays both structural and dynamic responses to anesthetic binding, which may mimic changes elicited by anesthetics in physiologic targets like ion channels.

  3. A Unitary Anesthetic-Binding Site at High Resolution

    Energy Technology Data Exchange (ETDEWEB)

    Vedula, L.; Brannigan, G; Economou, N; Xi, J; Hall, M; Liu, R; Rossi, M; Dailey, W; Grasty, K; et. al.

    2009-01-01

    Propofol is the most widely used injectable general anesthetic. Its targets include ligand-gated ion channels such as the GABAA receptor, but such receptor-channel complexes remain challenging to study at atomic resolution. Until structural biology methods advance to the point of being able to deal with systems such as the GABA{sub A} receptor, it will be necessary to use more tractable surrogates to probe the molecular details of anesthetic recognition. We have previously shown that recognition of inhalational general anesthetics by the model protein apoferritin closely mirrors recognition by more complex and clinically relevant protein targets; here we show that apoferritin also binds propofol and related GABAergic anesthetics, and that the same binding site mediates recognition of both inhalational and injectable anesthetics. Apoferritin binding affinities for a series of propofol analogs were found to be strongly correlated with the ability to potentiate GABA responses at GABA{sub A} receptors, validating this model system for injectable anesthetics. High resolution x-ray crystal structures reveal that, despite the presence of hydrogen bond donors and acceptors, anesthetic recognition is mediated largely by van der Waals forces and the hydrophobic effect. Molecular dynamics simulations indicate that the ligands undergo considerable fluctuations about their equilibrium positions. Finally, apoferritin displays both structural and dynamic responses to anesthetic binding, which may mimic changes elicited by anesthetics in physiologic targets like ion channels.

  4. Effects of high-frequency emphasis and compression time constants on speech intelligibility in noise.

    Science.gov (United States)

    van Toor, Thijs; Verschuure, Hans

    2002-10-01

    The objectives of the study were to evaluate the effect of different settings with regard to speech intelligibility in noise both objectively and subjectively and thus determine a favoured setting of compression time parameters, pre-set program (high-frequency emphasis) or combination for each individual user in a prospective study. Another objective was to evaluate the relationship between patient characteristics (e.g. slope of hearing loss) and favoured settings. In total, 38 subjects divided over five audiological centres were fitted with the Philips Spaceline D71-40 BTE digital hearing aid. Subjects were asked to compare three predefined compression algorithms with different time constants, slow (indicated by the manufacturer as AVC), intermediate (NORMAL) and fast (SYLLABIC) over two 4-week periods using the intermediate setting in both comparisons and randomizing over the fast and slow conditions. A randomization determined whether a subject started with the comfort-oriented pre-set program (AUTO) or the speech intelligibility-oriented setting with high-frequency emphasis (SPIN). In a third 4-week period, the pre-sets AUTO and SPIN were compared using the setting of the compression time constants that gave the best results during the first two periods. Comparisons were made using a standard speech-in-noise test with three types of noise: continuous speaker noise, modulated ICRA-4 noise, and car noise. The patients were also asked to fill in a Dutch translation and adaptation of the APHAB questionnaire to indicate their impression of performance. The results indicate that no compression algorithm, pre-set or combination is favoured overall. The largest improvement in speech-in-noise scores was found with syllabic compression. The advantageous effect of high-frequency emphasis after optimization of compression timing is small. The APHAB showed that users tend to prefer the SPIN setting. We found no relationship between favoured compression or pre-set and the

  5. High molecular weight polysaccharide that binds and inhibits virus

    Science.gov (United States)

    Konowalchuk, Thomas W

    2014-01-14

    This invention provides a high molecular weight polysaccharide capable of binding to and inhibiting virus and related pharmaceutical formulations and methods on inhibiting viral infectivity and/or pathogenicity, as well as immunogenic compositions. The invention further methods of inhibiting the growth of cancer cells and of ameliorating a symptom of aging. Additionally, the invention provides methods of detecting and/or quantifying and/or isolating viruses.

  6. High molecular weight polysaccharide that binds and inhibits virus

    Energy Technology Data Exchange (ETDEWEB)

    Konowalchuk, Thomas W.; Konowalchuk, Jack

    2017-07-18

    This invention provides a high molecular weight polysaccharide capable of binding to and inhibiting virus and related pharmaceutical formulations and methods of inhibiting viral infectivity and/or pathogenicity, as well as immunogenic compositions. The invention further includes methods of inhibiting the growth of cancer cells and of ameliorating a symptom of aging. Additionally, the invention provides methods of detecting and/or quantifying and/or isolating viruses.

  7. The high-affinity peptidoglycan binding domain of Pseudomonas phage endolysin KZ144

    Energy Technology Data Exchange (ETDEWEB)

    Briers, Yves [Division of Gene Technology, Department of Biosystems, Katholieke Universiteit Leuven, Kasteelpark Arenberg 21, B-3001 Leuven (Belgium); Schmelcher, Mathias; Loessner, Martin J. [Institute of Food Science and Nutrition, ETH Zuerich, Schmelzbergstrasse 7, CH-8092 Zuerich (Switzerland); Hendrix, Jelle; Engelborghs, Yves [Laboratory of Biomolecular Dynamics, Department of Chemistry, Katholieke Universiteit Leuven, Celestijnenlaan 200G, B-3001 Leuven (Belgium); Volckaert, Guido [Division of Gene Technology, Department of Biosystems, Katholieke Universiteit Leuven, Kasteelpark Arenberg 21, B-3001 Leuven (Belgium); Lavigne, Rob, E-mail: rob.lavigne@biw.kuleuven.be [Division of Gene Technology, Department of Biosystems, Katholieke Universiteit Leuven, Kasteelpark Arenberg 21, B-3001 Leuven (Belgium)

    2009-05-29

    The binding affinity of the N-terminal peptidoglycan binding domain of endolysin KZ144 (PBD{sub KZ}), originating from Pseudomonas aeruginosa bacteriophage {phi}KZ, has been examined using a fusion protein of PBD{sub KZ} and green fluorescent protein (PBD{sub KZ}-GFP). A fluorescence recovery after photobleaching analysis of bound PBD{sub KZ}-GFP molecules showed less than 10% fluorescence recovery in the bleached area within 15 min. Surface plasmon resonance analysis confirmed this apparent high binding affinity revealing an equilibrium affinity constant of 2.95 x 10{sup 7} M{sup -1} for the PBD{sub KZ}-peptidoglycan interaction. This unique domain, which binds to the peptidoglycan of all tested Gram-negative species, was harnessed to improve the specific activity of the peptidoglycan hydrolase domain KMV36C. The chimeric peptidoglycan hydrolase (PBD{sub KZ}-KMV36C) exhibits a threefold higher specific activity than the native catalytic domain (KMV36C). These results demonstrate that the modular assembly of functional domains is a rational approach to improve the specific activity of endolysins from phages infecting Gram-negatives.

  8. An investigation of constant pressure gas well testing influenced by high velocity flow

    Energy Technology Data Exchange (ETDEWEB)

    Berumen, S. [PEMEX Exploracion-Produccion, Mexico City (Mexico); Samaniego, F. [Universidad de Mexico, Mexico City (Mexico). Facultad de Ingeniera; Cinco-Ley, H. [Universidad de Mexico, Mexico City (Mexico). Facultad de Ingeniera; Bouhroum, A.

    1997-03-01

    This paper presents the results of a study of transient pressure analysis of gas flow under either constant bottom-hole conditions or the constant wellhead pressure conditions. The effect of formation damage, wellbore storage and high velocity flow are included in the model. The analysis of simulated well tests showed that the interpretation methods used for liquid flow are generally accurate when the p{sub p}(p) is used. For these conditions, a graph of 1/q{sub D} vs log t{sub D} presents gradually lower values of 1.1513 as the value of p{sub wf} decreases: For pressure buildup conditions a graph of p{sub pD}(1, {Delta}t{sub aD})/q{sub D}({Delta}t{sub aD}=0) vs (t{sub aD}+{Delta}t{sub aD})/{Delta}t{sub aD} shows values of this slope within 1% of the 1.1513 value. The maximum error was in the rate performance simulated cases that included high-velocity flows; being less than 13%. This upper limit occurs when the formation has a relatively `high` permeability (around 1 mD) and the rate performance test is affected by high-velocity flow. It was found that pressure buildup tests are superior to rate performance tests because high-velocity flow does not affect the slope of the straight line portion of the buildup curve. However, it was also found, through derivative analysis of simulated buildup tests, that the skin factor is sensibly miscalculated when the high-velocity flow effect is singificant. This problem could lead to errors in the calculation of the skin factor, s, up to 300%. (orig.) [Deutsch] Vorgestellt werden instationaere Testergebnisse an Gas-Sonden unter konstanten Bohrlochsohlenbedingungen bzw. konstantem Bohrlochkopfdruck. Folgende Stoereffekte: Sondennahe Tragerschaedigung, Speicherkapazitaet des Bohrloches und die bei der Gasstroemung eintretende hohe Fliessgeschwindigkeit werden beruecksichtigt. Die Auswertung von simulierten Testergebnissen zeigt, dass die zur Interpretation von Erdoelsonden bewaehrten Verfahren in der Darstellung p{sub p}(p) gute

  9. RELATIONSHIP BETWEEN BLOOD LACTATE AND HYPERVENTILATION DURING HIGH-INTENSITY CONSTANT-LOAD EXERCISE IN HEAT

    Directory of Open Access Journals (Sweden)

    T. Chiba

    2011-09-01

    Full Text Available The purpose of this study was to examine the relationship between hyperventilation and increase in blood lactate during high-intensity constant-load exercise in heat and normal conditions. Seven male volunteers exercised for 10 min on a cycle ergometer at 80%·VO2max in heat (40ºC, 50%relative humidity: HT and normal conditions (20ºC, 50% relative humidity: CON. Oxygen uptake, carbon dioxide output, ventilation, blood lactate and blood electrolytes (K , Na , Cl− were measured in HT and CON. We found that ventilation was significantly higher during exercise in HT compared with CON (p<0.05 and RER tends to be higher in HT than in CON. Blood lactate was significantly higher at 3 min during exercise in HT compared with CON (5.96 ± 0.57 mEq·l-1 5.00 ± 0.28 mEq·l-1, p<0.05. Change in strong ion difference [∆SID = (∆K ∆Na − (∆Cl− ∆La−], which affects ∆HCO3− in blood significantly, was lower at 5 min during exercise in HT compared with in CON (p<0.05. These results suggest that hyperventilation during exercise in heat would induce lower HCO3− in blood and consequently would result in an increase in blood lactate at an earlier time during high-intensity exercise in heat. It was concluded that hyperventilation during short-term high-intensity exercise in heat is temporarily associated with an increase in blood lactate.

  10. Extraordinary high dielectric constant, electrical and magnetic properties of ferrite nanoparticles at room temperature

    Science.gov (United States)

    Batoo, Khalid Mujasam; Mir, Feroz Ahmed; Abd El-sadek, M.-S.; Shahabuddin, Md.; Ahmed, Niyaz

    2013-11-01

    Nanoparticles of spinel ferrites of basic composition Ni1- x Co x Fe2O4 (0.0 ≤ x ≤ 0.05) were synthesized through modified co-precipitation method, and were characterized for structural, transport electrical and magnetic properties using XRD, HRTEM, FTIR, LCR meter and VSM techniques, respectively. XRD analysis showed that all the samples are single-phase cubic spinel in structure. The average crystallite sizes of the nanoparticles were found between 30 nm to 45 nm. Real and imaginary parts of the impedance ( Z' and Z″) suggested coexistence of two relaxation regimes: one was introduced by electrode polarization, while the other was attributed to the coeffect of grain and grain boundary effects. The dielectric constant of the samples was found very high, which showed non-Debye relaxation phenomena, while conductivity of the samples exhibited a two-segment behavior with frequency. The room temperature M-H curves suggested that the samples exhibit supermagnetism, and the saturation magnetization increases with increasing Co2+ ion substitution.

  11. Extraordinary high dielectric constant, electrical and magnetic properties of ferrite nanoparticles at room temperature

    Energy Technology Data Exchange (ETDEWEB)

    Batoo, Khalid Mujasam, E-mail: khalid.mujasam@gmail.com [King Saud University, King Abdullah Institute for Nanotechnology (Saudi Arabia); Mir, Feroz Ahmed [University of Kashmir, University Science Instrumentation Centre (India); Abd El-sadek, M.-S. [South Valley University, Nanomaterial Laboratory, Physics Department, Faculty of Science (Egypt); Shahabuddin, Md.; Ahmed, Niyaz [King Saud University, Department of Physics and Astronomy (Saudi Arabia)

    2013-11-15

    Nanoparticles of spinel ferrites of basic composition Ni{sub 1−x}Co{sub x}Fe{sub 2}O{sub 4} (0.0 ≤ x ≤ 0.05) were synthesized through modified co-precipitation method, and were characterized for structural, transport electrical and magnetic properties using XRD, HRTEM, FTIR, LCR meter and VSM techniques, respectively. XRD analysis showed that all the samples are single-phase cubic spinel in structure. The average crystallite sizes of the nanoparticles were found between 30 nm to 45 nm. Real and imaginary parts of the impedance (Z′ and Z″) suggested coexistence of two relaxation regimes: one was introduced by electrode polarization, while the other was attributed to the coeffect of grain and grain boundary effects. The dielectric constant of the samples was found very high, which showed non-Debye relaxation phenomena, while conductivity of the samples exhibited a two-segment behavior with frequency. The room temperature M–H curves suggested that the samples exhibit supermagnetism, and the saturation magnetization increases with increasing Co{sup 2+} ion substitution.

  12. A high-performance constant-temperature hot-wire anemometer

    Science.gov (United States)

    Watmuff, Jonathan H.

    1994-08-01

    A high-performance constant-temperature hot-wire anemometer has been designed based on a system theory analysis that can be extended to arbitrary order. A motivating factor behind the design was to achieve the highest possible frequency response while ensuring overall system stability. Based on these considerations, the design of the circuit and the selection of components is discussed in depth. Basic operating instructions are included in an operator's guide. The analysis is used to identify operating modes, observed in all anemometers, that are misleading in the sense that the operator can be deceived by interpreting an erroneous frequency response. Unlike other anemometers, this instrument provides front panel access to all the circuit parameters which affect system stability and frequency response. Instructions are given on how to identify and avoid these rather subtle and undesirable operating modes by appropriate adjustment of the controls. Details, such as fabrication drawings and a parts list, are provided to enable others to construct the instrument.

  13. A high-performance constant-temperature hot-wire anemometer

    Science.gov (United States)

    Watmuff, Jonathan H.

    1994-01-01

    A high-performance constant-temperature hot-wire anemometer has been designed based on a system theory analysis that can be extended to arbitrary order. A motivating factor behind the design was to achieve the highest possible frequency response while ensuring overall system stability. Based on these considerations, the design of the circuit and the selection of components is discussed in depth. Basic operating instructions are included in an operator's guide. The analysis is used to identify operating modes, observed in all anemometers, that are misleading in the sense that the operator can be deceived by interpreting an erroneous frequency response. Unlike other anemometers, this instrument provides front panel access to all the circuit parameters which affect system stability and frequency response. Instructions are given on how to identify and avoid these rather subtle and undesirable operating modes by appropriate adjustment of the controls. Details, such as fabrication drawings and a parts list, are provided to enable others to construct the instrument.

  14. A Calculation Approach to Elastic Constants of Crystallines at High Pressure and Finite Temperature

    Institute of Scientific and Technical Information of China (English)

    向士凯; 蔡灵仓; 张林; 经福谦

    2002-01-01

    Elastic constants of Na and Li metals are calculated successfully for temperatures up to 350K and pressures up to 30 GPa using a scheme without involving any adjustable parameter. Elastic constants are assumed to depend only on an effective pair potential that is only determined by the average interatomic distance. Temperature has an effect on elastic constants by way of charging the equilibrium. The elastic constants can be obtained by fitting the relationship between total energy and strain tensor using the new set of lattice parameters obtained by calculating displacement of atoms at the finite temperature and at a fixed pressure. The relationship between the effective pair potential and the interatomic distance is fitted by using a series of data of cohesive energy corresponding to lattice parameters.

  15. High-Precision Ephemerides of Planets—EPM and Determination of Some Astronomical Constants

    Science.gov (United States)

    Pitjeva, E. V.

    2005-05-01

    The latest version of the planetary part of the numerical ephemerides EPM (Ephemerides of Planets and the Moon) developed at the Institute of Applied Astronomy of the Russian Academy of Sciences is presented. The ephemerides of planets and the Moon were constructed by numerical integration in the post-Newtonian metric over a 140-year interval (from 1880 to 2020). The dynamical model of EPM2004 ephemerides includes the mutual perturbations from major planets and the Moon computed in terms of General Relativity with allowance for effects due to lunar physical libration, perturbations from 301 big asteroids, and dynamic perturbations due to the solar oblateness and the massive asteroid ring with uniform mass distribution in the plane of the ecliptic. The EPM2004 ephemerides resulted from a least-squares adjustment to more than 317000 position observations (1913 2003) of various types, including radiometric measurements of planets and spacecraft, CCD astrometric observations of the outer planets and their satellites, and meridian and photographic observations. The high-precision ephemerides constructed made it possible to determine, from modern radiometric measurements, a wide range of astrometric constants, including the astronomical unit AU = (149597870.6960 ± 0.0001) km, parameters of the rotation of Mars, the masses of the biggest asteroids, the solar quadrupole moment J 2 = (1.9 ± 0.3) × 10-7, and the parameters of the PPN formalism β and γ. Also given is a brief summary of the available state-of-the-art ephemerides with the same precision: various versions of EPM and DE ephemerides from the Jet Propulsion Laboratory (JPL) (USA) and the recent versions of these ephemerides—EPM2004 and DE410—are compared. EPM2004 ephemerides are available via FTP at ftp://qua-sar.ipa.nw.ru/incoming/EPM2004.

  16. A high-linearity digital-to-time converter technique: constant-slope charging

    NARCIS (Netherlands)

    Ru, Jiayoon Zhiyu; Palattella, Claudia; Geraedts, Paul; Klumperink, Eric; Nauta, Bram

    2015-01-01

    A digital-to-time converter (DTC) controls time delay by a digital code, which is useful, for example, in a sampling oscilloscope, fractional-N PLL, or time-interleaved ADC. This paper proposes constant-slope charging as a method to realize a DTC with intrinsically better integral non-linearity (INL

  17. Highly sensitive electrochemiluminescence displacement method for the study of DNA/small molecule binding interactions

    Energy Technology Data Exchange (ETDEWEB)

    Huang Rongfu; Wang Lirong [State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-environmental Sciences, Chinese Academy of Sciences, 18 Shuangqing Road, P.O. Box 2871, Beijing 100085 (China); Guo Lianghong, E-mail: LHGuo@rcees.ac.cn [State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-environmental Sciences, Chinese Academy of Sciences, 18 Shuangqing Road, P.O. Box 2871, Beijing 100085 (China)

    2010-08-31

    Non-covalent binding interactions of small molecules with DNA play important roles in regulating gene expression and gene function. In this work, a highly sensitive electrochemiluminescence (ECL) displacement method has been developed to investigate such interactions, particularly for weak DNA binders. This ECL method relies on a double-stranded DNA film deposited on an indium tin oxide electrode (ITO) surface by layer-by-layer self-assembly. A DNA intercalator, [Ru(bpy){sub 2}(dppz)]{sup 2+} (bpy = 2,2'-bipyridine, dppz = dipyrido[3,2-a:2'3'-c]phenazine), is employed as the ECL signal indicator. If a test compound competes with the indicator for the same binding sites in DNA, it would displace the indicator from the film and reduce ECL signal. The new method was validated by measuring five well-known DNA-binding organic molecules including quinacrine, H33258, thiazole orange, ethidium bromide and 4,6-diamidine-2-phenylindole dihydrochloride. Due to high ECL sensitivity, only 0.4 {mu}mol L{sup -1} [Ru(bpy){sub 2}(dppz)]{sup 2+} was needed in the ECL displacement measurement, which is about 75-fold less than the concentration in the voltammetric measurement. The lowered concentration permitted direct measurement of IC{sub 50} values of eight hydroxylated polycyclic aromatic hydrocarbons in their ECL displacement curves and subsequent calculation of their binding constants with DNA. The ECL displacement method is particularly useful for investigating weak DNA binders with limited aqueous solubility.

  18. Optical and electrical characterization of high resistivity semiconductors for constant-bias microbolometer devices

    Science.gov (United States)

    Saint John, David B.

    The commercial market for uncooled infrared imaging devices has expanded in the last several decades, following the declassification of pulse-biased microbolometer-based focal plane arrays (FPAs) using vanadium oxide as the sensing material. In addition to uncooled imaging platforms based on vanadium oxide, several constant-bias microbolometer FPAs have been developed using doped hydrogenated amorphous silicon (a-Si:H) as the active sensing material. While a-Si:H and the broader Si1-xGex:H system have been studied within the context of photovoltaic (PV) devices, only recently have these materials been studied with the purpose of qualifying and optimizing them for potential use in microbolometer applications, which demand thinner films deposited onto substrates different than those used in PV. The behavior of Ge:H is of particular interest for microbolometers due to its intrinsically low resistivity without the introduction of dopants, which alter the growth behavior and frustrate any attempt to address the merits of protocrystalline a-Ge:H. This work reports the optical, microstructural, and electrical characterization and qualification of a variety of Si:H, Si1-xGex:H, and Ge:H films deposited using a plasma enhanced chemical vapor deposition (PECVD) process, including a-Ge:H films which exhibit high TCR (4-6 -%/K) and low 1/f noise at resistivities of interest for microbolometers (4000 -- 6000 O cm). Thin film deposition has been performed simultaneously with real-time optical characterization of the growth evolution dynamics, providing measurement of optical properties and surface roughness evolutions relevant to controlling the growth process for deliberate variations in film microstructure. Infrared spectroscopic ellipsometry has been used to characterize the Si-H and Ge-H absorption modes allowing assessment of the hydrogen content and local bonding behavior in thinner films than measured traditionally. This method allows IR absorption analysis of hydrogen

  19. First example of a high-level correlated calculation of the indirect spin-spin coupling constants involving tellurium

    DEFF Research Database (Denmark)

    Rusakov, Yury Yu; Krivdin, Leonid B.; Østerstrøm, Freja From;

    2013-01-01

    This paper documents a very first example of a high-level correlated calculation of spin-spin coupling constants involving tellurium taking into account relativistic effects, vibrational corrections and solvent effects for the medium sized organotellurium molecules. The 125Te-1H spin-spin coupling...... of spin-spin coupling constants involving tellurium, was developed. The SOPPA methods show much better performance as compared to 15 those of DFT, if relativistic effects calculated within the ZORA scheme are taken into account. Vibrational and solvent corrections are next to negligible, while...

  20. High-level theoretical study of the reaction between hydroxyl and ammonia: Accurate rate constants from 200 to 2500 K

    Science.gov (United States)

    Nguyen, Thanh Lam; Stanton, John F.

    2017-10-01

    Hydrogen abstraction from NH3 by OH to produce H2O and NH2—an important reaction in combustion of NH3 fuel—was studied with a theoretical approach that combines high level quantum chemistry and advanced chemical kinetics methods. Thermal rate constants calculated from first principles agree well (within 5%-20%) with available experimental data over a temperature range that extends from 200 to 2500 K. Quantum mechanical tunneling effects were found to be important; they lead to a decided curvature and non-Arrhenius behavior for the rate constant.

  1. AB INITIO CALCULATIONS OF ELASTIC CONSTANTS OF BCC V-NB SYSTEM AT HIGH PRESSURES

    Energy Technology Data Exchange (ETDEWEB)

    Landa, A; Klepeis, J; Soderlind, P; Naumov, I; Velikokhatnyi, O; Vitos, L; Ruban, A

    2005-05-02

    First-principles total energy calculation based on the exact muffin-tin orbital and full potential linear muffin-tin orbital methods were used to calculate the equation of state and shear elastic constants of bcc V, Nb, and the V{sub 95}Nb{sub 05} disordered alloy as a function of pressure up to 6 Mbar. We found a mechanical instability in C{sub 44} and a corresponding softening in C at pressures {approx} 2 Mbar for V. Both shear elastic constants show softening at pressures {approx} 0.5 Mbar for Nb. Substitution of 5 at. % of V with Nb removes the instability of V with respect to trigonal distortions in the vicinity of 2 Mbar pressure, but still leaves the softening of C{sub 44} in this pressure region. We argue that the pressure induced shear instability (softening) of V (Nb) originates from the electronic system and can be explained by a combination of the Fermi surface nesting, electronic topological transition, and band Jahn-Teller effect.

  2. High-Fidelity DNA Sensing by Protein Binding Fluctuations

    CERN Document Server

    Tlusty, Tsvi; Libchaber, Albert; 10.1103/PhysRevLett.93.258103

    2010-01-01

    One of the major functions of RecA protein in the cell is to bind single-stranded DNA exposed upon damage, thereby triggering the SOS repair response.We present fluorescence anisotropy measurements at the binding onset, showing enhanced DNA length discrimination induced by adenosine triphosphate consumption. Our model explains the observed DNA length sensing as an outcome of out-of equilibrium binding fluctuations, reminiscent of microtubule dynamic instability. The cascade architecture of the binding fluctuations is a generalization of the kinetic proofreading mechanism. Enhancement of precision by an irreversible multistage pathway is a possible design principle in the noisy biological environment.

  3. New method evaluating currents keeping the voltage constant for fast and highly resolved measurement of Arrhenius relation and capacity fade

    Science.gov (United States)

    Lewerenz, Meinert; Käbitz, Stefan; Knips, Marcus; Münnix, Jens; Schmalstieg, Johannes; Warnecke, Alexander; Sauer, Dirk Uwe

    2017-06-01

    The evaluation of floating currents is a powerful method to characterize capacity fade induced by calendaric aging and enables a highly resolved representation of the Arrhenius relation. The test arrangement is simple and could constitute a cheap alternative to state-of-the-art calendaric aging tests including check-up tests. Therefore the currents to maintain a constant voltage are evaluated. This method is validated by analyzing nine cylindrical 8 Ah LiFePO4|Graphite battery cells during calendaric aging at 25 °C, 40 °C and 60 °C at 3.6 V (100% SOC). The 3.6 V are kept by applying constant voltage while the floating currents are logged. The floating currents correlate with the rate of capacity loss measured during capacity tests. The floating currents reveal to be rather constant at 25 °C, linearly increasing at 40 °C and decreasing from a higher level at 60 °C. Additional tests with three test cells, with the temperature rising from 40 to 60 °C in steps of 5 K, exhibit non-constant currents starting from 50 °C on with high variations amongst the tested cells. Once stored above 50 °C, the cells exhibit increased floating currents compared to the measurement at the same temperature before exceeding 50 °C.

  4. One-loop omega-potential of charged massive fields in a constant homogeneous magnetic field at high temperatures

    CERN Document Server

    Kalinichenko, I S

    2016-01-01

    The explicit expressions for the high-temperature expansions of the one-loop corrections to the omega-potential coming from the charged scalar and Dirac particles and, separately, from antiparticles in a constant homogeneous magnetic field are derived. The explicit expressions for the non-perturbative corrections to the effective action at finite temperature and density are obtained. The thermodynamic properties of a gas of charged scalars in a constant homogeneous magnetic field are analyzed in the one-loop approximation. It turns out that, in this approximation, the system suffers the first order phase transition from the diamagnetic to the superconducting state at sufficiently high densities. The improvement of the one-loop result by summing the ring diagrams is investigated. This improvement leads to a drastic change of the thermodynamic properties of the system. The gas of charged scalars passes to the ferromagnetic state in place of the superconducting one at high densities and sufficiently low temperat...

  5. Testing the running of the cosmological constant with Type Ia Supernovae at high z

    CERN Document Server

    Espana-Bonet, C; Shapiro, I L; Solà, J; Espana-Bonet, Cristina; Ruiz-Lapuente, Pilar; Shapiro, Ilya L.; Sola, Joan

    2004-01-01

    Within the Quantum Field Theory context the idea of a "cosmological constant" (CC) evolving with time looks quite natural as it just reflects the change of the vacuum energy with the typical energy of the universe. In the particular frame of Ref.[30], a "running CC" at low energies may arise from generic quantum effects near the Planck scale, M_P, provided there is a smooth decoupling of all massive particles below M_P. In this work we further develop the cosmological consequences of a "running CC" by addressing the accelerated evolution of the universe within that model. The rate of change of the CC stays slow, without fine-tuning, and is comparable to H^2 M_P^2. It can be described by a single parameter, \

  6. High magnetic exchange coupling constants: a density functional theory based study of substituted Schlenk diradicals.

    Science.gov (United States)

    Latif, Iqbal A; Hansda, Shekhar; Datta, Sambhu N

    2012-08-23

    The Schlenk diradical has been known since 1915. After a detailed experimental work by Rajca, its magnetic nature has remained more or less unexplored. We have investigated by quantum chemical calculations the nature of magnetic coupling in 11 substituted Schlenk diradicals. Substitution has been considered at the fifth carbon atom of the meta-phenylene moiety. The UB3LYP method has been used to study 12 diradicals including the original one. The 6-311G(d,p) basis set has been employed for optimization of molecular geometry in both singlet and triplet states for each species. The singlet optimization has led to the optimization of the broken-symmetry structure for 10 species including the unsubstituted one. This development makes it possible to carry out further broken symmetry calculations in two ways. The triplet calculation has been done using 6-311++G(d,p) basis set and the optimized triplet geometry in both procedures. The broken symmetry calculations have used the optimized geometries of either the triplet states or the broken symmetry solutions. The first method leads to the prediction of electron paramagnetic resonance (EPR) compatible magnetic exchange coupling constant (J) in the range 517-617 cm(-1). A direct optimization of the broken symmetry geometry gives rise to a lower estimate of J, in the range of 411-525 cm(-1) and compatible with macroscopic Curie studies. The calculated J for the unsubstituted Schlenk diradical is 512 cm(-1) that can be compared with 455 cm(-1) estimated by Rajca. In both cases, introduction of groups with +M and +I effects (Ingold's notation) decreases the J value from that for the unsubstituted Schlenk diradical while -I and -M groups at the same position increases J. These trends have been explained in terms of Hammett constants, atomic spin densities, and dihedral angles.

  7. Single-Crystal Elastic Constants of Yttria (Y2O3) Measured to High Temperatures

    Science.gov (United States)

    Sayir, Ali; Palko, James W.; Kriven, Waltraud M.; Sinogeikin, Sergey V.; Bass, Jay D.

    2001-01-01

    Yttria, or yttrium sesquioxide (Y2O3), has been considered for use in nuclear applications and has gained interest relatively recently for use in infrared optics. Single crystals of yttria have been grown successfully at the NASA Glenn Research Center using a laser-heated float zone technique in a fiber and rod. Such samples allow measurement of the single-crystal elastic properties, and these measurements provide useful property data for the design of components using single crystals. They also yield information as to what degree the elastic properties of yttria ceramics are a result of the intrinsic properties of the yttria crystal in comparison to characteristics that may depend on processing, such as microstructure and intergranular phases, which are common in sintered yttria. The single-crystal elastic moduli are valuable for designing such optical components. In particular, the temperature derivatives of elastic moduli allow the dimensional changes due to heating under physical constraints, as well as acoustic excitation, to be determined. The single-crystal elastic moduli of yttria were measured by Brillouin spectroscopy up to 1200 C. The room-temperature values obtained were C(sub 11) = 223.6 + 0.6 GPa, C(sub 44) = 74.6 + 0.5 GPa, and C(sub 12) = 112.4 + 1.0 GPa. The resulting bulk and (Voigt-Reuss-Hill) shear moduli were K = 149.5 + 1.0 GPa and G(sub VRH) = 66.3 + 0.8 GPa, respectively. Linear least-squares regressions to the variation of bulk and shear moduli with temperature resulted in derivatives of dK/dT = -17 + 2 MPa/C and dG(sub VRH)/dT = -8 + 2 MPa/ C. Elastic anisotropy was found to remain essentially constant over the temperature range studied.

  8. Complementary DNA display selection of high-affinity peptides binding the vacuolating toxin (VacA) of Helicobacter pylori.

    Science.gov (United States)

    Hayakawa, Yumiko; Matsuno, Mitsuhiro; Tanaka, Makoto; Wada, Akihiro; Kitamura, Koichiro; Takei, Osamu; Sasaki, Ryuzo; Mizukami, Tamio; Hasegawa, Makoto

    2015-09-01

    Artificial peptides designed for molecular recognition of a bacterial toxin have been developed. Vacuolating cytotoxin A protein (VacA) is a major virulence factor of Helicobacter pylori, a gram-negative microaerophilic bacterium inhabiting the upper gastrointestinal tract, particularly the stomach. This study attempted to identify specific peptide sequences with high affinity for VacA using systematic directed evolution in vitro, a cDNA display method. A surface plasmon resonance-based biosensor and fluorescence correlation spectroscopy to examine binding of peptides with VacA identified a peptide (GRVNQRL) with high affinity. Cyclization of the peptide by attaching cysteine residues to both termini improved its binding affinity to VacA, with a dissociation constant (Kd ) of 58 nm. This study describes a new strategy for the development of artificial functional peptides, which are promising materials in biochemical analyses and medical applications.

  9. Preparation of high dielectric constant thin films of CaCu3Ti4O12 by sol–gel

    Indian Academy of Sciences (India)

    Deepam Maurya; Devendra P Singh; D C Agrawal; Y N Mohapatra

    2008-02-01

    Preparation of sol–gel derived CaCu3Ti4O12 (CCTO) thin films using two different sols and their characterization including their dielectric response are reported. The properties of CCTO films depend heavily on solvents used to prepare the sols. Dielectric constant as high as ∼900 at 100 kHz could be obtained when acetic acid was used to prepare the sol; in contrast, use of hexanoic acid in the sol yielded films with a much lower dielectric constant. The variation in grain and grain boundary conductivities with temperature has been measured. Activation energies of 0.08 eV and 0.68 eV have been found for grain and grain boundary conduction, respectively.

  10. Cosmochemistry, cosmology and fundamental constants: High-resolution spectroscopy of damped Lyman-alpha systems

    CERN Document Server

    Quast, R; Smette, A; Garcet, O; Ledoux, C; López, S; Wisotzki, L

    2004-01-01

    Spectroscopy of QSO absorption lines provides essential observational input for the study of nucleosynthesis and chemical evolution of galaxies at high redshift. But new observations may indicate that present chemical abundance data are biased due to deficient spectral resolution and unknown selection effects: Recent high-resolution spectra reveal the hitherto unperceived chemical nonuniformity of molecular hydrogen-bearing damped Lyman-alpha (DLA) systems, and the novel H/ESO DLA survey produces compelling evidence for faint QSOs being attenuated by dust. We present a revised analysis of the molecular hydrogen-bearing DLA complex toward HE 0515-4414 showing nonuniform differential depletion of chemical elements onto dust grains, and introduce to the H/ESO DLA survey and its implications. Conclusively, we aim at starting an unbiased chemical abundance database established on high-resolution spectroscopic observations. New data to probe the temperature-redshift relation predicted by standard cosmology and to t...

  11. Design and Construction of Low Cost High Voltage dc Power Supply for Constant Power Operation

    Science.gov (United States)

    Kumar, N. S.; Jayasankar, V.

    2013-06-01

    Pulsed load applications like laser based systems need high voltage dc power supplies with better regulation characteristics. This paper presents the design, construction and testing of dc power supply with 1 kV output at 300 W power level. The designed converter has half bridge switched mode power supply (SMPS) configuration with 20 kHz switching. The paper covers the design of half bridge inverter, closed loop control, High frequency transformer and other related electronics. The designed power supply incorporates a low cost OPAMP based feedback controller which is designed using small signal modelling of the converter. The designed converter was constructed and found to work satisfactorily as per the specifications.

  12. A Comparison of Constant Time Delay Instruction with High and Low Treatment Integrity

    Science.gov (United States)

    Tekin Iftar, Elif; Kurt, Onur; Cetin, Ozlem

    2011-01-01

    Time delay (TD) procedure is an effective procedure in teaching various skills to children with developmental disabilities. Moreover, research has shown that it is used with high treatment integrity (HTI). However, there are several barriers which may prevent delivery instruction with HTI. Therefore, this study was designed to compare the…

  13. Hydrogenlike highly charged ions for tests of the time independence of fundamental constants.

    Science.gov (United States)

    Schiller, S

    2007-05-04

    Hyperfine transitions in the electronic ground state of cold, trapped hydrogenlike highly charged ions have attractive features for use as frequency standards because the majority of systematic frequency shifts are smaller by orders of magnitude compared to many microwave and optical frequency standards. Frequency measurements of these transitions hold promise for significantly improved laboratory tests of local position invariance of the electron and quark masses.

  14. Highly accurate analytical energy of a two-dimensional exciton in a constant magnetic field

    Energy Technology Data Exchange (ETDEWEB)

    Hoang, Ngoc-Tram D. [Department of Physics, Ho Chi Minh City University of Pedagogy 280, An Duong Vuong Street, District 5, Ho Chi Minh City (Viet Nam); Nguyen, Duy-Anh P. [Department of Natural Science, Thu Dau Mot University, 6, Tran Van On Street, Thu Dau Mot City, Binh Duong Province (Viet Nam); Hoang, Van-Hung [Department of Physics, Ho Chi Minh City University of Pedagogy 280, An Duong Vuong Street, District 5, Ho Chi Minh City (Viet Nam); Le, Van-Hoang, E-mail: levanhoang@tdt.edu.vn [Atomic Molecular and Optical Physics Research Group, Ton Duc Thang University, 19 Nguyen Huu Tho Street, Tan Phong Ward, District 7, Ho Chi Minh City (Viet Nam); Faculty of Applied Sciences, Ton Duc Thang University, 19 Nguyen Huu Tho Street, Tan Phong Ward, District 7, Ho Chi Minh City (Viet Nam)

    2016-08-15

    Explicit expressions are given for analytically describing the dependence of the energy of a two-dimensional exciton on magnetic field intensity. These expressions are highly accurate with the precision of up to three decimal places for the whole range of the magnetic field intensity. The results are shown for the ground state and some excited states; moreover, we have all formulae to obtain similar expressions of any excited state. Analysis of numerical results shows that the precision of three decimal places is maintained for the excited states with the principal quantum number of up to n=100.

  15. Effect Of A Video-Based Laboratory On The High School Pupils’ Understanding Of Constant Speed Motion

    Directory of Open Access Journals (Sweden)

    Louis Trudel, Abdeljalil Métioui

    2012-05-01

    Full Text Available Among the physical phenomena studied in high school, the kinematical concepts are important because they constitute a precondition for the study of subsequent concepts of mechanics. Our research aims at studying the effect of a computer-assisted scientific investigation on high school pupils’ understanding of the constant speed motion. Experimentation took place in a high school physics classroom. A repeated measures analysis of variance shows that, during the implementation of this strategy, the pupils’ understanding of kinematical concepts increased in a significant way. In conclusion, we specify advantages and limits of the study and give future research directions concerning the design of a computer-assisted laboratory in high school physics.

  16. High-Order Elastic Constants and Anharmonic Properties of NaBH4: First-Principles Calculations

    Institute of Scientific and Technical Information of China (English)

    ZHANG Xiao-Dong; JIANG Zhen-Yi; ZHOU Bo; HOU Zhu-Feng; HOU Yu-Qing

    2011-01-01

    We present theoretical studies for second- and third-order elastic constants in NaBH4 based on ab initio calculations. Our calculated second-order elastic constants agree well with available experimental results. The anharmonic properties of NaBH4,such as pressure derivative of the second-order elastic constants and the Grüneisen constants for long-wavelength acoustic modeγ(q,j),are characterized using the third-order elastic constants.

  17. Highly Accurate Quartic Force Fields, Vibrational Frequencies, and Spectroscopic Constants for Cyclic and Linear C3H3(+)

    Science.gov (United States)

    Huang, Xinchuan; Taylor, Peter R.; Lee, Timothy J.

    2011-01-01

    High levels of theory have been used to compute quartic force fields (QFFs) for the cyclic and linear forms of the C H + molecular cation, referred to as c-C H + and I-C H +. Specifically the 33 3333 singles and doubles coupled-cluster method that includes a perturbational estimate of connected triple excitations, CCSD(T), has been used in conjunction with extrapolation to the one-particle basis set limit and corrections for scalar relativity and core correlation have been included. The QFFs have been used to compute highly accurate fundamental vibrational frequencies and other spectroscopic constants using both vibrational 2nd-order perturbation theory and variational methods to solve the nuclear Schroedinger equation. Agreement between our best computed fundamental vibrational frequencies and recent infrared photodissociation experiments is reasonable for most bands, but there are a few exceptions. Possible sources for the discrepancies are discussed. We determine the energy difference between the cyclic and linear forms of C H +, 33 obtaining 27.9 kcal/mol at 0 K, which should be the most reliable available. It is expected that the fundamental vibrational frequencies and spectroscopic constants presented here for c-C H + 33 and I-C H + are the most reliable available for the free gas-phase species and it is hoped that 33 these will be useful in the assignment of future high-resolution laboratory experiments or astronomical observations.

  18. A reassessment of the association between azulene and [60]fullerene. Possible pitfalls in the determination of binding constants through fluorescence spectroscopy.

    Science.gov (United States)

    Stella, Lorenzo; Capodilupo, Agostina L; Bietti, Massimo

    2008-10-21

    We show here that the recently reported surprisingly large association constant (K = 7.6 x 10(4) M(-1)) between azulene and [60]fullerene is due to experimental artifacts, pointing out potential errors in the characterization of association equilibria by fluorescence spectroscopy, and suggesting the best experimental practices.

  19. Will the western Pacific subtropical high constantly intensify in the future?

    Science.gov (United States)

    Huang, Yanyan; Li, Xiaofan; Wang, Huijun

    2016-07-01

    The western Pacific subtropical high (WPSH) features lower-level southerlies or southwesterlies at its western and southern edges that transport amount of water vapor into East Asia, and it exerts a large influence on the East Asian summer climate. This paper evaluates the historical (1950-2005) spatial distribution and variability in the summer WPSH at 850 hPa using 28 general circulation models from the Coupled Model Intercomparison Project Phase 5 (CMIP5) relative to the NCEP-NCAR reanalysis data. To avoid the artificial influence caused by global warming, this study primarily investigates the 850-hPa eddy geopotential height and the horizontal winds. The results show that most of the CMIP5 models reliably reproduce the geographical distribution and spatial variability in the WPSH. Four models (bcc-csm1-1, CESM1-CAM5, GFDL-ESM2G and inmcm4) generally perform well in simulating the eastward-recessed interdecadal variation in the WPSH during 1979-2005 relative to 1950-1978, with a significant cyclone anomaly appearing over the western Pacific and a decreasing trend in the WPSH index. Based on these four models, a multi-model ensemble projects a weaker WPSH during 2026-2070 relative to 2010-2025 and 2071-2100 under the representative concentration pathway 8.5 scenario.

  20. A High-Affinity Binding Site for the AVR9 Peptide Elicitor of Cladosporium fulvum Is Present on Plasma Membranes of Tomato and Other Solanaceous Plants.

    Science.gov (United States)

    Kooman-Gersmann, M.; Honee, G.; Bonnema, G.; De Wit, PJGM.

    1996-05-01

    The race-specific Cladosporium fulvum peptide elicitor AVR9, which specifically induces a hypersensitive response in tomato genotypes carrying the Cf-9 resistance gene, was labeled with iodine-125 at the N-terminal tyrosine residue and used in binding studies. 125I-AVR9 showed specific, saturable, and reversible binding to plasma membranes isolated from leaves of tomato cultivar Moneymaker without Cf resistance genes (MM-Cf0) or from a near-isogenic genotype with the Cf-9 resistance gene (MM-Cf9). The dissociation constant was found to be 0.07 nM, and the receptor concentration was 0.8 pmol/mg microsomal protein. Binding was highly influenced by pH and the ionic strength of the binding buffer and by temperature, indicating the involvement of both electrostatic and hydrophobic interactions. Binding kinetics and binding capacity were similar for membranes of the MM-Cf0 and MM-Cf9 genotypes. In all solanaceous plant species tested, an AVR9 binding site was present, whereas in the nonsolanaceous species that were analyzed, such a binding site could not be identified. The ability of membranes isolated from different solanaceous plant species to bind AVR9 seems to correlate with the presence of members of the Cf-9 gene family, but whether this correlation is functional remains to be determined.

  1. Entrapment of alpha1-acid glycoprotein in high-performance affinity columns for drug-protein binding studies.

    Science.gov (United States)

    Bi, Cong; Jackson, Abby; Vargas-Badilla, John; Li, Rong; Rada, Giana; Anguizola, Jeanethe; Pfaunmiller, Erika; Hage, David S

    2016-05-15

    A slurry-based method was developed for the entrapment of alpha1-acid glycoprotein (AGP) for use in high-performance affinity chromatography to study drug interactions with this serum protein. Entrapment was achieved based on the physical containment of AGP in hydrazide-activated porous silica supports and by using mildly oxidized glycogen as a capping agent. The conditions needed for this process were examined and optimized. When this type of AGP column was used in binding studies, the association equilibrium constant (Ka) measured by frontal analysis at pH 7.4 and 37°C for carbamazepine with AGP was found to be 1.0 (±0.5)×10(5)M(-1), which agreed with a previously reported value of 1.0 (±0.1)×10(5)M(-1). Binding studies based on zonal elution were conducted for several other drugs with such columns, giving equilibrium constants that were consistent with literature values. An entrapped AGP column was also used in combination with a column containing entrapped HSA in a screening assay format to compare the binding of various drugs to AGP and HSA. These results also agreed with previous data that have been reported in literature for both of these proteins. The same entrapment method could be extended to other proteins and to the investigation of additional types of drug-protein interactions. Potential applications include the rapid quantitative analysis of biological interactions and the high-throughput screening of drug candidates for their binding to a given protein.

  2. Effects of endurance training on reduction of plasma glucose during high intensity constant and incremental speed tests in Wistar rats.

    Science.gov (United States)

    Abreu, P; Vitzel, K F; Monteiro, I C C R; Lima, T I; Queiroz, A N; Leal-Cardoso, J H; Hirabara, S M; Ceccatto, V M

    2016-10-24

    The aim of this research was to investigate the effects of endurance training on reduction of plasma glucose during high intensity constant and incremental speed tests in Wistar rats. We hypothesized that plasma glucose might be decreased in the exercised group during heavy (more intense) exercise. Twenty-four 10-week-old male Wistar rats were randomly assigned to sedentary and exercised groups. The prescription of endurance exercise training intensity was determined as 60% of the maximum intensity reached at the incremental speed test. The animals were trained by running on a motorized treadmill, five days/week for a total period of 67 weeks. Plasma glucose during the constant speed test in the exercised group at 20 m/min was reduced at the 14th, 21st and 28th min compared to the sedentary group, as well at 25 m/min at the 21st and 28th min. Plasma glucose during the incremental speed test was decreased in the exercised group at the moment of exhaustion (48th min) compared to the sedentary group (27th min). Endurance training positively modulates the mitochondrial activity and capacity of substrate oxidation in muscle and liver. Thus, in contrast to other studies on high load of exercise, the effects of endurance training on the decrease of plasma glucose during constant and incremental speed tests was significantly higher in exercised than in sedentary rats and associated with improved muscle and hepatic oxidative capacity, constituting an important non-pharmacological intervention tool for the prevention of insulin resistance, including type 2 diabetes mellitus.

  3. Effects of endurance training on reduction of plasma glucose during high intensity constant and incremental speed tests in Wistar rats

    Directory of Open Access Journals (Sweden)

    P. Abreu

    Full Text Available The aim of this research was to investigate the effects of endurance training on reduction of plasma glucose during high intensity constant and incremental speed tests in Wistar rats. We hypothesized that plasma glucose might be decreased in the exercised group during heavy (more intense exercise. Twenty-four 10-week-old male Wistar rats were randomly assigned to sedentary and exercised groups. The prescription of endurance exercise training intensity was determined as 60% of the maximum intensity reached at the incremental speed test. The animals were trained by running on a motorized treadmill, five days/week for a total period of 67 weeks. Plasma glucose during the constant speed test in the exercised group at 20 m/min was reduced at the 14th, 21st and 28th min compared to the sedentary group, as well at 25 m/min at the 21st and 28th min. Plasma glucose during the incremental speed test was decreased in the exercised group at the moment of exhaustion (48th min compared to the sedentary group (27th min. Endurance training positively modulates the mitochondrial activity and capacity of substrate oxidation in muscle and liver. Thus, in contrast to other studies on high load of exercise, the effects of endurance training on the decrease of plasma glucose during constant and incremental speed tests was significantly higher in exercised than in sedentary rats and associated with improved muscle and hepatic oxidative capacity, constituting an important non-pharmacological intervention tool for the prevention of insulin resistance, including type 2 diabetes mellitus.

  4. High molecular weight kininogen binds to laminin--characterization and kinetic analysis

    DEFF Research Database (Denmark)

    Schousboe, Inger; Nystrøm, Birthe

    2009-01-01

    proteins laminin, but not to vitronectin or fibronectin coated on microtiter plates. The binding to laminin was Zn2+ independent. However, at low but not at high concentrations of albumin, Zn2+ increased the affinity for the binding by abolishing an inhibitory effect of Ca2+. Recombinant human kininostatin...... encompassing the amino acid sequence, Arg439-Ser532 but not the endothelial cell binding peptide sequence (His479-His498; HKH20) within kininostatin inhibited the binding of HKa to laminin. This established that the amino acid sequence Arg439-Lys478 in domain 5 of HK is of importance for its binding to laminin....... Extensive proteolytic cleavage of HK and HKa with kallikrein abolished the binding to laminin, releasing a 12 kDa anti-kininostatin reacting peptide. On the basis of these results, we propose that the binding of HK to laminin is a primary event, which secures proper localization of the cleavage products...

  5. High-affinity binding of (/sup 3/H)acetylcholine to muscarinic cholinergic receptors

    Energy Technology Data Exchange (ETDEWEB)

    Kellar, K.J.; Martino, A.M.; Hall, D.P. Jr.; Schwartz, R.D.; Taylor, R.L.

    1985-06-01

    High-affinity binding of (/sup 3/H)acetylcholine to muscarinic cholinergic sites in rat CNS and peripheral tissues was measured in the presence of cytisin, which occupies nicotinic cholinergic receptors. The muscarinic sites were characterized with regard to binding kinetics, pharmacology, anatomical distribution, and regulation by guanyl nucleotides. These binding sites have characteristics of high-affinity muscarinic cholinergic receptors with a Kd of approximately 30 nM. Most of the muscarinic agonist and antagonist drugs tested have high affinity for the (/sup 3/H)acetylcholine binding site, but pirenzepine, an antagonist which is selective for M-1 receptors, has relatively low affinity. The ratio of high-affinity (/sup 3/H)acetylcholine binding sites to total muscarinic binding sites labeled by (/sup 3/H)quinuclidinyl benzilate varies from 9 to 90% in different tissues, with the highest ratios in the pons, medulla, and heart atrium. In the presence of guanyl nucleotides, (/sup 3/H) acetylcholine binding is decreased, but the extent of decrease varies from 40 to 90% in different tissues, with the largest decreases being found in the pons, medulla, cerebellum, and heart atrium. The results indicate that (/sup 3/H)acetylcholine binds to high-affinity M-1 and M-2 muscarinic receptors, and they suggest that most M-2 sites have high affinity for acetylcholine but that only a small fraction of M-1 sites have such high affinity.

  6. Structure and performance of dielectric films based on self-assembled nanocrystals with a high dielectric constant.

    Science.gov (United States)

    Huang, Limin; Liu, Shuangyi; Van Tassell, Barry J; Liu, Xiaohua; Byro, Andrew; Zhang, Henan; Leland, Eli S; Akins, Daniel L; Steingart, Daniel A; Li, Jackie; O'Brien, Stephen

    2013-10-18

    Self-assembled films built from nanoparticles with a high dielectric constant are attractive as a foundation for new dielectric media with increased efficiency and range of operation, due to the ability to exploit nanofabrication techniques and emergent electrical properties originating from the nanoscale. However, because the building block is a discrete one-dimensional unit, it becomes a challenge to capture potential enhancements in dielectric performance in two or three dimensions, frequently due to surface effects or the presence of discontinuities. This is a recurring theme in nanoparticle film technology when applied to the realm of thin film semiconductor and device electronics. We present the use of chemically synthesized (Ba,Sr)TiO3 nanocrystals, and a novel deposition-polymerization technique, as a means to fabricate the dielectric layer. The effective dielectric constant of the film is tunable according to nanoparticle size, and effective film dielectric constants of up to 34 are enabled. Wide area and multilayer dielectrics of up to 8 cm(2) and 190 nF are reported, for which the building block is an 8 nm nanocrystal. We describe models for assessing dielectric performance, and distinct methods for improving the dielectric constant of a nanocrystal thin film. The approach relies on evaporatively driven assembly of perovskite nanocrystals with uniform size distributions in a tunable 7-30 nm size range, coupled with the use of low molecular weight monomer/polymer precursor chemistry that can infiltrate the porous nanocrystal thin film network post assembly. The intercrystal void space (low k dielectric volume fraction) is minimized, while simultaneously promoting intercrystal connectivity and maximizing volume fraction of the high k dielectric component. Furfuryl alcohol, which has good affinity to the surface of (Ba,Sr)TiO3 nanocrystals and miscibility with a range of solvents, is demonstrated to be ideal for the production of nanocomposites. The

  7. High affinity binding of (/sup 3/H)cocaine to rat liver microsomes

    Energy Technology Data Exchange (ETDEWEB)

    El-Maghrabi, E.A.; Calligaro, D.O.; Eldefrawi, M.E.

    1988-01-01

    )/sup 3/H)cocaine bound reversible, with high affinity and stereospecificity to rat liver microsomes. Little binding was detected in the lysosomal, mitochondrial and nuclear fractions. The binding kinetics were slow and the kinetically calculated K/sub D/ was 2 nM. Induction of mixed function oxidases by phenobarbital did not produce significant change in (/sup 3/H)cocaine binding. On the other hand, chronic administration of cocaine reduced (/sup 3/H)cocaine binding drastically. Neither treatment affected the affinity of the liver binding protein for cocaine. Microsomes from mouse and human livers had less cocaine-binding protein and lower affinity for cocaine than those from rat liver. Binding of (/sup 3/H)cocaine to rat liver microsomes was insensitive to monovalent cations and > 10 fold less sensitive to biogenic amines than the cocaine receptor in rat striatum. However, the liver protein had higher affinity for cocaine and metabolites except for norcocaine. Amine uptake inhibitors displaced (/sup 3/H)cocaine binding to liver with a different rank order of potency than their displacement of (/sup 3/H)cocaine binding to striatum. This high affinity (/sup 3/H)cocaine binding protein in liver is not likely to be monooxygenase, but may have a role in cocaine-induced hepatotoxicity

  8. MATHEMATICAL CONSTANTS.

    Energy Technology Data Exchange (ETDEWEB)

    Robinson, H.P.; Potter, Elinor

    1971-03-01

    This collection of mathematical data consists of two tables of decimal constants arranged according to size rather than function, a third table of integers from 1 to 1000, giving some of their properties, and a fourth table listing some infinite series arranged according to increasing size of the coefficients of the terms. The decimal values of Tables I and II are given to 20 D.

  9. Hemaka's constant

    CERN Document Server

    Sparavigna, Amelia Carolina

    2012-01-01

    As proposed in a previous paper, the decorations of ancient objects can provide some information on the approximate evaluations of constant {\\pi}, the ratio of circumference to diameter. Here we discuss some disks found in the tomb of Hemaka, the chancellor of a king of the First Dynasty of Egypt, about 3000 BC. The discussion is based on measurements of the dimensionless ratio of lengths.

  10. Fault strength evolution during high velocity friction experiments with slip-pulse and constant-velocity loading

    Science.gov (United States)

    Liao, Z.; Chang, J. C.; Reches, Z.

    2014-12-01

    Seismic analyses show that slip during large earthquakes evolves in a slip-pulse mode that is characterized by abrupt, intense acceleration followed by moderate deceleration. We experimentally analyze the friction evolution under slip-pulse proxy of a large earthquake, and compare it with the evolution at loading modes of constant-velocity and changing-velocity. We present a series of 42 experiments conducted on granite samples sheared in a high-velocity rotary apparatus. The experiments were conducted on room-dry, solid granite samples at slip-velocities of 0.0006-1 m/s, and normal stress of 1-11.5 MPa. The constitutive relations are presented with respect to mechanical power-density: PD= [shear stress * slip velocity], with units of power per area (MW/m^2). The experimental constitutive relations strongly depend on the loading mode. Constant velocity mode displays initial weakening with increasing PD that is followed by strengthening for PD = 0.02-0.5 MW/m^2, and abrupt weakening at PD > 0.5 MW/m^2. Changing-velocity modes display gentle strengthening for PD < 0.2 MW/m^2 that is followed by abrupt weakening as PD reaches 0.7-0.8 MW/m^2. Beyond this level of power-density, the two loading modes diverge: in changing-velocity of quake-mode the experimental fault continues to weaken with friction coefficient approaching 0.2, whereas in changing-velocity of ramp-mode the fault strengthens with friction coefficient approaching 1.0. The analysis demonstrates that (1) the strength evolution and constitutive parameters of the granite fault strongly depend on the loading mode, and (2) the slip-pulse mode is energy efficient relatively to the constant-velocity mode as manifested by faster, more intense weakening and 50-90% lower energy dissipation. The results suggest that the frictional strength determined in slip-pulse experiments, is more relevant to simulations of earthquake rupture than frictional strength determined in constant-velocity experiments.Figure 1. Friction

  11. A High-Performance Control Method of Constant V/f-Controlled Induction Motor Drives for Electric Vehicles

    Directory of Open Access Journals (Sweden)

    Long Chen

    2014-01-01

    Full Text Available A three-phase induction motor used as a propulsion system for the electric vehicle (EV is a nonlinear, multi-input multi-output, and strong coupling system. For such a complicated model system with unmeasured and unavoidable disturbances, as well as parameter variations, the conventional vector control method cannot meet the demands of high-performance control. Therefore, a novel control strategy named least squares support vector machines (LSSVM inverse control is presented in the paper. Invertibility of the induction motor in the constant V/f control mode is proved to confirm its feasibility. The LSSVM inverse is composed of an LSSVM approximating the nonlinear mapping of the induction motor and two integrators. The inverse model of the constant V/f-controlled induction motor drive is obtained by using LSSVM, and then the optimal parameters of LSSVM are determined automatically by applying a modified particle swarm optimization (MPSO. Cascading the LSSVM inverse with the induction motor drive system, the pseudolinear system can be obtained. Thus, it is easy to design the closed-loop linear regulator. The simulation results verify the effectiveness of the proposed method.

  12. Alignment-free ultra-high-throughput comparison of druggable protein-ligand binding sites.

    Science.gov (United States)

    Weill, Nathanaël; Rognan, Didier

    2010-01-01

    Inferring the biological function of a protein from its three-dimensional structure as well as explaining why a drug may bind to various targets is of crucial importance to modern drug discovery. Here we present a generic 4833-integer vector describing druggable protein-ligand binding sites that can be applied to any protein and any binding cavity. The fingerprint registers counts of pharmacophoric triplets from the Calpha atomic coordinates of binding-site-lining residues. Starting from a customized data set of diverse protein-ligand binding site pairs, the most appropriate metric and a similarity threshold could be defined for similar binding sites. The method (FuzCav) has been used in various scenarios: (i) screening a collection of 6000 binding sites for similarity to different queries; (ii) classifying protein families (serine endopeptidases, protein kinases) by binding site diversity; (iii) discriminating adenine-binding cavities from decoys. The fingerprint generation and comparison supports ultra-high throughput (ca. 1000 measures/s), does not require prior alignment of protein binding sites, and is able to detect local similarity among subpockets. It is thus particularly well suited to the functional annotation of novel genomic structures with low sequence identity to known X-ray templates.

  13. Fc-Binding Ligands of Immunoglobulin G: An Overview of High Affinity Proteins and Peptides

    Directory of Open Access Journals (Sweden)

    Weonu Choe

    2016-12-01

    Full Text Available The rapidly increasing application of antibodies has inspired the development of several novel methods to isolate and target antibodies using smart biomaterials that mimic the binding of Fc-receptors to antibodies. The Fc-binding domain of antibodies is the primary binding site for e.g., effector proteins and secondary antibodies, whereas antigens bind to the Fab region. Protein A, G, and L, surface proteins expressed by pathogenic bacteria, are well known to bind immunoglobulin and have been widely exploited in antibody purification strategies. Several difficulties are encountered when bacterial proteins are used in antibody research and application. One of the major obstacles hampering the use of bacterial proteins is sample contamination with trace amounts of these proteins, which can invoke an immune response in the host. Many research groups actively develop synthetic ligands that are able to selectively and strongly bind to antibodies. Among the reported ligands, peptides that bind to the Fc-domain of antibodies are attractive tools in antibody research. Besides their use as high affinity ligands in antibody purification chromatography, Fc-binding peptides are applied e.g., to localize antibodies on nanomaterials and to increase the half-life of proteins in serum. In this review, recent developments of Fc-binding peptides are presented and their binding characteristics and diverse applications are discussed.

  14. New high-pressure phase of MgH2: An ab initio constant-pressure study

    Science.gov (United States)

    Durandurdu, Murat

    2014-02-01

    The stability of magnesium hydride (MgH2) at high pressure is studied using a constant-pressure ab initio technique. Two phase transformations are successfully observed through the simulations. The rutile structure undergoes a phase transformation into a CaCl2-type phase. Further increase in pressure results into a first-order phase transition into an orthorhombic state within Pbcm symmetry. This phase can be considered as a distorted CaF2-type crystal and does not correspond to the previously proposed MgH2 phases. The transformation mechanism of the CaCl2-Pbcm phase change at the atomistic level is successfully characterized and it is found that the CaCl2-to-Pbcm phase change proceeds via an ideal CaF2-type intermediate phase. These phase transformations are also analyzed using total energy-volume calculations.

  15. Decavanadate binding to a high affinity site near the myosin catalytic centre inhibits F-actin-stimulated myosin ATPase activity.

    Science.gov (United States)

    Tiago, Teresa; Aureliano, Manuel; Gutiérrez-Merino, Carlos

    2004-05-11

    Decameric vanadate (V(10)) inhibits the actin-stimulated myosin ATPase activity, noncompetitively with actin or with ATP upon interaction with a high-affinity binding site (K(i) = 0.27 +/- 0.05 microM) in myosin subfragment-1 (S1). The binding of V(10) to S1 can be monitored from titration with V(10) of the fluorescence of S1 labeled at Cys-707 and Cys-697 with N-iodo-acetyl-N'-(5-sulfo-1-naphthyl)ethylenediamine (IAEDANS) or 5-(iodoacetamido) fluorescein, which showed the presence of only one V(10) binding site per monomer with a dissociation constant of 0.16-0.7 microM, indicating that S1 labeling with these dyes produced only a small distortion of the V(10) binding site. The large quenching of AEDANS-labeled S1 fluorescence produced by V(10) indicated that the V(10) binding site is close to Cys-697 and 707. Fluorescence studies demonstrated the following: (i) the binding of V(10) to S1 is not competitive either with actin or with ADP.V(1) or ADP.AlF(4); (ii) the affinity of V(10) for the complex S1/ADP.V(1) and S1/ADP.AlF(4) is 2- and 3-fold lower than for S1; and (iii) it is competitive with the S1 "back door" ligand P(1)P(5)-diadenosine pentaphosphate. A local conformational change in S1 upon binding of V(10) is supported by (i) a decrease of the efficiency of fluorescence energy transfer between eosin-labeled F-actin and fluorescein-labeled S1, and (ii) slower reassociation between S1 and F-actin after ATP hydrolysis. The results are consistent with binding of V(10) to the Walker A motif of ABC ATPases, which in S1 corresponds to conserved regions of the P-loop which form part of the phosphate tube.

  16. Aluminium fluoride and magnesium, activators of heterotrimeric GTP-binding proteins, affect high-affinity binding of the fungal toxin fusicoccin to the fusicoccin-binding protein in oat root plasma membranes.

    NARCIS (Netherlands)

    de Boer, A.H.; Van der Molen, G.W.; Prins, H.B.A.; Korthout, H.A.A.J.; van der Hoeven, P.C.J.

    1994-01-01

    The fusicoccin-binding protein was solubilised from purified oat root plasma membranes. The solubilised protein retained full binding activity, provided that protease inhibitors were included. Sodium fluoride reduced the high-affinity [H-3]fusicoccin binding to almost zero in a concentration-depende

  17. Reconstitution of high-affinity opioid agonist binding in brain membranes

    Energy Technology Data Exchange (ETDEWEB)

    Remmers, A.E.; Medzihradsky, F. (Univ. of Michigan Medical School, Ann Arbor (United States))

    1991-03-15

    In synaptosomal membranes from rat brain cortex, the {mu} selective agonist ({sup 3}H)dihydromorphine in the absence of sodium, and the nonselective antagonist ({sup 3}H)naltrexone in the presence of sodium, bound to two populations of opioid receptor sites with K{sub d} values of 0.69 and 8.7 nM for dihydromorphine, and 0.34 and 5.5 nM for naltrexone. The addition of 5 {mu}M guanosine 5{prime}-({gamma}-thio)triphosphate (GTP({gamma}S)) strongly reduced high-affinity agonist but not antagonist binding. Exposure of the membranes to high pH reduced the number of GTP({gamma}-{sup 35}S) binding sites by 90% and low K{sub m}, opioid-sensitive GTPase activity by 95%. In these membranes, high-affinity agonist binding was abolished and modulation of residual binding by GTP({gamma}S) was diminished. Alkali treatment of the glioma cell membranes prior to fusion inhibited most of the low K{sub m} GTPase activity and prevented the reconstitution of agonist binding. The results show that high-affinity opioid agonist binding reflects the ligand-occupied receptor - guanine nucleotide binding protein complex.

  18. Generation of high-performance binding proteins for peptide motifs by affinity clamping

    OpenAIRE

    Koide, Shohei; Huang, Jin

    2013-01-01

    We describe concepts and methodologies for generating “Affinity Clamps”, a new class of recombinant binding proteins that achieve high affinity and high specificity toward short peptide motifs of biological importance, which is a major challenge in protein engineering. The Affinity Clamping concept exploits the potential of nonhomologous recombination of protein domains in generating large changes in protein function and the inherent binding affinity and specificity of the so-called modular i...

  19. The protective effect of alpha lipoic acid on Schwann cells exposed to constant or intermittent high glucose.

    Science.gov (United States)

    Sun, Lian-Qing; Chen, Ying-Ying; Wang, Xuan; Li, Xiao-Jin; Xue, Bing; Qu, Ling; Zhang, Ting-Ting; Mu, Yi-Ming; Lu, Ju-Ming

    2012-10-01

    Diabetic peripheral neuropathy (DPN) is one of the most common and costly microvascular complications of diabetes, and no effective therapy exists. Previous studies have demonstrated that oxidative stress may be the unifying factor for the damaging effect of hyperglycemia. The aim of this study was to examine the impact of treatment with Alpha lipoic acid (ALA) on the intermittent high glucose (IHG) or high glucose (HG)-induced oxidative stress-induced mitochondrial pathway activation and Schwann cells (SCs) apoptosis in vitro. Our results suggested that IHG and HG induced SCs apoptosis in both caspase-dependent and caspase-independent pathways related to oxidative stress. More importantly, the cytotoxic effect of IHG was significantly more potent than that of HG. Treatment with ALA inhibited the IHG and HG-induced oxidative stress and apoptosis in SCs. Furthermore, treatment with ALA down-regulated the Bax expression and the release of cytochrome c and AIF translocation, but up-regulated the Bcl-2 expression in SCs. Treatment with ALA attenuated the activation of caspase-3 and caspase-9 and minimized the cleavage of PARP in SCs. These findings suggest that variability in glycemic control could be more deleterious than a constant HG and ALA antagonized the IHG-induced oxidative stress, activation of mitochondrial pathway and apoptosis in SCs.

  20. Possible effects of delivering methionine to broilers in drinking water at constant low and high environmental temperatures

    Directory of Open Access Journals (Sweden)

    Sahin Cadirci

    2014-02-01

    Full Text Available An experiment was conducted to study the effects of water-soluble DL-methionine supplied through water and feed on the performance and carcass yield of broilers housed at two constant temperatures from 21 to 42 days of age. Birds were housed in two rooms (240 birds per room with temperatures set at 21±2 and 30±2oC, respectively. Birds were divided into five groups of equal number within each room and fed five different diets (G1-G5. A low-methionine basal diet without supplemental methionine was fed to group 1 (G1. The basal diet was fortified with 0.025% or 0.050% methionine, either in feed (G2 and G3, respectively, or in a water solution (G4 and G5, respectively. Mortality was not significantly altered by any dietary treatment. Neither feed nor water intake was affected adversely by DLmethionine inclusion in drinking water. Housing at high temperature showed deleterious effect on birds’ weight gain. Additional methionine intake both in feed and water was associated with significantly heavier body weight, weight gain and feed conversion ratio, than the basal diet at low and high environmental temperature. Carcass yields, as a percentage of live body weight, were not affected by dietary treatment. The results indicate that, under the experimental conditions, DLmethionine provided in drinking water can be effectively assimilated by broilers, at least from 21 to 42 days of age.

  1. A Physically-Intuitive Method for Calculation of the Local Lattice Constant from a High-Resolution Transmission Electron Microscopy Image by Fourier Analysis

    CERN Document Server

    Teherani, James T

    2013-01-01

    We have developed a physically-intuitive method to calculate the local lattice constant as a function of position in a high-resolution transmission electron microscopy image by performing a two-dimensional fast Fourier transform. We apply a Gaussian filter with appropriate spatial full-width-half-max (FWHM) bandwidth to the image centered at the desired location to calculate the local lattice constant (as opposed to the average lattice constant). Fourier analysis of the filtered image yields the vertical and horizontal lattice constants at this location. The process is repeated by stepping the Gaussian filter across the image to produce a set of local lattice constants in the vertical and horizontal direction as a function of position in the image. The method has been implemented in a freely available tool on nanoHUB.

  2. High-precision limit on variation in the fine-structure constant from a single quasar absorption system

    Science.gov (United States)

    Kotuš, S. M.; Murphy, M. T.; Carswell, R. F.

    2017-01-01

    The brightest southern quasar above redshift z = 1, HE 0515-4414, with its strong intervening metal absorption line system at zabs = 1.1508, provides a unique opportunity to precisely measure or limit relative variations in the fine-structure constant (Δα/α). A variation of just ˜3 parts per million (ppm) would produce detectable velocity shifts between its many strong metal transitions. Using new and archival observations from the Ultraviolet and Visual Echelle Spectrograph (UVES), we obtain an extremely high signal-to-noise ratio spectrum (peaking at S/N ≈ 250 pix-1). This provides the most precise measurement of Δα/α from a single absorption system to date, Δα/α = -1.42 ± 0.55stat ± 0.65sys ppm, comparable with the precision from previous, large samples of ˜150 absorbers. The largest systematic error in all (but one) previous similar measurements, including the large samples, was long-range distortions in the wavelength calibration. These would add an ˜2 ppm systematic error to our measurement and up to ˜10 ppm to other measurements using Mg and Fe transitions. However, we corrected the UVES spectra using well-calibrated spectra of the same quasar from the High Accuracy Radial velocity Planet Searcher, leaving a residual 0.59 ppm systematic uncertainty, the largest contribution to our total systematic error. A similar approach, using short observations on future well-calibrated spectrographs to correct existing high S/N spectra, would efficiently enable a large sample of reliable Δα/α measurements. The high-S/N UVES spectrum also provides insights into analysis difficulties, detector artefacts and systematic errors likely to arise from 25-40-m telescopes.

  3. High-precision limit on variation in the fine-structure constant from a single quasar absorption system

    CERN Document Server

    Kotuš, Srđan M; Carswell, Robert F

    2016-01-01

    The brightest southern quasar above redshift $z=1$, HE 0515$-$4414, with its strong intervening metal absorption-line system at $z_{abs}=1.1508$, provides a unique opportunity to precisely measure or limit relative variations in the fine-structure constant ($\\Delta\\alpha/\\alpha$). A variation of just $\\sim$3 parts per million (ppm) would produce detectable velocity shifts between its many strong metal transitions. Using new and archival observations from the Ultraviolet and Visual Echelle Spectrograph (UVES) we obtain an extremely high signal-to-noise ratio spectrum (peaking at S/N $\\approx250$ pix$^{-1}$). This provides the most precise measurement of $\\Delta\\alpha/\\alpha$ from a single absorption system to date, $\\Delta\\alpha/\\alpha=-1.42\\pm0.55_{\\rm stat}\\pm0.65_{\\rm sys}$ ppm, comparable with the precision from previous, large samples of $\\sim$150 absorbers. The largest systematic error in all (but one) previous similar measurements, including the large samples, was long-range distortions in the wavelengt...

  4. Catalytic antibody light chain capable of cleaving a chemokine receptor CCR-5 peptide with a high reaction rate constant.

    Science.gov (United States)

    Mitsuda, Yukie; Hifumi, Emi; Tsuruhata, Kumi; Fujinami, Hiroko; Yamamoto, Naoki; Uda, Taizo

    2004-04-20

    A monoclonal antibody (MAb), ECL2B-2, was obtained by immunizing a peptide possessing a part of a sequence of a chemokine receptor, CCR-5, which is present as a membrane protein on the macrophage surface, and which plays an important role in human immunodeficiency virus (HIV) infection. From the DNA and the deduced amino acid sequences of the light and heavy chains of ECL2B-2 MAb, molecular modeling was conducted to calculate the steric conformation of the antibody. Modeling suggested that the structure of ECL2B-2 could possess one or two catalytic triad(s), composed of Asp(1), Ser(27a) (or Ser(27e)), and His(93) (or His(27d)), in the light chain of ECL2B-2. The three amino acid residues, Asp(1), Ser(27a), and His(93), are identical to those of catalytic antibody light chains such as VIPase and i41SL1-2. The light chain of ECL2B-2 MAb degraded the antigenic peptide CCR-5 within about 100 h. Surprisingly, the light chain had a very high catalytic reaction rate constant (k(cat)) of 2.23 min(-1), which is greater by factors of tens to hundreds than those of natural catalytic antibodies obtained previously. The heavy chain of ECL2B-2 MAb, which has no catalytic triad because of a lack of His residue, did not degrade the CCR-5 peptide.

  5. Optimal planning of series resistor to control time constant of test circuit for high-voltage AC circuit-breakers

    Directory of Open Access Journals (Sweden)

    Yoon-Ho Kim

    2016-01-01

    Full Text Available The equivalent test circuit that can deliver both short-circuit current and recovery voltage is used to verify the performance of high-voltage circuit breakers. Most of the parameters in this circuit can be obtained by using a simple calculation or a simulation program. The ratings of the circuit breaker include rated short-circuit breaking current, rated short-circuit making current, rated operating sequence of the circuit breaker and rated short-time current. Among these ratings, the short-circuit making capacity of the circuit breaker is expressed in peak value and not in RMS value similar to breaking capacity. A series resistor or super-excitation is used to control the peak value of the short-circuit current in the equivalent test circuit. When using a series resistor, a higher rating of circuit breakers leads to a higher thermal capacity, thereby requiring additional space. Therefore, an effective, optimal design of the series resistor is essential. This paper proposes a method for reducing thermal capacity and selecting the optimal resistance to limit the making current by controlling the DC time constant of the test circuit.

  6. Specific high-affinity binding sites for a synthetic gliadin heptapeptide of human peripheral blood lymphocytes

    Energy Technology Data Exchange (ETDEWEB)

    Payan, D.G.; Horvath, K.; Graf, L.

    1987-03-23

    The synthetic peptide containing residues 43-49 of ..cap alpha..-gliadin, the major protein component of gluten, has previously been shown to inhibit the production of lymphokine activities by mononuclear leukocytes. The authors demonstrate using radiolabeled ..cap alpha..-gliadin(43-49) that human peripheral blood lymphocytes express approximately 20,000-25,000 surface receptors for this peptide, with a dissociation constant (K/sub D/) of 20 nM. In addition, binding is inhibited by naloxone and an enkephalin analog, thus confirming the functional correlate which demonstrates inhibition by these agents of ..cap alpha..-gliadin(43-49) functional effects. Furthermore, B-lymphocytes bind specifically a greater amount of (/sup 125/I)..cap alpha..-gliadin(43-49) than T-lymphocytes. The lymphocyte ..cap alpha..-gliadin(43-49) receptor may play an important role in mediating the immunological response to ..cap alpha..-gliadin. 16 references, 4 figures.

  7. Identification of highly reactive cysteine residues at less exposed positions in the Fab constant region for site-specific conjugation.

    Science.gov (United States)

    Shiraishi, Yasuhisa; Muramoto, Takashige; Nagatomo, Kazutaka; Shinmi, Daisuke; Honma, Emiko; Masuda, Kazuhiro; Yamasaki, Motoo

    2015-06-17

    Engineered cysteine residues are currently used for the site-specific conjugation of antibody-drug conjugates (ADC). In general, positions on the protein surface have been selected for substituting a cysteine as a conjugation site; however, less exposed positions (with less than 20% of accessible surface area [ASA]) have not yet been evaluated. In this study, we engineered original cysteine positional variants of a Fab fragment, with less than 20% of ASA, and evaluated their thiol reactivities through conjugation with various kinds of payloads. As a result, we have identified three original cysteine positional variants (heavy chain: Hc-A140C, light chain: Lc-Q124C and Lc-L201C), which exhibited similar monomer content, thermal stability, and antigen binding affinity in comparison to the wild-type Fab. In addition, the presence of cysteine in these positions made it possible for the Fab variants to react with variable-sized molecules with high efficiency. The favorable physical properties of the cysteine positional variants selected in our study suggest that less exposed positions, with less than 20% of ASA, provide an alternative for creating conjugation sites.

  8. A High-Affinity Metal-Binding Peptide From Escherichia Coli Hypb

    Energy Technology Data Exchange (ETDEWEB)

    Chung, K.C.Chan; Cao, L.; Dias, A.V.; Pickering, I.J.; George, G.N.; Zamble, D.B.

    2009-05-12

    The high-affinity nickel-binding site of the Escherichia coli [NiFe]-hydrogenase accessory protein HypB was localized to residues at the immediate N-terminus of the protein. Modification of a metal-binding fusion protein, site-directed mutagenesis experiments, and DFT calculations were used to identify the N-terminal amine as a ligand as well as the three cysteine residues in the CXXCGCXXX motif. This sequence can be removed from the protein and both a synthesized peptide and a protein fusion bind nickel with a similar affinity and the same structure as the parent metalloprotein, indicating the self-sufficiency of this high-affinity nickel-binding sequence.

  9. High-throughput engineering and analysis of peptide binding to class II MHC.

    Science.gov (United States)

    Jiang, Wei; Boder, Eric T

    2010-07-27

    Class II major histocompatibility complex (MHC-II) proteins govern stimulation of adaptive immunity by presenting antigenic peptides to CD4+ T lymphocytes. Many allelic variants of MHC-II exist with implications in peptide presentation and immunity; thus, high-throughput experimental tools for rapid and quantitative analysis of peptide binding to MHC-II are needed. Here, we present an expression system wherein peptide and MHC-II are codisplayed on the surface of yeast in an intracellular association-dependent manner and assayed by flow cytometry. Accordingly, the relative binding of different peptides and/or MHC-II variants can be assayed by genetically manipulating either partner, enabling the application of directed evolution approaches for high-throughput characterization or engineering. We demonstrate the application of this tool to map the side-chain preference for peptides binding to HLA-DR1 and to evolve novel HLA-DR1 mutants with altered peptide-binding specificity.

  10. Fast high-throughput method for the determination of acidity constants by capillary electrophoresis: I. Monoprotic weak acids and bases.

    Science.gov (United States)

    Fuguet, Elisabet; Ràfols, Clara; Bosch, Elisabeth; Rosés, Martí

    2009-04-24

    A new and fast method to determine acidity constants of monoprotic weak acids and bases by capillary zone electrophoresis based on the use of an internal standard (compound of similar nature and acidity constant as the analyte) has been developed. This method requires only two electrophoretic runs for the determination of an acidity constant: a first one at a pH where both analyte and internal standard are totally ionized, and a second one at another pH where both are partially ionized. Furthermore, the method is not pH dependent, so an accurate measure of the pH of the buffer solutions is not needed. The acidity constants of several phenols and amines have been measured using internal standards of known pK(a), obtaining a mean deviation of 0.05 pH units compared to the literature values.

  11. Design of high-linear CMOS circuit using a constant transconductance method for gamma-ray spectroscopy system

    Energy Technology Data Exchange (ETDEWEB)

    Jung, I.I. [School of Electrical and Electronics Engineering, Chung-Ang University, Seoul 156-756 (Korea, Republic of); Lee, J.H. [Institute of Innovative Functional Imaging, Chung-Ang University, Seoul 156-756 (Korea, Republic of); Lee, C.S. [Department of Physics, Chung-Ang University, Seoul 156-756 (Korea, Republic of); Choi, Y.-W., E-mail: ychoi@cau.ac.k [School of Electrical and Electronics Engineering, Chung-Ang University, Seoul 156-756 (Korea, Republic of)

    2011-02-11

    We propose a novel circuit to be applied to the front-end integrated circuits of gamma-ray spectroscopy systems. Our circuit is designed as a type of current conveyor (ICON) employing a constant-g{sub m} (transconductance) method which can significantly improve the linearity in the amplified signals by using a large time constant and the time-invariant characteristics of an amplifier. The constant-g{sub m} method is obtained by a feedback control which keeps the transconductance of the input transistor constant. To verify the performance of the propose circuit, the time constant variations for the channel resistances are simulated with the TSMC 0.18{mu}m transistor parameters using HSPICE, and then compared with those of a conventional ICON. As a result, the proposed ICON shows only 0.02% output linearity variation and 0.19% time constant variation for the input amplitude up to 100 mV. These are significantly small values compared to a conventional ICON's 1.39% and 19.43%, respectively, for the same conditions.

  12. Computational Calorimetry: High-Precision Calculation of Host-Guest Binding Thermodynamics.

    Science.gov (United States)

    Henriksen, Niel M; Fenley, Andrew T; Gilson, Michael K

    2015-09-08

    We present a strategy for carrying out high-precision calculations of binding free energy and binding enthalpy values from molecular dynamics simulations with explicit solvent. The approach is used to calculate the thermodynamic profiles for binding of nine small molecule guests to either the cucurbit[7]uril (CB7) or β-cyclodextrin (βCD) host. For these systems, calculations using commodity hardware can yield binding free energy and binding enthalpy values with a precision of ∼0.5 kcal/mol (95% CI) in a matter of days. Crucially, the self-consistency of the approach is established by calculating the binding enthalpy directly, via end point potential energy calculations, and indirectly, via the temperature dependence of the binding free energy, i.e., by the van't Hoff equation. Excellent agreement between the direct and van't Hoff methods is demonstrated for both host-guest systems and an ion-pair model system for which particularly well-converged results are attainable. Additionally, we find that hydrogen mass repartitioning allows marked acceleration of the calculations with no discernible cost in precision or accuracy. Finally, we provide guidance for accurately assessing numerical uncertainty of the results in settings where complex correlations in the time series can pose challenges to statistical analysis. The routine nature and high precision of these binding calculations opens the possibility of including measured binding thermodynamics as target data in force field optimization so that simulations may be used to reliably interpret experimental data and guide molecular design.

  13. New percolative BaTiO{sub 3}-Ni composites with a high and frequency-independent dielectric constant ({epsilon}{sub r}{approx}80000)

    Energy Technology Data Exchange (ETDEWEB)

    Pecharroman, C.; Esteban-Betegon, F.; Bartolome, J.F.; Lopez-Esteban, S.; Moya, J.S. [Consejo Superior de Investigaciones Cientificas, Madrid (Spain). Inst. de Ciencia de Materiales

    2001-10-16

    A novel capacitor device with a high, frequency-independent dielectric constant has been fabricated by a simple powder processing route. A mixture of microparticles of ferroelectric BaTiO{sub 3} and de-passivated metallic nickel was sintered to yield percolative composites in which the system of conducting particles separated by insulator layers exhibits a dielectric constant higher than that of either component. (orig.)

  14. High-throughput prediction of RNA, DNA and protein binding regions mediated by intrinsic disorder.

    Science.gov (United States)

    Peng, Zhenling; Kurgan, Lukasz

    2015-10-15

    Intrinsically disordered proteins and regions (IDPs and IDRs) lack stable 3D structure under physiological conditions in-vitro, are common in eukaryotes, and facilitate interactions with RNA, DNA and proteins. Current methods for prediction of IDPs and IDRs do not provide insights into their functions, except for a handful of methods that address predictions of protein-binding regions. We report first-of-its-kind computational method DisoRDPbind for high-throughput prediction of RNA, DNA and protein binding residues located in IDRs from protein sequences. DisoRDPbind is implemented using a runtime-efficient multi-layered design that utilizes information extracted from physiochemical properties of amino acids, sequence complexity, putative secondary structure and disorder and sequence alignment. Empirical tests demonstrate that it provides accurate predictions that are competitive with other predictors of disorder-mediated protein binding regions and complementary to the methods that predict RNA- and DNA-binding residues annotated based on crystal structures. Application in Homo sapiens, Mus musculus, Caenorhabditis elegans and Drosophila melanogaster proteomes reveals that RNA- and DNA-binding proteins predicted by DisoRDPbind complement and overlap with the corresponding known binding proteins collected from several sources. Also, the number of the putative protein-binding regions predicted with DisoRDPbind correlates with the promiscuity of proteins in the corresponding protein-protein interaction networks. Webserver: http://biomine.ece.ualberta.ca/DisoRDPbind/.

  15. Characterization of high affinity binding motifs for the discoidin domain receptor DDR2 in collagen.

    Science.gov (United States)

    Konitsiotis, Antonios D; Raynal, Nicolas; Bihan, Dominique; Hohenester, Erhard; Farndale, Richard W; Leitinger, Birgit

    2008-03-14

    The discoidin domain receptors, DDR1 and DDR2, are receptor tyrosine kinases that are activated by native triple-helical collagen. Here we have located three specific DDR2 binding sites by screening the entire triple-helical domain of collagen II, using the Collagen II Toolkit, a set of overlapping triple-helical peptides. The peptide sequence that bound DDR2 with highest affinity interestingly contained the sequence for the high affinity binding site for von Willebrand factor in collagen III. Focusing on this sequence, we used a set of truncated and alanine-substituted peptides to characterize the sequence GVMGFO (O is hydroxyproline) as the minimal collagen sequence required for DDR2 binding. Based on a recent NMR analysis of the DDR2 collagen binding domain, we generated a model of the DDR2-collagen interaction that explains why a triple-helical conformation is required for binding. Triple-helical peptides comprising the DDR2 binding motif not only inhibited DDR2 binding to collagen II but also activated DDR2 transmembrane signaling. Thus, DDR2 activation may be effected by single triple-helices rather than fibrillar collagen.

  16. Room temperature plasma oxidation: A new process for preparation of ultrathin layers of silicon oxide, and high dielectric constant materials

    Energy Technology Data Exchange (ETDEWEB)

    Tinoco, J.C. [Seccion de Electronica del Estado Solido, Depto. Ingenieria Electrica, CINVESTAV-IPN, Av. IPN, No. 2508, CP 07730, Mexico D.F. (Mexico) and Facultad de Ingenieria y Arquitectura, Universidad de San Martin de Porres (FIA-USMP), Av. La Fontana 1250, La Molina, Lima 12 (Peru)]. E-mail: jcesartinoco@yahoo.com.mx; Estrada, M. [Seccion de Electronica del Estado Solido, Depto. Ingenieria Electrica, CINVESTAV-IPN, Av. IPN, No. 2508, CP 07730, Mexico D.F. (Mexico)]. E-mail: mestrada@mail.cinvestav.mx; Baez, H. [Seccion de Electronica del Estado Solido, Depto. Ingenieria Electrica, CINVESTAV-IPN, Av. IPN, No. 2508, CP 07730, Mexico D.F. (Mexico); Cerdeira, A. [Seccion de Electronica del Estado Solido, Depto. Ingenieria Electrica, CINVESTAV-IPN, Av. IPN, No. 2508, CP 07730, Mexico D.F. (Mexico)

    2006-02-21

    In this paper we present basic features and oxidation law of the room temperature plasma oxidation (RTPO), as a new process for preparation of less than 2 nm thick layers of SiO{sub 2}, and high-k layers of TiO{sub 2}. We show that oxidation rate follows a potential law dependence on oxidation time. The proportionality constant is function of pressure, plasma power, reagent gas and plasma density, while the exponent depends only on the reactive gas. These parameters are related to the physical phenomena occurring inside the plasma, during oxidation. Metal-Oxide-Semiconductor (MOS) capacitors fabricated with these layers are characterized by capacitance-voltage, current-voltage and current-voltage-temperature measurements. Less than 2.5 nm SiO{sub 2} layers with surface roughness similar to thermal oxide films, surface state density below 3 x 10{sup 11} cm{sup -2} and current density in the expected range for each corresponding thickness, were obtained by RTPO in a parallel-plate reactor, at 180 mW/cm{sup 2} and pressure range between 9.33 and 66.5 Pa (0.07 and 0.5 Torr) using O{sub 2} and N{sub 2}O as reactive gases. MOS capacitors with TiO{sub 2} layers formed by RTPO of sputtered Ti layers are also characterized. Finally, MOS capacitors with stacked layers of TiO{sub 2} over SiO{sub 2}, both layers obtained by RTPO, were prepared and evaluated to determine the feasibility of the use of TiO{sub 2} as a candidate for next technology nodes.

  17. High Purity DNA Extraction with a SPE Microfluidic Chip Using KI as the Binding Salt

    Institute of Scientific and Technical Information of China (English)

    Xing CHEN; Da Fu CUI; Chang Chun LIU

    2006-01-01

    Based on solid phase extraction method, a novel silicon-PDMS-glass microchip for high purity DNA extraction has been developed by using KI as the binding salt. The microfluidic chip fabricated by MEMS technology was composed of a silicon substrate with a coiled channel and a compounded PDMS-glass cover. With this microfluidic chip, the wall of the coiled channel was used as solid phase matrix for binding DNA and DNA was extracted by the fluxion of the binding buffer, washing buffer and elution buffer. KI as a substitute for guanidine, was used successfully as binding salt for purification DNA, obtaining higher purity of genomic DNA and about 13.9 ng DNA from 1 μL rat whole blood in 35 minutes.

  18. Maintenance of energy expenditure on high-protein vs. high-carbohydrate diets at a constant body weight may prevent a positive energy balance.

    Science.gov (United States)

    Martens, E A; Gonnissen, H K; Gatta-Cherifi, B; Janssens, P L; Westerterp-Plantenga, M S

    2015-10-01

    Relatively high-protein diets are effective for body weight loss, and subsequent weight maintenance, yet it remains to be shown whether these diets would prevent a positive energy balance. Therefore, high-protein diet studies at a constant body weight are necessary. The objective was to determine fullness, energy expenditure, and macronutrient balances on a high-protein low-carbohydrate (HPLC) diet compared with a high-carbohydrate low-protein (HCLP) diet at a constant body weight, and to assess whether effects are transient or sustained after 12 weeks. A randomized parallel study was performed in 14 men and 18 women [mean ± SD age: 24 ± 5 y; BMI (in kg/m(2)): 22.8 ± 2.0] on diets containing 30/35/35 (HPLC) or 5/60/35 (HCLP) % of energy from protein/carbohydrate/fat. Significant interactions between dietary intervention and time on total energy expenditure (TEE) (P = 0.013), sleeping metabolic rate (SMR) (P = 0.040), and diet-induced thermogenesis (DIT) (P = 0.027) appeared from baseline to wk 12. TEE was maintained in the HPLC diet group, while it significantly decreased throughout the intervention period in the HCLP diet group (wk 1: P = 0.002; wk 12: P = 0.001). Energy balance was maintained in the HPLC diet group, and became positive in the HCLP diet group at wk 12 (P = 0.008). Protein balance varied directly according to the amount of protein in the diet, and diverged significantly between the diets (P = 0.001). Fullness ratings were significantly higher in the HPLC vs. the HCLP diet group at wk 1 (P = 0.034), but not at wk 12. Maintenance of energy expenditure on HPLC vs. HCLP diets at a constant body weight may prevent development of a positive energy balance, despite transiently higher fullness. The study was registered on clinicaltrials.gov with Identifier: NCT01551238. Copyright © 2014 Elsevier Ltd and European Society for Clinical Nutrition and Metabolism. All rights reserved.

  19. Quantification aspects of constant pressure (ultra) high pressure liquid chromatography using mass-sensitive detectors with a nebulizing interface.

    Science.gov (United States)

    Verstraeten, M; Broeckhoven, K; Lynen, F; Choikhet, K; Landt, K; Dittmann, M; Witt, K; Sandra, P; Desmet, G

    2013-01-25

    The present contribution investigates the quantitation aspects of mass-sensitive detectors with nebulizing interface (ESI-MSD, ELSD, CAD) in the constant pressure gradient elution mode. In this operation mode, the pressure is controlled and maintained at a set value and the liquid flow rate will vary according to the inverse mobile phase viscosity. As the pressure is continuously kept at the allowable maximum during the entire gradient run, the average liquid flow rate is higher compared to that in the conventional constant flow rate operation mode, thus shortening the analysis time. The following three mass-sensitive detectors were investigated: mass spectrometry detector (MS), evaporative light scattering detector (ELSD) and charged aerosol detector (CAD) and a wide variety of samples (phenones, polyaromatic hydrocarbons, wine, cocoa butter) has been considered. It was found that the nebulizing efficiency of the LC-interfaces of the three detectors under consideration changes with the increasing liquid flow rate. For the MS, the increasing flow rate leads to a lower peak area whereas for the ELSD the peak area increases compared to the constant flow rate mode. The peak area obtained with a CAD is rather insensitive to the liquid flow rate. The reproducibility of the peak area remains similar in both modes, although variation in system permeability compromises the 'long-term' reproducibility. This problem can however be overcome by running a flow rate program with an optimized flow rate and composition profile obtained from the constant pressure mode. In this case, the quantification remains reproducibile, despite any occuring variations of the system permeability. Furthermore, the same fragmentation pattern (MS) has been found in the constant pressure mode compared to the customary constant flow rate mode.

  20. High Efficiency Binding Aptamers for a Wide Range of Bacterial Sepsis Agents.

    Science.gov (United States)

    Graziani, Ana Cláudia; Stets, Maria Isabel; Lopes, Ana Luisa Kalb; Schluga, Pedro Henrique Caires; Marton, Soledad; Mendes, Ieda Ferreira; Andrade, Antero Silva Ribeiro de; Krieger, Marco Aurélio; Cardoso, Josiane

    2017-04-28

    Sepsis is a major health problem worldwide, with an extremely high rate of morbidity and mortality, partly due to delayed diagnosis during early disease. Currently, sepsis diagnosis requires bacterial culturing of blood samples over several days, whereas PCR-based molecular diagnosis methods are faster but lack sensitivity. The use of biosensors containing nucleic acid aptamers that bind targets with high affinity and specificity could accelerate sepsis diagnosis. Previously, we used the systematic evolution of ligands by exponential enrichment technique to develop the aptamers Antibac1 and Antibac2, targeting the ubiquitous bacterial peptidoglycan. Here, we show that these aptamers bind to four gram-positive and seven gram-negative bacterial sepsis agents with high binding efficiency. Thus, these aptamers could be used in combination as biological recognition elements in the development of biosensors that are an alternative to rapid bacteria detection, since they could provide culture and amplification-free tests for rapid clinical sepsis diagnosis.

  1. High affinity binding of /sup 125/I-labeled mouse interferon to a specific cell surface receptor. II. Analysis of binding properties

    Energy Technology Data Exchange (ETDEWEB)

    Aguet, M.; Blanchard, B.

    1981-12-01

    Direct ligand-binding studies with highly purified /sup 125/I-labeled virus-induced mouse interferon on mouse lymphoma L 1210 cells revealed a direct correlation of specific high-affinity binding with the biologic response to interferon. Neutralization of the antiviral effect by anti-interferon gamma globulin occurred at the same antibody concentration as the inhibition of specific binding. These results suggest that specific high-affinity binding of /sup 125/I-interferon occurred at a biologically functional interferon receptor. Competitive inhibition experiments using /sup 125/I- and /sup 127/I-labeled interferon provided strong evidence that the fraction of /sup 125/I-interferon inactivated upon labeling did not bind specifically. Scatchard analysis of the binding data yielded linear plots and thus suggested that interferon binds to homogeneous noncooperative receptor sites. In contrast to a characteristic property of several peptide hormone systems, binding of /sup 125/I-interferon to its specific receptor did not induce subsequent ligand degradation. At 37/sup o/ bound interferon was rapidly released in a biologically active form without evidence for molecular degradation. The expression of interferon receptors was not modified by treatment with interferon. Trypsin treatment of target cells and inhibition of protein synthesis abolished the specific binding of /sup 125/I-interferon. Three major molecular weight species of Newcastle disease virus-induced mouse C 243 cell interferon were isolated, separated, and identified as mouse ..cap alpha.. and ..beta.. interferons. These interferons were shown to inhibit competitively the specific binding of the highly purified labeled starting material thus providing evidence for a common receptor site for mouse interferon.

  2. Colorimetry and constant-potential coulometry determinations of transferrin-bound iron, total iron-binding capacity, and total iron in serum containing iron-dextran, with use of sodium dithionite and alumina columns.

    Science.gov (United States)

    Jacobs, J C; Alexander, N M

    1990-10-01

    After the parenteral administration of iron-dextran (imferon), the increased total iron concentrations in serum can be determined by atomic absorption spectroscopy and by colorimetric methods involving sodium dithionite, which reductively dissociates iron from the dextran complex. We report that constant-potential coulometry detects only about 55-70% of dextran-bound iron before dithionite reduction and variable amounts after reaction with the reducing agent. In addition, we have developed a procedure for determining transferrin-bound iron, total iron-binding capacity (TIBC), total iron, and dextran-bound iron with the Kodak Ektachem colorimetric system. In determining total serum iron, the sample is first mixed with sodium dithionite, which rapidly dissociates all dextran-bound iron, but does not remove iron from either transferrin or hemoglobin. After the mixture is applied to an Ektachem slide, transferrin-bound iron is released at pH 4 and is detected together with the iron previously bound to dextran. TIBC is determined by mixing serum with ferric citrate in moderate excess and filtering through a small alumina (Al2O3) column, which binds excess free iron and iron-dextran; the iron in the column eluate represents the TIBC. Transferrin-bound iron is determined by applying diluted serum without added ferric citrate to an alumina column and measuring the iron in the column eluate. Dextran-bound iron is equivalent to the difference between total and transferrin-bound iron. Using this method, we found that transferrin iron-binding sites are saturated in vitro by excess iron-dextran less efficiently than by ferric citrate.

  3. Detrimental effects of constant light exposure and high-fat diet on circadian energy metabolism and insulin sensitivity

    NARCIS (Netherlands)

    Coomans, C.P.; Berg, S.A.A. van den; Houben, T.; Klinken, J.B. van; Berg, R. van den; Pronk, A.C.M.; Havekes, L.M.; Romijn, J.A.; Dijk, K.W. van; Biermasz, N.R.; Meijer, J.H.

    2013-01-01

    Circadian rhythm disturbances are observed in, e.g., aging and neurodegenerative diseases and are associated with an increased incidence of obesity and diabetes. We subjected male C57Bl/6J mice to constant light [12-h light-light (LL) cycle] to examine the effects of a disturbed circadian rhythm on

  4. Further characterization of the low and high affinity binding components of the thyrotropin receptor.

    Science.gov (United States)

    McQuade, R; Thomas, C G; Nayfeh, S N

    1986-05-29

    Following cross-linking with disuccinimidyl suberate and analysis by SDS-PAGE and autoradiography, both the high- and low-affinity TSH binding components exhibited two similar 125I-TSH-labeled bands, with Mr values of 80,000 and 68,000. IgG fractions from patients with Graves' disease inhibited 125I-TSH binding to both components, while normal IgG had no effect. Although not entirely conclusive, these results suggest that the high- and low-affinity components share similar subunit composition and antigenic determinants.

  5. Further characterization of the low and high affinity binding components of the thyrotropin receptor

    Energy Technology Data Exchange (ETDEWEB)

    McQuade, R.; Thomas, C.G. Jr.; Nayfeh, S.N.

    1986-05-29

    Following cross-linking with disuccinimdiyl suberate and analysis by SDS-PAGE and autoradiography, both the high- and low-affinity TSH binding components exhibited two similar /sup 125/I-TSH-labeled bands, with Mr values of 80,000 and 68,000. IgG fractions from patients with Graves' disease inhibited /sup 125/I-TSH binding to both components, while normal IgG had no effect. Although not entirely conclusive, these results suggest that the high- and low-affinity components share similar subunit composition and antigenic determinants.

  6. CSI-FID: high throughput label-free detection of DNA binding molecules.

    Science.gov (United States)

    Hauschild, Karl E; Stover, James S; Boger, Dale L; Ansari, Aseem Z

    2009-07-15

    Determining the sequence specifity of DNA binding molecules is a non-trivial task. Here we describe the development of a platform for assaying the sequence specificity of DNA ligands using label free detection on high density DNA microarrays. This is achieved by combining Cognate Site Identification (CSI) with Fluorescence Intercalation Displacement (FID) to create CSI-FID. We use the well-studied small molecule DNA ligand netropsin to develop this high throughput platform. Analysis of the DNA binding properties of protein- and small molecule-based libraries with CSI-FID will advance the development of genome-anchored molecules for therapeutic purposes.

  7. High-affinity cannabinoid binding site in brain: A possible marijuana receptor

    Energy Technology Data Exchange (ETDEWEB)

    Nye, J.S.

    1988-01-01

    The mechanism by which delta{sup 9} tetrahydrocannabinol (delta{sup 9}THC), the major psychoactive component of marijuana or hashish, produces its potent psychological and physiological effects is unknown. To find receptor binding sites for THC, we designed a water-soluble analog for use as a radioligand. 5{prime}-Trimethylammonium-delta{sup 8}THC (TMA) is a positively charged analog of delta-{sup 8}THC modified on the 5{prime} carbon, a portion of the molecule not important for its psychoactivity. We have studied the binding of ({sup 3}H)-5{prime}-trimethylammonium-delta-{sup 8}THC (({sup 3}H)TMA) to rat neuronal membranes. ({sup 3}H)TMA binds saturably and reversibly to brain membranes with high affinity to apparently one class of sites. Highest binding site density occurs in brain, but several peripheral organs also display specific binding. Detergent solubilizes the sites without affecting their pharmacologial properties. Molecular sieve chromatography reveals a bimodal peak of ({sup 3}H)TMA binding activity of approximately 60,000 daltons apparent molecular weight.

  8. Specific binding of eukaryotic ORC to DNA replication origins depends on highly conserved basic residues.

    Science.gov (United States)

    Kawakami, Hironori; Ohashi, Eiji; Kanamoto, Shota; Tsurimoto, Toshiki; Katayama, Tsutomu

    2015-10-12

    In eukaryotes, the origin recognition complex (ORC) heterohexamer preferentially binds replication origins to trigger initiation of DNA replication. Crystallographic studies using eubacterial and archaeal ORC orthologs suggested that eukaryotic ORC may bind to origin DNA via putative winged-helix DNA-binding domains and AAA+ ATPase domains. However, the mechanisms how eukaryotic ORC recognizes origin DNA remain elusive. Here, we show in budding yeast that Lys-362 and Arg-367 residues of the largest subunit (Orc1), both outside the aforementioned domains, are crucial for specific binding of ORC to origin DNA. These basic residues, which reside in a putative disordered domain, were dispensable for interaction with ATP and non-specific DNA sequences, suggesting a specific role in recognition. Consistent with this, both residues were required for origin binding of Orc1 in vivo. A truncated Orc1 polypeptide containing these residues solely recognizes ARS sequence with low affinity and Arg-367 residue stimulates sequence specific binding mode of the polypeptide. Lys-362 and Arg-367 residues of Orc1 are highly conserved among eukaryotic ORCs, but not in eubacterial and archaeal orthologs, suggesting a eukaryote-specific mechanism underlying recognition of replication origins by ORC.

  9. Quinine binding by the cocaine-binding aptamer. Thermodynamic and hydrodynamic analysis of high-affinity binding of an off-target ligand.

    Science.gov (United States)

    Reinstein, Oren; Yoo, Mina; Han, Chris; Palmo, Tsering; Beckham, Simone A; Wilce, Matthew C J; Johnson, Philip E

    2013-12-03

    The cocaine-binding aptamer is unusual in that it tightly binds molecules other than the ligand it was selected for. Here, we study the interaction of the cocaine-binding aptamer with one of these off-target ligands, quinine. Isothermal titration calorimetry was used to quantify the quinine-binding affinity and thermodynamics of a set of sequence variants of the cocaine-binding aptamer. We find that the affinity of the cocaine-binding aptamer for quinine is 30-40 times stronger than it is for cocaine. Competitive-binding studies demonstrate that both quinine and cocaine bind at the same site on the aptamer. The ligand-induced structural-switching binding mechanism of an aptamer variant that contains three base pairs in stem 1 is retained with quinine as a ligand. The short stem 1 aptamer is unfolded or loosely folded in the free form and becomes folded when bound to quinine. This folding is confirmed by NMR spectroscopy and by the short stem 1 construct having a more negative change in heat capacity of quinine binding than is seen when stem 1 has six base pairs. Small-angle X-ray scattering (SAXS) studies of the free aptamer and both the quinine- and the cocaine-bound forms show that, for the long stem 1 aptamers, the three forms display similar hydrodynamic properties, and the ab initio shape reconstruction structures are very similar. For the short stem 1 aptamer there is a greater variation among the SAXS-derived ab initio shape reconstruction structures, consistent with the changes expected with its structural-switching binding mechanism.

  10. Novel cyclen-based linear polymer as a high-affinity binding material for DNA condensation

    Institute of Scientific and Technical Information of China (English)

    XIANG YongZhe; WANG Na; ZHANG Ji; LI Kun; ZHANG ZhongWei; LIN HongHui; YU XiaoQi

    2009-01-01

    A novel cyclen-based linear polyamine (POGEC) was designed and synthesized from the reaction be-tween 1,3-propanediol diglycidyl ether and 1,7-bis(diethoxyphosphory)-1,4,7,10-tetraazacyclod- odecane.High-affinity binding between POGEC and DNA was demonstrated by agarose gel electrophoresis and scanning electron microscopy (SEM). Moreover, the formed POGEC/DNA complex (termed polyplex) could be disassociated to release the free DNA through addition of the physiological concentration of NaCl solution. Fluorescence spectrum was used to measure the high-affinity binding and DNA con-densation capability of POGEC. Circular dichroism (CD) spectrum indicates that the DNA conformation did not change after binding to POEGC.

  11. Novel cyclen-based linear polymer as a high-affinity binding material for DNA condensation

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    A novel cyclen-based linear polyamine (POGEC) was designed and synthesized from the reaction between 1,3-propanediol diglycidyl ether and 1,7-bis(diethoxyphosphory)-1,4,7,10-tetraazacyclod-odecane. High-affinity binding between POGEC and DNA was demonstrated by agarose gel electrophoresis and scanning electron microscopy (SEM). Moreover,the formed POGEC/DNA complex (termed polyplex) could be disassociated to release the free DNA through addition of the physiological concentration of NaCl solution. Fluorescence spectrum was used to measure the high-affinity binding and DNA condensation capability of POGEC. Circular dichroism (CD) spectrum indicates that the DNA conformation did not change after binding to POEGC.

  12. Functional recombinant MHC class II molecules and high-throughput peptide-binding assays

    DEFF Research Database (Denmark)

    Justesen, Sune; Harndahl, Mikkel; Lamberth, Kasper

    2009-01-01

    of peptide-binding assay were developed including a homogeneous, non-radioactive, high-throughput (HTS) binding assay. Binding isotherms were generated allowing the affinities of interaction to be determined. The affinities of the best binders were found to be in the low nanomolar range. Recombinant MHC...... in the generation of MHC-II molecules as reagents to study and manipulate specific T helper cell responses. Methods to generate functional MHC-II molecules recombinantly, and measure their interaction with peptides, would be highly desirable; however, no consensus methodology has yet emerged. RESULTS: We generated....... CONCLUSION: We have successfully developed versatile MHC-II resources, which may assist in the generation of MHC class II -wide reagents, data, and tools....

  13. Neurotensin decreases high affinity [3H]-ouabain binding to cerebral cortex membranes.

    Science.gov (United States)

    Rosin, Carina; Ordieres, María Graciela López; Arnaiz, Georgina Rodríguez de Lores

    2011-12-10

    Previous work from this laboratory showed the ability of neurotensin to inhibit synaptosomal membrane Na(+), K(+)-ATPase activity, the effect being blocked by SR 48692, a non-peptidic antagonist for high affinity neurotensin receptor (NTS1) [López Ordieres and Rodríguez de Lores Arnaiz 2000; 2001]. To further study neurotensin interaction with Na(+), K(+)-ATPase, peptide effect on high affinity [(3)H]-ouabain binding was studied in cerebral cortex membranes. It was observed that neurotensin modified binding in a dose-dependent manner, leading to 80% decrease with 1 × 10(-4)M concentration. On the other hand, the single addition of 1 × 10(-6)M, 1 × 10(-5)M and 1 × 10(-4)M SR 48692 (Sanofi-Aventis, U.S., Inc.) decreased [(3)H]-ouabain binding (in %) to 87 ± 16; 74 ± 16 and 34 ± 17, respectively. Simultaneous addition of neurotensin and SR 48692 led to additive or synergic effects. Partial NTS2 agonist levocabastine inhibited [(3)H]-ouabain binding likewise. Saturation assays followed by Scatchard analyses showed that neurotensin increased K(d) value whereas failed to modify B(max) value, indicating a competitive type interaction of the peptide at Na(+), K(+)-ATPase ouabain site. At variance, SR 48692 decreased B(max) value whereas it did not modify K(d) value. [(3)H]-ouabain binding was also studied in cerebral cortex membranes obtained from rats injected i. p. 30 min earlier with 100 μg and 250 μg/kg SR 48692. It was observed that the 250 μg/kg SR 48692 dose led to 19% decrease in basal [(3)H]-ouabain binding. After SR 48692 treatments, addition of 1 × 10(-6)M led to additive or synergic effect. Results suggested that [(3)H]-ouabain binding inhibition by neurotensin hardly involves NTS1 receptor.

  14. GHB receptor targets in the CNS: focus on high-affinity binding sites.

    Science.gov (United States)

    Bay, Tina; Eghorn, Laura F; Klein, Anders B; Wellendorph, Petrine

    2014-01-15

    γ-Hydroxybutyric acid (GHB) is an endogenous compound in the mammalian brain with both low- and high-affinity receptor targets. GHB is used clinically in the treatment of symptoms of narcolepsy and alcoholism, but also illicitly abused as the recreational drug Fantasy. Major pharmacological effects of exogenous GHB are mediated by GABA subtype B (GABAB) receptors that bind GHB with low affinity. The existence of GHB high-affinity binding sites has been known for more than three decades, but the uncovering of their molecular identity has only recently begun. This has been prompted by the generation of molecular tools to selectively study high-affinity sites. These include both genetically modified GABAB knock-out mice and engineered selective GHB ligands. Recently, certain GABA subtype A (GABAA) receptor subtypes emerged as high-affinity GHB binding sites and potential physiological mediators of GHB effects. In this research update, a description of the various reported receptors for GHB is provided, including GABAB receptors, certain GABAA receptor subtypes and other reported GHB receptors. The main focus will thus be on the high-affinity binding targets for GHB and their potential functional roles in the mammalian brain.

  15. Integrated microfluidic approach for quantitative high-throughput measurements of transcription factor binding affinities.

    Science.gov (United States)

    Glick, Yair; Orenstein, Yaron; Chen, Dana; Avrahami, Dorit; Zor, Tsaffrir; Shamir, Ron; Gerber, Doron

    2016-04-07

    Protein binding to DNA is a fundamental process in gene regulation. Methodologies such as ChIP-Seq and mapping of DNase I hypersensitive sites provide global information on this regulation in vivo In vitro methodologies provide valuable complementary information on protein-DNA specificities. However, current methods still do not measure absolute binding affinities. There is a real need for large-scale quantitative protein-DNA affinity measurements. We developed QPID, a microfluidic application for measuring protein-DNA affinities. A single run is equivalent to 4096 gel-shift experiments. Using QPID, we characterized the different affinities of ATF1, c-Jun, c-Fos and AP-1 to the CRE consensus motif and CRE half-site in two different genomic sequences on a single device. We discovered that binding of ATF1, but not of AP-1, to the CRE half-site is highly affected by its genomic context. This effect was highly correlated with ATF1 ChIP-seq and PBM experiments. Next, we characterized the affinities of ATF1 and ATF3 to 128 genomic CRE and CRE half-site sequences. Our affinity measurements explained that in vivo binding differences between ATF1 and ATF3 to CRE and CRE half-sites are partially mediated by differences in the minor groove width. We believe that QPID would become a central tool for quantitative characterization of biophysical aspects affecting protein-DNA binding.

  16. High affinity binding of proteins HMG1 and HMG2 to semicatenated DNA loops

    Directory of Open Access Journals (Sweden)

    Strauss François

    2000-10-01

    Full Text Available Abstract Background Proteins HMG1 and HMG2 are two of the most abundant non histone proteins in the nucleus of mammalian cells, and contain a domain of homology with many proteins implicated in the control of development, such as the sex-determination factor Sry and the Sox family of proteins. In vitro studies of interactions of HMG1/2 with DNA have shown that these proteins can bind to many unusual DNA structures, in particular to four-way junctions, with binding affinities of 107 to 109 M-1. Results Here we show that HMG1 and HMG2 bind with a much higher affinity, at least 4 orders of magnitude higher, to a new structure, Form X, which consists of a DNA loop closed at its base by a semicatenated DNA junction, forming a DNA hemicatenane. The binding constant of HMG1 to Form X is higher than 5 × 1012 M-1, and the half-life of the complex is longer than one hour in vitro. Conclusions Of all DNA structures described so far with which HMG1 and HMG2 interact, we have found that Form X, a DNA loop with a semicatenated DNA junction at its base, is the structure with the highest affinity by more than 4 orders of magnitude. This suggests that, if similar structures exist in the cell nucleus, one of the functions of these proteins might be linked to the remarkable property of DNA hemicatenanes to associate two distant regions of the genome in a stable but reversible manner.

  17. Determination of equilibrium constant of amino carbamate adduct formation in sisomicin by a high pH based high performance liquid chromatography.

    Science.gov (United States)

    Wlasichuk, Kenneth B; Tan, Li; Guo, Yushen; Hildebrandt, Darin J; Zhang, Hao; Karr, Dane E; Schmidt, Donald E

    2015-01-01

    Amino carbamate adduct formation from the amino group of an aminoglycoside and carbon dioxide has been postulated as a mechanism for reducing nephrotoxicity in the aminoglycoside class compounds. In this study, sisomicin was used as a model compound for amino carbamate analysis. A high pH based reversed-phase high performance liquid chromatography (RP-HPLC) method is used to separate the amino carbamate from sisomicin. The carbamate is stable as the breakdown is inhibited at high pH and any reactive carbon dioxide is removed as the carbonate. The amino carbamate was quantified and the molar fraction of amine as the carbamate of sisomicin was obtained from the HPLC peak areas. The equilibrium constant of carbamate formation, Kc, was determined to be 3.3 × 10(-6) and it was used to predict the fraction of carbamate over the pH range in a typical biological systems. Based on these results, the fraction of amino carbamate at physiological pH values is less than 13%, and the postulated mechanism for nephrotoxicity protection is not valid. The same methodology is applicable for other aminoglycosides. Copyright © 2015 Elsevier B.V. All rights reserved.

  18. ACYLTRANSFERASE ACTIVITIES OF THE HIGH-MOLECULAR-MASS ESSENTIAL PENICILLIN-BINDING PROTEINS

    NARCIS (Netherlands)

    ADAM, M; DAMBLON, C; JAMIN, M; ZORZI, W; DUSART, [No Value; GALLENI, M; ELKHARROUBI, A; PIRAS, G; SPRATT, BG; KECK, W; COYETTE, J; GHUYSEN, JM; NGUYENDISTECHE, M; FRERE, JM

    1991-01-01

    The high-molecular-mass penicillin-binding proteins (HMM-PBPs), present in the cytoplasmic membranes of all eubacteria, are involved in important physiological events such as cell elongation, septation or shape determination. Up to now it has, however, been very difficult or impossible to study the

  19. Haptoglobin-related protein is a high-affinity hemoglobin-binding plasma protein

    DEFF Research Database (Denmark)

    Nielsen, Marianne Jensby; Petersen, Steen Vang; Jacobsen, Christian

    2006-01-01

    Haptoglobin-related protein (Hpr) is a primate-specific plasma protein associated with apolipoprotein L-I (apoL-I)-containing high-density lipoprotein (HDL) particles shown to be a part of the innate immune defense. Despite the assumption hitherto that Hpr does not bind to hemoglobin, the present...

  20. Simple and Rapid Hollow Fiber Liquid Phase Microextraction Followed by High Performance Liquid Chromatography Method for Determination of Drug-protein Binding

    Institute of Scientific and Technical Information of China (English)

    XI Guo-chen; HU Shuang; BAI Xiao-hong

    2011-01-01

    A method was established using hollow fiber-liquid phase microextraction(HF-LPME) followed by high performance liquid chromatography(HPLC) to determine the concentration of the free(unbound) drug in the solution of the drug and protein.Measurements of drug-protein binding ratios and free drug concentrations were then analyzed with the Klotz equation to determine the equilibrium binding constant and number of binding sites for drug-protein interaction.The optimized method allows one to perform the efficient extraction and separation of free drug from protein-bound drug,protein,and other interfering substances.This approach was used to characterize the binding of the anticholinergic drugs atropine sulfate and scopolamine hydrobromide to proteins in human plasma and bovine serum albumin(BSA).The results demonstrate the utility of HF-LPME method for measuring free drug concentrations in protein-drug mixtures and determining the protein binding parameters of a pharmacologically important class of drugs.

  1. RPM3: a multifunctional microporous MOF with recyclable framework and high H2 binding energy.

    Science.gov (United States)

    Lan, Anjian; Li, Kunhao; Wu, Haohan; Kong, Lingzhu; Nijem, Nour; Olson, David H; Emge, Thomas J; Chabal, Yves J; Langreth, David C; Hong, Maochun; Li, Jing

    2009-08-03

    A microporous metal organic framework structure, Zn(2)(bpdc)(2)(bpee).2DMF (DMF: N,N-dimethylformamide), has been synthesized via solvothermal reactions. The compound is a new member of the RPM series (RPM = Rutgers Recyclable Porous Material) that possesses a flexible and recyclable three-dimensional framework containing one-dimensional channels. It exhibits interesting and multifold functionality, including porosity, commensurate adsorption for hydrocarbons, high hydrogen binding energy (determined by isosteric heats of hydrogen adsorption and confirmed by van der Waals density functional calculations) as a result of multifold binding to aromatic ligands (determined by IR spectroscopy), strong photoluminescence emission, and reversible fluorescence quenching properties.

  2. A polyaromatic molecular tube that binds long hydrocarbons with high selectivity

    Science.gov (United States)

    Yazaki, Kohei; Sei, Yoshihisa; Akita, Munetaka; Yoshizawa, Michito

    2014-10-01

    Long hydrocarbon chains are essential components of biomolecules used for structure and function in living organisms. The selective recognition and effective binding of hydrocarbons within synthetic host compounds are problematic owing to their conformational flexibility and the lack of specific binding sites. Here we report a molecular tube with polyaromatic frameworks prepared by the Zincke cross-coupling reaction. The tube has a well-defined cylindrical cavity with a diameter and length of ~1 nm encircled by multiple anthracene panels and thereby binds long hydrocarbons containing branched methyl groups and/or unsaturated carbon-carbon double bonds (for example, heptamethylnonane, nervonic acid ester and squalene) with high selectivity in aqueous solutions.

  3. The anti-cancer IgM monoclonal antibody PAT-SM6 binds with high avidity to the unfolded protein response regulator GRP78.

    Directory of Open Access Journals (Sweden)

    Zachary Rosenes

    Full Text Available The monoclonal IgM antibody PAT-SM6 derived from human tumours induces apoptosis in tumour cells and is considered a potential anti-cancer agent. A primary target for PAT-SM6 is the unfolded protein response regulator GRP78, over-expressed externally on the cell surface of tumour cells. Small angle X-ray scattering (SAXS studies of human GRP78 showed a two-domain dumbbell-shaped monomer, while SAXS analysis of PAT-SM6 revealed a saucer-shaped structure accommodating five-fold symmetry, consistent with previous studies of related proteins. Sedimentation velocity analysis of GRP78 and PAT-SM6 mixtures indicated weak complex formation characterized by dissociation constants in the high micromolar concentration range. In contrast, enzyme-linked immunosorbant assays (ELISAs showed strong and specific interactions between PAT-SM6 and immobilized GRP78. The apparent binding constant estimated from a PAT-SM6 saturation curve correlated strongly with the concentration of GRP78 used to coat the microtiter tray. Experiments using polyclonal antiGRP78 IgG antibodies or a monoclonal IgG derivative of PAT-SM6 did not show a similar dependence. Competition experiments with soluble GRP78 indicated more effective inhibition of PAT-SM6 binding at low GRP78 coating concentrations. These observations suggest an avidity-based binding mechanism that depends on the multi-point attachment of PAT-SM6 to GRP78 clustered on the surface of the tray. Analysis of ELISA data at high GRP78 coating concentrations yielded an apparent dissociation constant of approximately 4 nM. We propose that the biological action of PAT-SM6 in tumour cell apoptosis may depend on the multivalent nature of PAT-SM6 and the high avidity of its interaction with multiple GRP78 molecules clustered on the tumour cell surface.

  4. Purification of high affinity benzodiazepine receptor binding site fragments from rat brain

    Energy Technology Data Exchange (ETDEWEB)

    Klotz, K.L.

    1984-01-01

    In central nervous system benzodiazepine recognition sites occur on neuronal cell surfaces as one member of a multireceptor complex, including recognition sites for benzodiazepines, gamma aminobutyric acid (GABA), barbiturates and a chloride ionophore. During photoaffinity labelling, the benzodiazepine agonist, /sup 3/H-flunitrazepam, is irreversibly bound to central benzodiazepine high affinity recognition sites in the presence of ultraviolet light. In these studies a /sup 3/H-flunitrazepam radiolabel was used to track the isolation and purification of high affinity agonist binding site fragments from membrane-bound benzodiazepine receptor in rat brain. The authors present a method for limited proteolysis of /sup 3/H-flunitrazepam photoaffinity labeled rat brain membranes, generating photolabeled benzodiazepine receptor fragments containing the agonist binding site. Using trypsin chymotrypsin A/sub 4/, or a combination of these two proteases, they have demonstrated the extent and time course for partial digestion of benzodiazepine receptor, yielding photolabeled receptor binding site fragments. These photolabeled receptor fragments have been further purified on the basis of size, using ultrafiltration, gel permeation chromatography, and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) as well as on the basis of hydrophobicity, using a high performance liquid chromatography (HPLC) precolumn, several HPLC elution schemes, and two different HPLC column types. Using these procedures, they have purified three photolabeled benzodiazepine receptor fragments containing the agonist binding site which appear to have a molecular weight of less than 2000 daltons each.

  5. Combinatorial Synthesis, Screening, and Binding Studies of Highly Functionalized Polyamino-amido Oligomers for Binding to Folded RNA

    Directory of Open Access Journals (Sweden)

    Jonathan K. Pokorski

    2012-01-01

    Full Text Available Folded RNA molecules have recently emerged as critical regulatory elements in biological pathways, serving not just as carriers of genetic information but also as key components in enzymatic assemblies. In particular, the transactivation response element (TAR of the HIV genome regulates transcriptional elongation by interacting specifically with the Tat protein, initiating the recruitment of the elongation complex. Preventing this interaction from occurring in vivo halts HIV replication, thus making RNA-binding molecules an intriguing pharmaceutical target. Using α-amino acids as starting materials, we have designed and synthesized a new class of polyamino-amido oligomers, called PAAs, specifically for binding to folded RNA structures. The PAA monomers were readily incorporated into a 125-member combinatorial library of PAA trimers. In order to rapidly assess RNA binding, a quantum dot-based fluorescent screen was developed to visualize RNA binding on-resin. The binding affinities of hits were quantified using a terbium footprinting assay, allowing us to identify a ligand (SFF with low micromolar affinity (kd=14 μM for TAR RNA. The work presented herein represents the development of a flexible scaffold that can be easily synthesized, screened, and subsequently modified to provide ligands specific for binding to folded RNAs.

  6. Elastic constants and Debye temperature of wz-AlN and wz-GaN semiconductors under high pressure from first-principles

    Indian Academy of Sciences (India)

    B P Pandey; V Kumar; Eduardo Menendez Proupin

    2014-09-01

    First-principles calculations were performed to study the elastic stiffness constants ($C_{ij}$) and Debye temperature ($_D$) of wurzite (wz) AlN and GaN binary semiconductors at high pressure. The lattice constants were calculated from the optimized structure of these materials. The band gaps were calculated at point using local density approximation (LDA) approach. The unit cell volume, lattice parameters, /, internal parameter (), elastic constant ($C_{ij}$), Debye temperature ($_D$), Hubbard parameter () and band gap ($E_g$) were studied under different pressures. The bulk modulus ($B_0$), reduced bulk modulus ($B'_0$) and Poisson ratio ($\\vee$) were also calculated. The calculated values of these parameters are in fair agreement with the available experimental and reported values.

  7. Possible effects of delivering methionine to broilers in drinking water at constant low and high environmental temperatures

    OpenAIRE

    Sahin Cadirci; Seyrani Koncagul

    2014-01-01

    An experiment was conducted to study the effects of water-soluble DL-methionine supplied through water and feed on the performance and carcass yield of broilers housed at two constant temperatures from 21 to 42 days of age. Birds were housed in two rooms (240 birds per room) with temperatures set at 21±2 and 30±2oC, respectively. Birds were divided into five groups of equal number within each room and fed five different diets (G1-G5). A low-methionine basal diet without supplemental methionin...

  8. High quality factor and high sensitivity photonic crystal rectangular holes slot nanobeam cavity with parabolic modulated lattice constant for refractive index sensing

    Science.gov (United States)

    Sun, Fujun; Zhou, Jian; Huang, Lijun; Fu, Zhongyuan; Tian, Huiping

    2017-09-01

    In this paper, we present a novel optical sensor based on photonic crystal slot nanobeam cavity (PCSNC) with rectangular air holes. By introducing a continuous slot and quadratically modulated hole spacing (lattice constant a) structure, the majority of the optical field is localized in the slot region, which enhances the light-matter interaction. With applying the three dimensional finite-difference time-domain (3D-FDTD) simulations, three key geometric parameters (hole width wx, slot width ws and the number of the holes N) are optimized to achieve a high sensitivity (S) while keeping a high quality (Q) factor. The highest S over 1000 nm/RIU (refractive index unit) is achieved when the slot width equals to 200 nm. The highest Q-factor of 2.15×107 is obtained when 30 holes are placed on both sides of the host waveguide with the slot width of 80 nm. Considering the transmission efficiency and the trade-off between S and Q-factor, the slot width and the number of the tapered region are chosen as 80 nm and 20, respectively. A high S approximately 835 nm/RIU and a Q-factor about 5.50×105 with small effective mode volume of 0.03(λ/nair)3 are achieved simultaneously, resulting in an ultra-high figure-of-merit (FOM) above 2.92×105. Furthermore, the active sensing region of the optimized structure occupies only about 12 μm×0.08 μm, which makes the device attractive for realizing on-chip integrated sensor arrays.

  9. Mivazerol, a novel compound with high specificity for alpha 2 adrenergic receptors: binding studies on different human and rat membrane preparations.

    Science.gov (United States)

    Noyer, M; de Laveleye, F; Vauquelin, G; Gobert, J; Wülfert, E

    1994-03-01

    Mivazerol, 3-[1(H-imidazol-4-yl)methyl]-2-hydroxybenzamide hydrochloride, a new potential anti-ischemic drug designed by UCB S.A. Pharma Sector, has been studied in binding experiments on adrenergic, dopaminergic, serotoninergic, muscarinic and idazoxan binding sites. Our results indicate that this compound displays high affinity and marked specificity for alpha 2 adrenoceptors. Mivazerol displaced the binding of the alpha 2 adrenoceptor antagonist [3H]RX 821002 to the alpha 2A adrenoceptors in human frontal cortex membranes with an apparent Ki value of 37 nM. The competition curve was shallow (nH = 0.55), suggesting that this compound acts as an alpha 2 adrenergic agonist. Mivazerol was also a potent competitor for [3H]RX 821002 binding to human platelet membranes (containing alpha 2A adrenoceptors) and rat kidney membranes (75% of the alpha 2 adrenoceptors of the alpha 2B subtype), indicating that this compound is not alpha 2 adrenoceptor subtype selective. Equilibrium dissociation constants for alpha 1 adrenoceptors (displacement of [3H]prazosin) and 5-HT1A receptors (displacement of [3H]rauwolscine) were respectively about 120 times (Ki = 4.4 microM) and 14 times (Ki = 530 nM) higher than that for the alpha 2 adrenoceptors. Equilibrium dissociation constants were approximately 1000 times higher for all other receptors tested in this study; namely beta 1 and beta 2 adrenoceptors, D1- and D2-dopamine receptors, M1-, M2- and M3-muscarinic receptors, 5-HT2 receptors and non-adrenergic idazoxan binding sites.

  10. Constant-pressure Blowers

    Science.gov (United States)

    Sorensen, E

    1940-01-01

    The conventional axial blowers operate on the high-pressure principle. One drawback of this type of blower is the relatively low pressure head, which one attempts to overcome with axial blowers producing very high pressure at a given circumferential speed. The Schicht constant-pressure blower affords pressure ratios considerably higher than those of axial blowers of conventional design with approximately the same efficiency.

  11. Crystallographically mapped ligand binding differs in high and low IgE binding isoforms of birch pollen allergen bet v 1.

    Science.gov (United States)

    Kofler, Stefan; Asam, Claudia; Eckhard, Ulrich; Wallner, Michael; Ferreira, Fátima; Brandstetter, Hans

    2012-09-07

    The ability of pathogenesis-related proteins of family 10 to bind a broad spectrum of ligands is considered to play a key role for their physiological and pathological functions. In particular, Bet v 1, an archetypical allergen from birch pollen, is described as a highly promiscuous ligand acceptor. However, the detailed recognition mechanisms, including specificity factors discriminating binding properties of naturally occurring Bet v 1 variants, are poorly understood. Here, we report crystal structures of Bet v 1 variants in complex with an array of ligands at a resolution of up to 1.2 Å. Residue 30 within the hydrophobic pocket not only discriminates in high and low IgE binding Bet v 1 isoforms but also induces a drastic change in the binding mode of the model ligand deoxycholate. Ternary crystal structure complexes of Bet v 1 with several ligands together with the fluorogenic reporter 1-anilino-8-naphthalene sulfonate explain anomalous fluorescence binding curves obtained from 1-anilino-8-naphthalene sulfonate displacement assays. The structures reveal key interaction residues such as Tyr83 and rationalize both the binding specificity and promiscuity of the so-called hydrophobic pocket in Bet v 1. The intermolecular interactions of Bet v 1 reveal an unexpected complexity that will be indispensable to fully understand its roles within the physiological and allergenic context.

  12. The Nucleotide-Free State of the Multidrug Resistance ABC Transporter LmrA: Sulfhydryl Cross-Linking Supports a Constant Contact, Head-to-Tail Configuration of the Nucleotide-Binding Domains.

    Directory of Open Access Journals (Sweden)

    Peter M Jones

    Full Text Available ABC transporters are integral membrane pumps that are responsible for the import or export of a diverse range of molecules across cell membranes. ABC transporters have been implicated in many phenomena of medical importance, including cystic fibrosis and multidrug resistance in humans. The molecular architecture of ABC transporters comprises two transmembrane domains and two ATP-binding cassettes, or nucleotide-binding domains (NBDs, which are highly conserved and contain motifs that are crucial to ATP binding and hydrolysis. Despite the improved clarity of recent structural, biophysical, and biochemical data, the seemingly simple process of ATP binding and hydrolysis remains controversial, with a major unresolved issue being whether the NBD protomers separate during the catalytic cycle. Here chemical cross-linking data is presented for the bacterial ABC multidrug resistance (MDR transporter LmrA. These indicate that in the absence of nucleotide or substrate, the NBDs come into contact to a significant extent, even at 4°C, where ATPase activity is abrogated. The data are clearly not in accord with an inward-closed conformation akin to that observed in a crystal structure of V. cholerae MsbA. Rather, they suggest a head-to-tail configuration 'sandwich' dimer similar to that observed in crystal structures of nucleotide-bound ABC NBDs. We argue the data are more readily reconciled with the notion that the NBDs are in proximity while undergoing intra-domain motions, than with an NBD 'Switch' mechanism in which the NBD monomers separate in between ATP hydrolysis cycles.

  13. High-accuracy Complete Active Space multiconfiguration Dirac-Hartree-Fock calculations of hyperfine structure constants of the gold atom

    CERN Document Server

    Bierón, Jacek; Indelicato, Paul; Jönsson, Per; Pyykkö, Pekka

    2009-01-01

    The multiconfiguration Dirac-Hartree-Fock (MCDHF) model has been employed to calculate the expectation values for the hyperfine splittings of the 5d96s2 2D3/2 and 5d96s2 2D5/2 levels of atomic gold. One-, two-, and three-body electron correlation effects involving all 79 electrons have been included in a systematic manner. The approximation employed in this study is equivalent to a Complete Active Space (CAS) approach. Calculated electric field gradients, together with experimental values of the electric quadrupole hyperfine structure constants, allow us to extract a nuclear electric quadrupole moment Q(197Au)=521.5(5.0) mb.

  14. Microwave assisted sol-gel synthesis of high dielectric constant CCTO and BFN ceramics for MLC applications

    Directory of Open Access Journals (Sweden)

    Sonia

    2017-06-01

    Full Text Available Ba(Fe1/2Nb1/2O3 (BFN and CaCu3Ti4O12 (CCTO ceramic powders were synthesized by microwave assisted sol-gel synthesis technique and sintered at 1100°C and 1000°C, respectively. Calcination and sintering processes were carried out in a microwave furnace. Dielectric constant (εr~2450 and dielectric loss (tan δ~0.5 at frequency of 1 kHz and 20°C were observed for the BFN ceramic samples. Higher value of εr ~ 3600 and lower value of tan δ ~ 0.07 at frequency of 1 kHz and in 20-60°C temperature range for the CCTO ceramic samples suggested its utility for MLC applications. Sharp decrease of εr and sharp increase of tan δ at higher frequencies of BFN ceramic samples indicated the presence of Debye like relaxation.

  15. Identification of fluorescent compounds with non-specific binding property via high throughput live cell microscopy.

    Directory of Open Access Journals (Sweden)

    Sangeeta Nath

    Full Text Available INTRODUCTION: Compounds exhibiting low non-specific intracellular binding or non-stickiness are concomitant with rapid clearing and in high demand for live-cell imaging assays because they allow for intracellular receptor localization with a high signal/noise ratio. The non-stickiness property is particularly important for imaging intracellular receptors due to the equilibria involved. METHOD: Three mammalian cell lines with diverse genetic backgrounds were used to screen a combinatorial fluorescence library via high throughput live cell microscopy for potential ligands with high in- and out-flux properties. The binding properties of ligands identified from the first screen were subsequently validated on plant root hair. A correlative analysis was then performed between each ligand and its corresponding physiochemical and structural properties. RESULTS: The non-stickiness property of each ligand was quantified as a function of the temporal uptake and retention on a cell-by-cell basis. Our data shows that (i mammalian systems can serve as a pre-screening tool for complex plant species that are not amenable to high-throughput imaging; (ii retention and spatial localization of chemical compounds vary within and between each cell line; and (iii the structural similarities of compounds can infer their non-specific binding properties. CONCLUSION: We have validated a protocol for identifying chemical compounds with non-specific binding properties that is testable across diverse species. Further analysis reveals an overlap between the non-stickiness property and the structural similarity of compounds. The net result is a more robust screening assay for identifying desirable ligands that can be used to monitor intracellular localization. Several new applications of the screening protocol and results are also presented.

  16. An integrated bioanalytical platform for supporting high-throughput serum protein binding screening.

    Science.gov (United States)

    Zhang, Jun; Shou, Wilson Z; Vath, Marianne; Kieltyka, Kasia; Maloney, Jennifer; Elvebak, Larry; Stewart, Jeremy; Herbst, John; Weller, Harold N

    2010-12-30

    Quantification of small molecules using liquid chromatography/tandem mass spectrometry (LC/MS/MS) on a triple quadrupole mass spectrometer has become a common practice in bioanalytical support of in vitro adsorption, distribution, metabolism and excretion (ADME) screening. The bioanalysis process involves primarily three indispensable steps: MS/MS optimization for a large number of new chemical compounds undergoing various screening assays in early drug discovery, high-throughput sample analysis with LC/MS/MS for those chemically diverse compounds using the optimized MS/MS conditions, and post-acquisition data review and reporting. To improve overall efficiency of ADME bioanalysis, an integrated system was proposed featuring an automated and unattended MS/MS optimization, a staggered parallel LC/MS/MS for high-throughput sample analysis, and a sophisticated software tool for LC/MS/MS raw data review as well as biological data calculation and reporting. The integrated platform has been used in bioanalytical support of a serum protein binding screening assay with high speed, high capacity, and good robustness. In this new platform, a unique sample dilution scheme was also introduced. With this dilution design, the total number of analytical samples was reduced; therefore, the total operation time was reduced and the overall throughput was further improved. The performance of the protein binding screening assay was monitored with two controls representing high and low binding properties and an acceptable inter-assay consistency was achieved. This platform has been successfully used for the determination of serum protein binding in multiple species for more than 4000 compounds.

  17. Neutrophil recruitment limited by high-affinity bent β2 integrin binding ligand in cis.

    Science.gov (United States)

    Fan, Zhichao; McArdle, Sara; Marki, Alex; Mikulski, Zbigniew; Gutierrez, Edgar; Engelhardt, Britta; Deutsch, Urban; Ginsberg, Mark; Groisman, Alex; Ley, Klaus

    2016-08-31

    Neutrophils are essential for innate immunity and inflammation and many neutrophil functions are β2 integrin-dependent. Integrins can extend (E(+)) and acquire a high-affinity conformation with an 'open' headpiece (H(+)). The canonical switchblade model of integrin activation proposes that the E(+) conformation precedes H(+), and the two are believed to be structurally linked. Here we show, using high-resolution quantitative dynamic footprinting (qDF) microscopy combined with a homogenous conformation-reporter binding assay in a microfluidic device, that a substantial fraction of β2 integrins on human neutrophils acquire an unexpected E(-)H(+) conformation. E(-)H(+) β2 integrins bind intercellular adhesion molecules (ICAMs) in cis, which inhibits leukocyte adhesion in vitro and in vivo. This endogenous anti-inflammatory mechanism inhibits neutrophil aggregation, accumulation and inflammation.

  18. High-Dose Mannose-Binding Lectin Therapy for Ebola Virus Infection

    Science.gov (United States)

    2010-06-01

    are heavily glycosylated and contain high-mannose. As a result, MBL binds to Ebola and Marburg viruses and mediates com- plement-dependent virus ...host cells. Therefore, MBL preferentially recognizes glycosylated viruses including influenza virus , human immunodeficiency virus , severe acute...respiratory syndrome coronovirus (SARS-CoV), Ebola virus , and Marburg virus . It also recognizes many glycosylated gram- positive and gram-negative bacteria [1

  19. New perovskite-related oxides having high dielectric constant: Ln2Ba2CaZn2Ti3O14 (Ln = La and Pr)

    Indian Academy of Sciences (India)

    Pika Jha; Ashok K Ganguli

    2003-10-01

    Two new oxides, La2Ba2CaZn2Ti3O14 and Pr2Ba2CaZn2Ti3O14, have been synthesized by the ceramic route at 1100°C. These oxides crystallize in the disordered cubic structure with an `a’ lattice parameter of 3.9728 (2) and 3.9448 (5) respectively. These oxides show high dielectric constant (70 and 57) and low loss (0.003 and 0.013 at 100 kHz) for La2Ba2CaZn2Ti)3O14 and Pr2Ba2CaZn2Ti3O14 respectively. The dielectric constant is highly stable with frequency and temperature.

  20. Design of numerical constant current source with high stability%高稳定性数控恒流源设计

    Institute of Scientific and Technical Information of China (English)

    王波; 戴曙光

    2016-01-01

    In order to improve the signal-to-noise ratio of the measurement,a constant current source of high stability is usually required,and to extent the measuring range,the constant current source can be used for adjusting and controlling,Therefore,this paper presents a highly stable constant current source method.Through the differential amplification and feedback of the voltage on the sampling resistor,on the one hand the common mode noise can be reduced and the accuracy of feedback can be improved,on the other hand also the heat consumption of the sampling resistor can be reduced,these aspects provide the guarantee for the constant current source of long time stability,in addition,the constant current source can be linearly adj usted by a DAC.After a long period of testing,the output ripple of the current is less than 0.1mA,and under different voltage the error of theory and actual in about one thousandth.So the constant current source is of high stability,and high control linearity.%在阻抗的测量过程中,为了提高测量的信噪比,通常需要使用高稳定性的恒流源,同时为了扩展测量的范围,也需要对恒流源进行控制调节,因此本文提出了一种高稳定性数控恒流源的设计方法。本文的恒流源通过对采样电阻端的电压进行差分放大反馈,一方面降低了共模噪声的干扰,提高了反馈精度,另一方面也降低了采样电阻端的热耗,为整个恒流源的长时间稳定性提供了保障,并且通过DAC可以实现对电流的线性调节。经过长时间的测试,该恒流源的输出电流的纹波小于0.1 mA,不同电压下理论电流与实际电流误差在千分之一左右,具有较高的稳定性以及高的控制线性度等特点。

  1. A Highly Expressed High-Molecular-Weight S-Layer Complex of Pelosinus sp. Strain UFO1 Binds Uranium

    Energy Technology Data Exchange (ETDEWEB)

    Thorgersen, Michael P. [Univ. of Georgia, Athens, GA (United States). Dept. of Biochemistry and Molecular Biology; Lancaster, W. Andrew [Univ. of Georgia, Athens, GA (United States). Dept. of Biochemistry and Molecular Biology; Rajeev, Lara [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Biological Systems and Engineering Division; Ge, Xiaoxuan [Univ. of Georgia, Athens, GA (United States). Dept. of Biochemistry and Molecular Biology; Vaccaro, Brian J. [Univ. of Georgia, Athens, GA (United States). Dept. of Biochemistry and Molecular Biology; Poole, Farris L. [Univ. of Georgia, Athens, GA (United States). Dept. of Biochemistry and Molecular Biology; Arkin, Adam P. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Biological Systems and Engineering Division; Mukhopadhyay, Aindrila [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Biological Systems and Engineering Division; Adams, Michael W. W. [Univ. of Georgia, Athens, GA (United States). Dept. of Biochemistry and Molecular Biology

    2016-12-02

    Cell suspensions of Pelosinus sp. strain UFO1 were previously shown, using spectroscopic analysis, to sequester uranium as U(IV) complexed with carboxyl and phosphoryl group ligands on proteins. The goal of our present study was to characterize the proteins involved in uranium binding. Virtually all of the uranium in UFO1 cells was associated with a heterodimeric protein, which was termed the uranium-binding complex (UBC). The UBC was composed of two S-layer domain proteins encoded by UFO1_4202 and UFO1_4203. Samples of UBC purified from the membrane fraction contained 3.3 U atoms/heterodimer, but significant amounts of phosphate were not detected. The UBC had an estimated molecular mass by gel filtration chromatography of 15 MDa, and it was proposed to contain 150 heterodimers (UFO1_4203 and UFO1_4202) and about 500 uranium atoms. The UBC was also the dominant extracellular protein, but when purified from the growth medium, it contained only 0.3 U atoms/heterodimer. The two genes encoding the UBC were among the most highly expressed genes within the UFO1 genome, and their expressions were unchanged by the presence or absence of uranium. Therefore, the UBC appears to be constitutively expressed and is the first line of defense against uranium, including by secretion into the extracellular medium. Although S-layer proteins were previously shown to bind U(VI), here we showed that U(IV) binds to S-layer proteins, we identified the proteins involved, and we quantitated the amount of uranium bound. Widespread uranium contamination from industrial sources poses hazards to human health and to the environment. Here in this paper, we identified a highly abundant uranium-binding complex (UBC) from Pelosinus sp. strain UFO1. The complex makes up the primary protein component of the S-layer of strain UFO1 and binds 3.3 atoms of U(IV) per heterodimer. Finally, while other bacteria have been shown to bind U(VI) on their S-layer, we demonstrate here an example of U(IV) bound by

  2. Synthesis and characterization of flexible and high-temperature resistant polyimide aerogel with ultra-low dielectric constant

    Directory of Open Access Journals (Sweden)

    X. M. Zhang

    2016-10-01

    Full Text Available A polyimide (PI aerogel with excellent combined thermal and dielectric properties was successfully prepared by the polycondensation of 3,3′,4,4′-biphenyltetracarboxylic dianhydride (BPDA, 5-amino-2-(4-aminophenylbenzoxazole (APBO and octa(amino-phenylsilsesquioxane (OAPS crosslinker, followed by a supercritical carbon dioxide (scCO2 drying treatment. The developed PI aerogel exhibited an ultra-low dielectric constant (k of 1.15 at a frequency of 2.75 GHz, a volume resistivity of 5.45·1014 Ω·cm, and a dielectric strength of 132 kV/cm. The flexible PI aerogel exhibited an openpore microstructure consisting of three-dimensional network with tangled nanofibers morphology with a porosity of 85.6% (volume ratio, an average pore diameter of 19.2 nm, and a Brunauer-Emmet-Teller (BET surface area of 428.6 m2/g. In addition, the PI aerogel showed excellent thermal stability with a glass transition temperature (Tg of 358.3 °C, a 5% weight loss temperature over 500 °C, and a residual weight ratio of 66.7% at 750 °C in nitrogen.

  3. Histidine-rich glycoprotein binds fibrin(ogen) with high affinity and competes with thrombin for binding to the gamma'-chain.

    Science.gov (United States)

    Vu, Trang T; Stafford, Alan R; Leslie, Beverly A; Kim, Paul Y; Fredenburgh, James C; Weitz, Jeffrey I

    2011-09-01

    Histidine-rich glycoprotein (HRG) is an abundant protein that binds fibrinogen and other plasma proteins in a Zn(2+)-dependent fashion but whose function is unclear. HRG has antimicrobial activity, and its incorporation into fibrin clots facilitates bacterial entrapment and killing and promotes inflammation. Although these findings suggest that HRG contributes to innate immunity and inflammation, little is known about the HRG-fibrin(ogen) interaction. By immunoassay, HRG-fibrinogen complexes were detected in Zn(2+)-supplemented human plasma, a finding consistent with a high affinity interaction. Surface plasmon resonance determinations support this concept and show that in the presence of Zn(2+), HRG binds the predominant γ(A)/γ(A)-fibrinogen and the γ-chain elongated isoform, γ(A)/γ'-fibrinogen, with K(d) values of 9 nm. Likewise, (125)I-labeled HRG binds γ(A)/γ(A)- or γ(A)/γ'-fibrin clots with similar K(d) values when Zn(2+) is present. There are multiple HRG binding sites on fibrin(ogen) because HRG binds immobilized fibrinogen fragment D or E and γ'-peptide, an analog of the COOH terminus of the γ'-chain that mediates the high affinity interaction of thrombin with γ(A)/γ'-fibrin. Thrombin competes with HRG for γ'-peptide binding and displaces (125)I-HRG from γ(A)/γ'-fibrin clots and vice versa. Taken together, these data suggest that (a) HRG circulates in complex with fibrinogen and that the complex persists upon fibrin formation, and (b) by competing with thrombin for γ(A)/γ'-fibrin binding, HRG may modulate coagulation. Therefore, the HRG-fibrin interaction may provide a novel link between coagulation, innate immunity, and inflammation.

  4. Control of lipopolysaccharide-high density lipoprotein binding by acute phase protein(s).

    Science.gov (United States)

    Tobias, P S; Ulevitch, R J

    1983-10-01

    When Salmonella minnesota R595 lipopolysaccharide (LPS) is mixed with serum, the LPS eventually forms a complex with high density lipoprotein (HDL). Complex formation is conveniently followed by CsCl equilibrium density gradient centrifugation. When mixing 10 micrograms LPS with normal rabbit serum (NRS) at 37 degrees C in the presence of 20 mM EDTA, the half-life for LPS binding to HDL is typically 2 to 3 min. When the same experiment is performed with the use of acute phase rabbit serum (APRS; collected 24 hr post-induction with silver nitrate), the half-life for LPS binding to HDL is typically 40 to 100 min. Thus LPS binding to HDL occurs some 20- to 40-fold slower in APRS than in NRS. Two other phenomena have been found, the time dependencies of which correlate well with the time dependency of LPS binding to HDL in APRS. If LPS-APRS reaction mixtures are cooled to 4 degrees C shortly after mixing and are dialyzed against 2.5 mM HEPES, 15 mM NaCl, pH 7.4 buffer, LPS is recovered in the washed precipitates ("euglobulin precipitate") if, and only if, the LPS-HDL binding reaction is not complete. The amount of LPS in the precipitate correlates well with the amount of LPS that has not bound to HDL. The second phenomenon we observe is that the LPS-containing euglobulin precipitate prepared from LPS-acute phase serum reaction mixtures shortly after mixing also contains a protein, gp60, the concentration of which in the euglobulin precipitate correlates well with the amount of LPS in the precipitate. Thus three phenomena are kinetically well correlated in APRS: the degree of binding of LPS to HDL, the degree of appearance of LPS in a euglobulin fraction, and the concentration of protein gp60 in the euglobulin fraction. We were unable to precipitate gp60 from APRS in the absence of LPS, from APRS after the LPS has fully bound to HDL, or from normal serum in the presence or absence of LPS. The known properties of gp60 are not reminiscent of any other known acute phase

  5. Analysis of RNA folding and ligand binding by conventional and high-throughput calorimetry.

    Science.gov (United States)

    Sokoloski, Joshua E; Bevilacqua, Philip C

    2012-01-01

    Noncoding RNAs serve myriad functions in the cell, but their biophysical properties are not well understood. Calorimetry offers direct and label-free means for characterizing the ligand-binding and thermostability properties of these RNA. We apply two main types of calorimetry--isothermal titration calorimetry (ITC) and differential scanning calorimetry (DSC)--to the characterization of these functional RNA molecules. ITC can describe ligand binding in terms of stoichiometry, affinity, and heat (enthalpy), while DSC can provide RNA stability in terms of heat capacity, melting temperature, and folding enthalpy. Here, we offer detailed experimental protocols for studying such RNA systems with commercially available conventional and high-throughput ITC and DSC instruments.

  6. New Synthesis and Tritium Labeling of a Selective Ligand for Studying High-affinity γ-Hydroxybutyrate (GHB) Binding Sites

    OpenAIRE

    Vogensen, Stine B.; Marek, Aleš; Bay, Tina; Wellendorph, Petrine; Kehler, Jan; Bundgaard, Christoffer; Frølund, Bente; Pedersen, Martin H. F.; Clausen, Rasmus P.

    2013-01-01

    3-Hydroxycyclopent-1-enecarboxylic acid (HOCPCA, 1) is a potent ligand for the high-affinity GHB binding sites in the CNS. An improved synthesis of 1 together with a very efficient synthesis of [3H]-1 is described. The radiosynthesis employs in situ generated lithium trimethoxyborotritide. Screening of 1 against different CNS targets establishes a high selectivity and we demonstrate in vivo brain penetration. In vitro characterization of [3H]-1 binding shows high specificity to the high-affin...

  7. Adjuvant effects of mannose-binding lectin ligands on the immune response to infectious bronchitis vaccine in chickens with high or low serum mannose-binding lectin concentrations

    DEFF Research Database (Denmark)

    Kjærup, Rikke Munkholm; Dalgaard, Tina Sørensen; Norup, Liselotte Rothmann;

    2014-01-01

    in the pathogenesis of IBV infection and the production of IBV-specific antibodies, which may be exploited in optimising IBV vaccine strategies. The present study shows that MBL has the capability to bind to IBV in vitro. Chickens from two inbred lines (L10H and L10L) selected for high or low MBL serum concentrations...

  8. Theoretical study of high-pressure phase stability of NaZr2(PO4)3 via elastic constants and equation of state

    Science.gov (United States)

    Chinnappan, Ravi; Panigrahi, B. K.

    2017-03-01

    Phase stability of NaZr_2(PO_4)_3 has been studied through density functional theory calculations of elastic constants, equation of state and enthalpies. The changes in elastic constants as a function of pressure show that the ambient rhombohedral (Rbar{3}c) NaZr_2(PO_4)_3 becomes unstable above 8 GPa and this instability is driven by a softening of C_{44} elastic constant through one of the Born stability criteria. High-pressure equation-of-state and enthalpy calculations further show that the ambient rhombohedral (Rbar{3}c) structure transforms first into another rhombohedral ( R3) phase and subsequently to LiZr_2(PO_4)_3-type orthorhombic phase at pressures above 6 and 8 GPa respectively which are in agreement with a recent x-ray diffraction study. Analysis of interatomic distances show that LiZr_2(PO_4)_3-type orthorhombic structure allows for shorter Na-O and Zr-P bonds at high pressures which appears to enable strong bonding and stability. Calculated formation enthalpy and bulk modulus of the ambient phase of NaZr_2(PO_4)_3 are found to be in reasonable agreement with the respective experimental values.

  9. New Synthesis and Tritium Labeling of a Selective Ligand for Studying High-affinity γ-Hydroxybutyrate (GHB) Binding Sites

    Science.gov (United States)

    Vogensen, Stine B.; Marek, Aleš; Bay, Tina; Wellendorph, Petrine; Kehler, Jan; Bundgaard, Christoffer; Frølund, Bente; Pedersen, Martin H.F.; Clausen, Rasmus P.

    2013-01-01

    3-Hydroxycyclopent-1-enecarboxylic acid (HOCPCA, 1) is a potent ligand for the high-affinity GHB binding sites in the CNS. An improved synthesis of 1 together with a very efficient synthesis of [3H]-1 is described. The radiosynthesis employs in situ generated lithium trimethoxyborotritide. Screening of 1 against different CNS targets establishes a high selectivity and we demonstrate in vivo brain penetration. In vitro characterization of [3H]-1 binding shows high specificity to the high-affinity GHB binding sites. PMID:24053696

  10. New synthesis and tritium labeling of a selective ligand for studying high-affinity γ-hydroxybutyrate (GHB) binding sites.

    Science.gov (United States)

    Vogensen, Stine B; Marek, Aleš; Bay, Tina; Wellendorph, Petrine; Kehler, Jan; Bundgaard, Christoffer; Frølund, Bente; Pedersen, Martin H F; Clausen, Rasmus P

    2013-10-24

    3-Hydroxycyclopent-1-enecarboxylic acid (HOCPCA, 1) is a potent ligand for the high-affinity GHB binding sites in the CNS. An improved synthesis of 1 together with a very efficient synthesis of [(3)H]-1 is described. The radiosynthesis employs in situ generated lithium trimethoxyborotritide. Screening of 1 against different CNS targets establishes a high selectivity, and we demonstrate in vivo brain penetration. In vitro characterization of [(3)H]-1 binding shows high specificity to the high-affinity GHB binding sites.

  11. New Synthesis and Tritium Labeling of a Selective Ligand for Studying High-Affinity γ-Hydroxybutyrate (GHB) Binding Sites

    DEFF Research Database (Denmark)

    Vogensen, Stine B.; Marek, Ales; Bay, Tina

    2013-01-01

    3-Hydroxycyclopent-1-enecarboxylic acid (HOCPCA, 1) is a potent ligand for the high-affinity GHB binding sites in the CNS. An improved synthesis of 1 together with a very efficient synthesis of [3H]-1 is described. The radiosynthesis employs in situ generated lithium trimethoxyborotritide....... Screening of 1 against different CNS targets establishes a high selectivity, and we demonstrate in vivo brain penetration. In vitro characterization of [3H]-1 binding shows high specificity to the high-affinity GHB binding sites....

  12. Unique Physically Anchored Cryptographic Theoretical Calculation of the Fine-Structure Constant {\\alpha} Matching both the g/2 and Interferometric High-Precision Measurements

    CERN Document Server

    Rhodes, Charles Kirkham

    2010-01-01

    The fine-structure constant {\\alpha}, the dimensionless number that represents the strength of electromagnetic coupling in the limit of sufficiently low energy interactions, is the crucial fundamental physical parameter that governs a nearly limitless range of phenomena involving the interaction of radiation with materials. Ideally, the apparatus of physical theory should be competent to provide a calculational procedure that yields a quantitatively correct value for {\\alpha} and the physical basis for its computation. This study presents the first demonstration of an observationally anchored theoretical procedure that predicts a unique value for {\\alpha} that stands in full agreement with the best (~370 ppt) high-precision experimental determinations. In a directly connected cryptographic computation, the method that gives these results also yields the magnitude of the cosmological constant {\\Omega}{\\Lambda} in conformance with the observational data and the condition of perfect flatness ({\\Omega}{\\Lambda} +...

  13. Mismatch of dielectric constants at the interface of nanometer metal-oxide-semiconductor devices with high- gate dielectric impacts on the inversion charge density

    Indian Academy of Sciences (India)

    Ling-Feng Mao

    2011-04-01

    The comparison of the inversion electron density between a nanometer metal-oxidesemiconductor (MOS) device with high- gate dielectric and a SiO2 MOS device with the same equivalent oxide thickness has been discussed. A fully self-consistent solution of the coupled Schrödinger–Poisson equations demonstrates that a larger dielectric-constant mismatch between the gate dielectric and silicon substrate can reduce electron density in the channel of a MOS device under inversion bias. Such a reduction in inversion electron density of the channel will increase with increase in gate voltage. A reduction in the charge density implies a reduction in the inversion electron density in the channel of a MOS device. It also implies that a larger dielectric constant of the gate dielectric might result in a reduction in the source–drain current and the gate leakage current.

  14. High-Resolution Infrared Fourier-Transform Spectroscopy and Rotational Constants of the ν 4Band of BrNO 2

    Science.gov (United States)

    Orphal, J.; Frenzel, A.; Grothe, H.; Redlich, B.; Scheffler, D.; Willner, H.; Zetzsch, C.

    1998-09-01

    For the first time, high-resolution infrared gas-phase absorption spectra of the BrNO2molecule were recorded using a Fourier-transform spectrometer. In this paper, the ν4bands of the79BrNO2and81BrNO2isotopomers around 1670 cm-1are investigated. Although the spectra are highly congested, rotational and centrifugal distortion constants for the ground andv4= 1 states of79BrNO2and81BrNO2were determined. The results show that BrNO2is a planar molecule ofC2νsymmetry and confirm predictions from a recentab initiostudy.

  15. Binding the Mammalian High Mobility Group Protein AT-hook 2 to AT-Rich Deoxyoligonucleotides: Enthalpy-Entropy Compensation

    OpenAIRE

    2009-01-01

    HMGA2 is a DNA minor-groove binding protein. We previously demonstrated that HMGA2 binds to AT-rich DNA with very high binding affinity where the binding of HMGA2 to poly(dA-dT)2 is enthalpy-driven and to poly(dA)poly(dT) is entropy-driven. This is a typical example of enthalpy-entropy compensation. To further study enthalpy-entropy compensation of HMGA2, we used isothermal-titration-calorimetry to examine the interactions of HMGA2 with two AT-rich DNA hairpins: 5′-CCAAAAAAAAAAAAAAAGCCCCCGCTT...

  16. Space-charge-limited leakage current in high dielectric constant and ferroelectric thin films considering the field-dependent permittivity

    Science.gov (United States)

    Sun, J.; Zheng, X. J.; Yin, W.; Tang, M. H.; Li, W.

    2010-12-01

    Distinguishing from the traditional characterization on high-field leakage current density-voltage relationship, the field-dependent permittivity from the polarization derivative is used to solve the space-charge-limited conduction, and the simulated leakage current densities are compared with the previous experimental observations. The influences of the mobility, ferroelectric parameters, and film thickness on the leakage current densities are discussed. The results verify that the high-field quasi-Ohmic region observed experimentally may result from the field-dependent permittivity, and that the leakage current can be influenced by the ferroelectric polarization.

  17. Autoradiographic imaging and quantification of the high-affinity GHB binding sites in rodent brain using (3)H-HOCPCA

    DEFF Research Database (Denmark)

    Klein, A B; Bay, T; Villumsen, I S

    2016-01-01

    analogue, 3-hydroxycyclopent-1-enecarboxylic acid (HOCPCA) as a tritiated version ((3)H-HOCPCA) to radioactively label the specific GHB high-affinity binding site and gain further insight into the density, distribution and developmental profile of this protein. We show that, in low nanomolar concentrations......, (3)H-HOCPCA displays excellent signal-to-noise ratios using rodent brain autoradiography, which makes it a valuable ligand for anatomical quantification of native GHB binding site levels. Our data confirmed that (3)H-HOCPCA labels only the high-affinity specific GHB binding site, found in high...... density in cortical and hippocampal regions. The experiments revealed markedly stronger binding at pH 6.0 (Kd 73.8 nM) compared to pH 7.4 (Kd 2312 nM), as previously reported for other GHB radioligands but similar Bmax values. Using (3)H-HOCPCA we analyzed the GHB binding protein profile during mouse...

  18. Characteristics of DNA-binding proteins determine the biological sensitivity to high-linear energy transfer radiation

    NARCIS (Netherlands)

    H. Wang (Hong); X. Zhang (Xiangming); P. Wang (Ping); X. Yu (Xiaoyan); J. Essers (Jeroen); D.J. Chen (David); R. Kanaar (Roland); S. Takeda (Shiunichi); Y. Wang (Ya)

    2010-01-01

    textabstractNon-homologous end-joining (NHEJ) and homologous recombination repair (HRR), contribute to repair ionizing radiation (IR)-induced DNA double-strand breaks (DSBs). Mre11 binding to DNA is the first step for activating HRR and Ku binding to DNA is the first step for initiating NHEJ. High-l

  19. Amino Terminal Region of Dengue Virus NS4A Cytosolic Domain Binds to Highly Curved Liposomes.

    Science.gov (United States)

    Hung, Yu-Fu; Schwarten, Melanie; Hoffmann, Silke; Willbold, Dieter; Sklan, Ella H; Koenig, BerndW

    2015-07-21

    Dengue virus (DENV) is an important human pathogen causing millions of disease cases and thousands of deaths worldwide. Non-structural protein 4A (NS4A) is a vital component of the viral replication complex (RC) and plays a major role in the formation of host cell membrane-derived structures that provide a scaffold for replication. The N-terminal cytoplasmic region of NS4A(1-48) is known to preferentially interact with highly curved membranes. Here, we provide experimental evidence for the stable binding of NS4A(1-48) to small liposomes using a liposome floatation assay and identify the lipid binding sequence by NMR spectroscopy. Mutations L6E;M10E were previously shown to inhibit DENV replication and to interfere with the binding of NS4A(1-48) to small liposomes. Our results provide new details on the interaction of the N-terminal region of NS4A with membranes and will prompt studies of the functional relevance of the curvature sensitive membrane anchor at the N-terminus of NS4A.

  20. Binding the mammalian high mobility group protein AT-hook 2 to AT-rich deoxyoligonucleotides: enthalpy-entropy compensation.

    Science.gov (United States)

    Joynt, Suzanne; Morillo, Victor; Leng, Fenfei

    2009-05-20

    HMGA2 is a DNA minor-groove binding protein. We previously demonstrated that HMGA2 binds to AT-rich DNA with very high binding affinity where the binding of HMGA2 to poly(dA-dT)(2) is enthalpy-driven and to poly(dA)poly(dT) is entropy-driven. This is a typical example of enthalpy-entropy compensation. To further study enthalpy-entropy compensation of HMGA2, we used isothermal-titration-calorimetry to examine the interactions of HMGA2 with two AT-rich DNA hairpins: 5'-CCAAAAAAAAAAAAAAAGCCCCCGCTTTTTTTTTTTTTTTGG-3' (FL-AT-1) and 5'-CCATATATATATATATAGCCCCCGCTATATATATATATATGG-3' (FL-AT-2). Surprisingly, we observed an atypical isothermal-titration-calorimetry-binding curve at low-salt aqueous solutions whereby the apparent binding-enthalpy decreased dramatically as the titration approached the end. This unusual behavior can be attributed to the DNA-annealing coupled to the ligand DNA-binding and is eliminated by increasing the salt concentration to approximately 200 mM. At this condition, HMGA2 binding to FL-AT-1 is entropy-driven and to FL-AT-2 is enthalpy-driven. Interestingly, the DNA-binding free energies for HMGA2 binding to both hairpins are almost temperature independent; however, the enthalpy-entropy changes are dependent on temperature, which is another aspect of enthalpy-entropy compensation. The heat capacity change for HMGA2 binding to FL-AT-1 and FL-AT-2 are almost identical, indicating that the solvent displacement and charge-charge interaction in the coupled folding/binding processes for both binding reactions are similar.

  1. On-column entrapment of alpha1-acid glycoprotein for studies of drug-protein binding by high-performance affinity chromatography.

    Science.gov (United States)

    Anguizola, Jeanethe; Bi, Cong; Koke, Michelle; Jackson, Abby; Hage, David S

    2016-08-01

    An on-column approach for protein entrapment was developed to immobilize alpha1-acid glycoprotein (AGP) for drug-protein binding studies based on high-performance affinity chromatography. Soluble AGP was physically entrapped by using microcolumns that contained hydrazide-activated porous silica and by employing mildly oxidized glycogen as a capping agent. Three on-column entrapment methods were evaluated and compared to a previous slurry-based entrapment method. The final selected method was used to prepare 1.0 cm × 2.1 mm I.D. affinity microcolumns that contained up to 21 (±4) μg AGP and that could be used over the course of more than 150 sample applications. Frontal analysis and zonal elution studies were performed on these affinity microcolumns to examine the binding of various drugs with the entrapped AGP. Site-selective competition studies were also conducted for these drugs. The results showed good agreement with previous observations for these drug-protein systems and with binding constants that have been reported in the literature. The entrapment method developed in this study should be useful for future work in the area of personalized medicine and in the high-throughput screening of drug interactions with AGP or other proteins. Graphical abstract On-column protein entrapment using a hydrazide-activated support and oxidized glycogen as a capping agent.

  2. 一种小功率高效恒流电源设计%A Low Power and High Efficiency Constant Current Power

    Institute of Scientific and Technical Information of China (English)

    王林; 夏婷; 张立生; 王荃禹

    2015-01-01

    针对传统恒流电源转换效率较低的问题,采用带有自抖动功能的电流模式 PWM 控制器 SD4843P,通过反激拓扑结构、反馈网络的设计实现恒流驱动电源的高效转换,同时给出了具体参数、详细的设计依据和过程。通过样机实验验证在设计的工作范围内,实现了恒流电源的转化效率大于90%,同时具有优质电气性能。%In view of transfer efficiency of the traditional constant flow power source is low,using the current mode PWM controller SD4843P with jitter function,the high-efficiency transfer of constant current power is realized through the design of fly-back topology circuit and feedback network,the fly-back topology circuit and feedback network have been giv-en the detailed design basis and the process of the concrete parameter.Experiments showed that the efficiency is greater than 90% in the design of the scope of work,and stable electrical performance is supplied by the constant current power.

  3. Vicinal 1H-1H NMR coupling constants from density functional theory as reliable tools for stereochemical analysis of highly flexible multichiral center molecules.

    Science.gov (United States)

    López-Vallejo, Fabian; Fragoso-Serrano, Mabel; Suárez-Ortiz, Gloria Alejandra; Hernández-Rojas, Adriana C; Cerda-García-Rojas, Carlos M; Pereda-Miranda, Rogelio

    2011-08-05

    A protocol for stereochemical analysis, based on the systematic comparison between theoretical and experimental vicinal (1)H-(1)H NMR coupling constants, was developed and applied to a series of flexible compounds (1-8) derived from the 6-heptenyl-5,6-dihydro-2H-pyran-2-one framework. The method included a broad conformational search, followed by geometry optimization at the DFT B3LYP/DGDZVP level, calculation of the vibrational frequencies, thermochemical parameters, magnetic shielding tensors, and the total NMR spin-spin coupling constants. Three scaling factors, depending on the carbon atom hybridizations, were found for the (1)H-C-C-(1)H vicinal coupling constants: f((sp3)-(sp3)) = 0.910, f((sp3)-(sp2)) = 0.929, and f((sp2)-(sp2))= 0.977. A remarkable correlation between the theoretical (J(pre)) and experimental (1)H-(1)H NMR (J(exp)) coupling constants for spicigerolide (1), a cytotoxic natural product, and some of its synthetic stereoisomers (2-4) demonstrated the predictive value of this approach for the stereochemical assignment of highly flexible compounds containing multiple chiral centers. The stereochemistry of two natural 6-heptenyl-5,6-dihydro-2H-pyran-2-ones (14 and 15) containing diverse functional groups in the heptenyl side chain was also analyzed by application of this combined theoretical and experimental approach, confirming its reliability. Additionally, a geometrical analysis for the conformations of 1-8 revealed that weak hydrogen bonds substantially guide the conformational behavior of the tetraacyloxy-6-heptenyl-2H-pyran-2-ones.

  4. Structure-affinity properties of a high-affinity ligand of FKBP12 studied by molecular simulations of a binding intermediate.

    Directory of Open Access Journals (Sweden)

    Lilian Olivieri

    Full Text Available With a view to explaining the structure-affinity properties of the ligands of the protein FKBP12, we characterized a binding intermediate state between this protein and a high-affinity ligand. Indeed, the nature and extent of the intermolecular contacts developed in such a species may play a role on its stability and, hence, on the overall association rate. To find the binding intermediate, a molecular simulation protocol was used to unbind the ligand by gradually decreasing the biasing forces introduced. The intermediate was subsequently refined with 17 independent stochastic boundary molecular dynamics simulations that provide a consistent picture of the intermediate state. In this state, the core region of the ligand remains stable, notably because of the two anchoring oxygen atoms that correspond to recurrent motifs found in all FKBP12 ligand core structures. Besides, the non-core regions participate in numerous transient intermolecular and intramolecular contacts. The dynamic aspect of most of the contacts seems important both for the ligand to retain at least a part of its configurational entropy and for avoiding a trapped state along the binding pathway. Since the transient and anchoring contacts contribute to increasing the stability of the intermediate, as a corollary, the dissociation rate constant [Formula: see text] of this intermediate should be decreased, resulting in an increase of the affinity constant [Formula: see text]. The present results support our previous conclusions and provide a coherent rationale for explaining the prevalence in high-affinity ligands of (i the two oxygen atoms found in carbonyl or sulfonyl groups of dissimilar core structures and of (ii symmetric or pseudo-symmetric mobile groups of atoms found as non-core moieties. Another interesting aspect of the intermediate is the distortion of the flexible 80 s loop of the protein, mainly in its tip region, that promotes the accessibility to the bound state.

  5. Stiff, strong, yet tough free-standing dielectric films of graphene nanosheets-polyurethane nanocomposites with very high dielectric constant and loss

    Science.gov (United States)

    Jan, Rahim; Habib, Amir; Gul, Iftikhar Hussain

    2016-01-01

    In this study, graphene nanosheets (GNS) prepared through a liquid exfoliation technique are dispersed in thermoplastic polyurethane (TPU) at a volume fraction (Vf) of up to 0.19. Then, the electrical and mechanical properties of the obtained composites are characterized. The dielectric spectroscopy shows an excessive variation in dielectric constant (1.1 to 3.53 × 107) and dielectric tangent loss (0.03 to 2515) with varying Vf over the frequency range of 25 kHz to 5 MHz. A considerable enhancement in electrical conductivity (DC) is found, from 3.87 × 10-10 S/m (base polymer) to 53.5 S/m for the 0.19 Vf GNS-TPU nanocomposite. The GNS-TPU composites are mechanically robust, with a considerable increase in stiffness (˜4-fold) and strength (almost twice), maintaining its ductility up to 0.09 Vf GNS. The high dielectric constant at lower frequencies is attributed to the well-established Maxwell-Wagner polarization effect, whereas the high dielectric tangent loss is due to leakage currents as a physical conducting network is formed at high filler loadings. The layered structure, high aspect ratio, and improved dispersion of GNS are the main reasons for the improvement in both the dielectric characteristics and the mechanical properties of the host polymer. [Figure not available: see fulltext.

  6. A Highly Expressed High-Molecular-Weight S-Layer Complex of Pelosinus sp. Strain UFO1 Binds Uranium.

    Science.gov (United States)

    Thorgersen, Michael P; Lancaster, W Andrew; Rajeev, Lara; Ge, Xiaoxuan; Vaccaro, Brian J; Poole, Farris L; Arkin, Adam P; Mukhopadhyay, Aindrila; Adams, Michael W W

    2017-02-15

    Cell suspensions of Pelosinus sp. strain UFO1 were previously shown, using spectroscopic analysis, to sequester uranium as U(IV) complexed with carboxyl and phosphoryl group ligands on proteins. The goal of our present study was to characterize the proteins involved in uranium binding. Virtually all of the uranium in UFO1 cells was associated with a heterodimeric protein, which was termed the uranium-binding complex (UBC). The UBC was composed of two S-layer domain proteins encoded by UFO1_4202 and UFO1_4203. Samples of UBC purified from the membrane fraction contained 3.3 U atoms/heterodimer, but significant amounts of phosphate were not detected. The UBC had an estimated molecular mass by gel filtration chromatography of 15 MDa, and it was proposed to contain 150 heterodimers (UFO1_4203 and UFO1_4202) and about 500 uranium atoms. The UBC was also the dominant extracellular protein, but when purified from the growth medium, it contained only 0.3 U atoms/heterodimer. The two genes encoding the UBC were among the most highly expressed genes within the UFO1 genome, and their expressions were unchanged by the presence or absence of uranium. Therefore, the UBC appears to be constitutively expressed and is the first line of defense against uranium, including by secretion into the extracellular medium. Although S-layer proteins were previously shown to bind U(VI), here we showed that U(IV) binds to S-layer proteins, we identified the proteins involved, and we quantitated the amount of uranium bound.

  7. Halophilic properties of metal binding protein characterized by high histidine content from Chromohalobacter salexigens DSM3043.

    Science.gov (United States)

    Yamaguchi, Rui; Arakawa, Tsutomu; Tokunaga, Hiroko; Ishibashi, Matsujiro; Tokunaga, Masao

    2012-02-01

    Periplasmic metal binding protein characterized by high histidine content was cloned from moderate halophile, Chromohalobacter salexigens. The protein, termed histidine-rich metal binding protein (HP), was expressed in and purified from E. coli as a native form. HP bound to Ni- and Cu-loaded chelate columns with high affinity, and Co- and Zn-columns with moderate affinity. Although the secondary structure was not grossly altered by the addition of 0.2-2.0 M NaCl, the thermal transition pattern was considerably shifted to higher temperature with increasing salt concentration: melting temperature was raised by ~20 °C at 2.0 M NaCl over the melting temperature at 0.2 M NaCl. HP showed reversible refolding from thermal melting in 0.2-1.15 M NaCl, while it formed irreversible aggregates upon thermal melting at 2 M NaCl. Addition of 0.01-0.1 mM NiSO₄ stabilized HP against thermal melting with high reversibility, while addition above 0.5 mM resulted in irreversible melting due to aggregation.

  8. The Power Control System of Constant Power Super-high Pressure Mercury Lamp%超高压汞灯功率控制研究

    Institute of Scientific and Technical Information of China (English)

    刘卫平; 葛劢冲

    2012-01-01

    A super-high pressure mercury lamp power of mask aligner is introduced in this paper,using analogue power feedback control,effectively improving power stability.The principle and major circuit structure of constant power control are emphatically expounded in this paper.%介绍了一种光刻机超高压汞灯电源,采用模拟功率反馈控制,有效提高了光源稳定性;重点阐述了恒功率控制的原理和主要电路结构。

  9. Electrical analysis of high dielectric constant insulator and metal gate metal oxide semiconductor capacitors on flexible bulk mono-crystalline silicon

    KAUST Repository

    Ghoneim, Mohamed T.

    2015-06-01

    We report on the electrical study of high dielectric constant insulator and metal gate metal oxide semiconductor capacitors (MOSCAPs) on a flexible ultra-thin (25 μm) silicon fabric which is peeled off using a CMOS compatible process from a standard bulk mono-crystalline silicon substrate. A lifetime projection is extracted using statistical analysis of the ramping voltage (Vramp) breakdown and time dependent dielectric breakdown data. The obtained flexible MOSCAPs operational voltages satisfying the 10 years lifetime benchmark are compared to those of the control MOSCAPs, which are not peeled off from the silicon wafer. © 2014 IEEE.

  10. The use of high resolution electron-energy-loss spectroscopy for refining the infrared optical constants of GaS, GaSe, and InSe

    Science.gov (United States)

    Yu, Li-Ming; Thiry, P. A.; Degiovanni, A.; Conard, Th.; Leclerc, G.; Caudano, R.; Lambin, Ph.; Debever, J.-M.

    1994-06-01

    Cleaved surfaces of III-VI lamellar semiconducting compounds GaS, GaSe, and InSe have been studied by high resolution electron-energy-loss spectroscopy (HREELS). The infrared optical constants of the materials were retrieved by using the dielectric theory taking account of the resonance frequencies published from infrared reflectivity (IRS) data. The limitations of the HREELS and IRS measurements in the case of these materials are discussed in detail. However, it is shown that, by combining the informations from both spectroscopies, it is possible to refine some of the oscillator strengths of these materials.

  11. High-affinity binding of fungal beta-glucan fragments to soybean (Glycine max L.) microsomal fractions and protoplasts.

    Science.gov (United States)

    Cosio, E G; Pöpperl, H; Schmidt, W E; Ebel, J

    1988-08-01

    We have recently reported the existence of binding sites in soybean membranes for a beta-glucan fraction derived from the fungal pathogen Phytophthora megasperma f. sp. glycinea, which may play a role in the elicitor-mediated phytoalexin response of this plant [Schmidt, W. E. & Ebel, J. (1987) Proc. Natl Acad. Sci. USA 84, 4117-4121]. The specificity of beta-glucan binding to soybean membranes has now been investigated using a variety of competing polyglucans and oligoglucans of fungal origin. P. megasperma beta-glucan binding showed high apparent affinity for branched glucans with degrees of polymerization greater than 12. Binding affinity showed good correlation with elicitor activity as measured in a soybean cotyledon bioassay. Modification of the glucans at the reducing end with phenylalkylamine reagents had no effect on binding affinity. This characteristic was used to synthesize an oligoglucosyl tyramine derivative suitable for radioiodination. The 125I-glucan (15-30 Ci/mmol) provided higher sensitivity and lower detection limits for the binding assays while behaving in a manner identical to the [3H]glucan used previously. More accurate determinations of the Kd value for glucan binding indicated a higher affinity than previously shown (37 nM versus 200 nM). The 125I-glucan was used to provide the first reported evidence of specific binding of a fungal beta-glucan fraction in vivo to soybean protoplasts. The binding affinity to protoplasts proved identical to that found in microsomal fractions.

  12. Nanoscale Synaptic Membrane Mimetic Allows Unbiased High Throughput Screen That Targets Binding Sites for Alzheimer's-Associated Aβ Oligomers.

    Directory of Open Access Journals (Sweden)

    Kyle C Wilcox

    Full Text Available Despite their value as sources of therapeutic drug targets, membrane proteomes are largely inaccessible to high-throughput screening (HTS tools designed for soluble proteins. An important example comprises the membrane proteins that bind amyloid β oligomers (AβOs. AβOs are neurotoxic ligands thought to instigate the synapse damage that leads to Alzheimer's dementia. At present, the identities of initial AβO binding sites are highly uncertain, largely because of extensive protein-protein interactions that occur following attachment of AβOs to surface membranes. Here, we show that AβO binding sites can be obtained in a state suitable for unbiased HTS by encapsulating the solubilized synaptic membrane proteome into nanoscale lipid bilayers (Nanodiscs. This method gives a soluble membrane protein library (SMPL--a collection of individualized synaptic proteins in a soluble state. Proteins within SMPL Nanodiscs showed enzymatic and ligand binding activity consistent with conformational integrity. AβOs were found to bind SMPL Nanodiscs with high affinity and specificity, with binding dependent on intact synaptic membrane proteins, and selective for the higher molecular weight oligomers known to accumulate at synapses. Combining SMPL Nanodiscs with a mix-incubate-read chemiluminescence assay provided a solution-based HTS platform to discover antagonists of AβO binding. Screening a library of 2700 drug-like compounds and natural products yielded one compound that potently reduced AβO binding to SMPL Nanodiscs, synaptosomes, and synapses in nerve cell cultures. Although not a therapeutic candidate, this small molecule inhibitor of synaptic AβO binding will provide a useful experimental antagonist for future mechanistic studies of AβOs in Alzheimer's model systems. Overall, results provide proof of concept for using SMPLs in high throughput screening for AβO binding antagonists, and illustrate in general how a SMPL Nanodisc system can

  13. Characterization of high affinity (/sup 3/H)triazolam binding in rat brain

    Energy Technology Data Exchange (ETDEWEB)

    Earle, M.; Concas, A.; Yamamura, H.I.

    1986-03-01

    The hypnotic Triazolam (TZ), a triazolo (1,4)-benzodiazepine, displays a short physiological half life and has been used for the treatment of insomnia related to anxiety states. Specific binding properties of this recently tritiated TZ were characterized. The authors major objectives were the direct measurement of the temperature dependence and the GABA effect on (/sup 3/H)TZ binding. Saturation studies showed a shift to lower affinity at 37/sup 0/C (K/sub d/ = 0.25 +/- 0.01 nM at O/sup 0/C; K/sub d/ = 1.46 +/- 0.03 nM at 37/sup 0/C) while the B/sub max/ values remained unchanged (1003 +/- 37 fmoles/mg prot. at 0/sup 0/C and 1001 +/- 43 fmoles/mg prot. at 37/sup 0/C). Inhibition studies showed that (/sup 3/H)TZ binding displayed no GABA shift at 0/sup 0/C(K/sub i/ 0.37 +/- 0.03 nM/- GABA and K/sub i/ = 0.55 +/- 0.13 nM/+GABA) but a nearly two-fold shift was apparent at 37/sup 0/C (K/sub i/ = 2.92 +/- 0.2 nM/-GABA; K/sub i/ = 1.37 +/- 0.11 mM/+GABA). These results were also confirmed by saturation studies in the presence or absence of GABA showing a shift to higher affinity in the presence of GABA only at 37/sup 0/C. In Ro 15-1788/(/sup 3/H)TZ competition experiments the presence of GABA did not affect the inhibitory potency of Ro 15-1788 on (/sup 3/H)TZ binding at both temperatures. In conclusion (/sup 3/H)TZ binding showed an extremely high affinity for benzodiazepine receptors. In contrast to reported literature, the findings suggest that TZ interacts with benzodiazepine receptors similar to other benzodiazepine agonists.

  14. Metal Decorated Multi-Walled Carbon Nanotube/Polyimide Composites with High Dielectric Constants and Low Loss Factors

    Science.gov (United States)

    Ghose, Sayata; Watson, Kent A.; Dudley, Kenneth L.; Elliott, Holly A.; Smith, Joseph G.; Connell, John W.

    2009-01-01

    The measurement of observable electromagnetic phenomena in materials and their derived intrinsic electric material properties are of prime importance in the discovery and development of material systems for electronic and aerospace applications. Nanocomposite materials comprised of metal decorated multi-walled carbon nanotubes (MWCNTs) were prepared by a facile method and characterized. Metal particles such as silver, platinum and palladium with diameters ranging from less than 5 to over 50 nanometers were distributed randomly on the MWCNTs. The metal-containing MWCNTs were then melt mixed into a polymer matrix and the mixture extruded as ribbons. These extruded ribbons exhibited a moderate to high degree of MWCNT alignment as determined by HRSEM. These ribbons were then fabricated into test specimens while maintaining MWCNT alignment and subsequently characterized for electromagnetic properties at 8-12 GHz. The present study is focused on silver decorated MWCNTs dispersed in an Ultem polyimide matrix. The results of the electromagnetic characterization showed that certain sample configurations exhibited a decoupling of the permittivity and loss factor (?? and ??) indicating that these properties could be tailored within certain limits. The decoupling and independent control of these fundamental electrical material parameters offer a new class of materials with potential applications in electronics, microwave engineering and optics.

  15. Spaces of constant curvature

    CERN Document Server

    Wolf, Joseph A

    2010-01-01

    This book is the sixth edition of the classic Spaces of Constant Curvature, first published in 1967, with the previous (fifth) edition published in 1984. It illustrates the high degree of interplay between group theory and geometry. The reader will benefit from the very concise treatments of riemannian and pseudo-riemannian manifolds and their curvatures, of the representation theory of finite groups, and of indications of recent progress in discrete subgroups of Lie groups. Part I is a brief introduction to differentiable manifolds, covering spaces, and riemannian and pseudo-riemannian geomet

  16. Nucleotide binding by the widespread high-affinity cyclic di-GMP receptor MshEN domain.

    Science.gov (United States)

    Wang, Yu-Chuan; Chin, Ko-Hsin; Tu, Zhi-Le; He, Jin; Jones, Christopher J; Sanchez, David Zamorano; Yildiz, Fitnat H; Galperin, Michael Y; Chou, Shan-Ho

    2016-01-01

    C-di-GMP is a bacterial second messenger regulating various cellular functions. Many bacteria contain c-di-GMP-metabolizing enzymes but lack known c-di-GMP receptors. Recently, two MshE-type ATPases associated with bacterial type II secretion system and type IV pilus formation were shown to specifically bind c-di-GMP. Here we report crystal structure of the MshE N-terminal domain (MshEN1-145) from Vibrio cholerae in complex with c-di-GMP at a 1.37 Å resolution. This structure reveals a unique c-di-GMP-binding mode, featuring a tandem array of two highly conserved binding motifs, each comprising a 24-residue sequence RLGxx(L/V/I)(L/V/I)xxG(L/V/I)(L/V/I)xxxxLxxxLxxQ that binds half of the c-di-GMP molecule, primarily through hydrophobic interactions. Mutating these highly conserved residues markedly reduces c-di-GMP binding and biofilm formation by V. cholerae. This c-di-GMP-binding motif is present in diverse bacterial proteins exhibiting binding affinities ranging from 0.5 μM to as low as 14 nM. The MshEN domain contains the longest nucleotide-binding motif reported to date.

  17. Low Frequency Noise Degradation in 45 nm High-k nMOSFETs due to Hot Carrier and Constant Voltage Stress

    Science.gov (United States)

    Rahman, M. Shahriar; ćelik-Butler, Zeynep; Quevedo-Lopez, M. A.; Shanware, Ajit; Colombo, Luigi

    2009-04-01

    Hafnium based materials are the leading candidates to replace conventional SiON as the gate dielectric in complementary metal-oxide-semiconductor devices. Hot carrier and constant voltage stress induced 1/f noise behavior is presented for HfSiON nMOSFETs. The additional low-frequency noise introduced through stressing was evaluated on nMOSFETs with TiN metal gate and HfSiON gate dielectric. Nitridation of HfSiO was achieved either by high temperature thermal nitridation or by relatively lower temperature plasma nitridation. The difference in stress induced noise behavior is attributed to the nitrogen profile across high-k/Si interface and bulk of high-k gate oxide caused by different nitridation techniques.

  18. Acylated heptapeptide binds albumin with high affinity and application as tag furnishes long-acting peptides

    Science.gov (United States)

    Zorzi, Alessandro; Middendorp, Simon J.; Wilbs, Jonas; Deyle, Kaycie; Heinis, Christian

    2017-07-01

    The rapid renal clearance of peptides in vivo limits this attractive platform for the treatment of a broad range of diseases that require prolonged drug half-lives. An intriguing approach for extending peptide circulation times works through a `piggy-back' strategy in which peptides bind via a ligand to the long-lived serum protein albumin. In accordance with this strategy, we developed an easily synthesized albumin-binding ligand based on a peptide-fatty acid chimera that has a high affinity for human albumin (Kd=39 nM). This ligand prolongs the elimination half-life of cyclic peptides in rats 25-fold to over seven hours. Conjugation to a peptide factor XII inhibitor developed for anti-thrombotic therapy extends the half-life from 13 minutes to over five hours, inhibiting coagulation for eight hours in rabbits. This high-affinity albumin ligand could potentially extend the half-life of peptides in human to several days, substantially broadening the application range of peptides as therapeutics.

  19. A novel and highly specific phage endolysin cell wall binding domain for detection of Bacillus cereus.

    Science.gov (United States)

    Kong, Minsuk; Sim, Jieun; Kang, Taejoon; Nguyen, Hoang Hiep; Park, Hyun Kyu; Chung, Bong Hyun; Ryu, Sangryeol

    2015-09-01

    Rapid, specific and sensitive detection of pathogenic bacteria is crucial for public health and safety. Bacillus cereus is harmful as it causes foodborne illness and a number of systemic and local infections. We report a novel phage endolysin cell wall-binding domain (CBD) for B. cereus and the development of a highly specific and sensitive surface plasmon resonance (SPR)-based B. cereus detection method using the CBD. The newly discovered CBD from endolysin of PBC1, a B. cereus-specific bacteriophage, provides high specificity and binding capacity to B. cereus. By using the CBD-modified SPR chips, B. cereus can be detected at the range of 10(5)-10(8) CFU/ml. More importantly, the detection limit can be improved to 10(2) CFU/ml by using a subtractive inhibition assay based on the pre-incubation of B. cereus and CBDs, removal of CBD-bound B. cereus, and SPR detection of the unbound CBDs. The present study suggests that the small and genetically engineered CBDs can be promising biological probes for B. cereus. We anticipate that the CBD-based SPR-sensing methods will be useful for the sensitive, selective, and rapid detection of B. cereus.

  20. High affinity binding site-mediated prevention of chemical absorption across the gastrointestinal tract.

    Science.gov (United States)

    Rasmussen, M V; Barker, T T; Silbart, L K

    2001-12-15

    Preventing mucosal absorption of low-molecular weight compounds such as carcinogens, toxins and drugs could help prevent many diseases. To characterize the effects of dose and timing on high-affinity binding site mediated sequestration of specific chemical ligands in the gastrointestinal tract, avidin was perorally-administered to mice either prior to or mixed with 3H-biotin. Avidin enhanced fecal 3H-biotin excretion in a dose-dependent manner, consistent with the accepted mechanism of egg white-induced biotin deficiency syndrome. Avidin administration up to 4 h before 3H-biotin administration also enhanced fecal 3H-biotin excretion. Activated charcoal (AC) reduced 3H-biotin absorption when mixed with 3H-biotin before ingestion, but was ineffective when ingested prior to 3H-biotin. These studies suggest that ingestion of high-affinity protein binding sites can establish an absorptive barrier at the gastrointestinal mucosa to prevent the uptake of unwanted low molecular-weight chemicals.

  1. An HIV-1 encoded peptide mimics the DNA binding loop of NF-{kappa}B and binds thioredoxin with high affinity

    Energy Technology Data Exchange (ETDEWEB)

    Su Guoping [Department of Pharmaceutical and Biomedical Sciences, College of Pharmacy, University of Georgia, Athens, GA 30602-2352 (United States)]. E-mail: gsu@u.washington.edu; Wang Min [Department of Pathology, Yale University School of Medicine, New Haven, CT 06520-8023 (United States)]. E-mail: wang.min@yale.edu; Taylor, Ethan Will [Department of Pharmaceutical and Biomedical Sciences, College of Pharmacy, University of Georgia, Athens, GA 30602-2352 (United States)]. E-mail: wtaylor@rx.uga.edu

    2005-11-11

    Pro-fs is a human immunodeficiency virus type 1 (HIV-l)-encoded putative selenoprotein, predicted by a theoretical analysis of the viral genome; it is potentially expressed by a -1 frameshift from the protease coding region. Pro-fs has significant sequence similarity to the DNA binding loop of nuclear factor kappa B (NF-{kappa}B), which is known to bind thioredoxin (Trx). We hypothesize that the putative HIV-1 pro-fs gene product functions by mimicry of NF-{kappa}B via binding to Trx. The hypothesis was tested in vitro by co-immunoprecipitation and GST-pull down assays, using a purified mutant pro-fs protein, in which the two potential selenocysteine residues were mutated to cysteines, in order to permit expression in bacteria. Both experiments showed that pro-fs binds to human wild type Trx (Trx-wt) with high affinity. Mutation of the two conserved cysteine residues in the Trx active site redox center to serine (Ser) (Trx-CS) weakened but failed to abolish the interaction. In pro-fs-transfected 293T cells, using confocal microscopy and fluorescence resonance energy transfer (FRET), we have observed that pro-fs localizes in cell nuclei and forms oligomers. Upon stimulation by phorbol 12-myristate 13-acetate (PMA), Trx translocates into cell nuclei. Significant FRET efficiency was detected in the nuclei of PMA-stimulated 293T cells co-expressing fluorescence-tagged pro-fs and Trx-wt or Trx-CS. These results indicate that in living cells the double cysteine mutant of pro-fs binds to both Trx and Trx-CS with high affinity, suggesting that Trx-pro-fs binding is a structurally-specific interaction, involving more of the Trx molecule than just its active site cysteine residues. These results establish the capacity for functional mimicry of the Trx binding ability of the NF-{kappa}B/Rel family of transcription factors by the putative HIV-1 pro-fs protein.

  2. High- and low-affinity binding of S-citalopram to the human serotonin transporter mutated at 20 putatively important amino acid positions

    DEFF Research Database (Denmark)

    Plenge, Per; Wiborg, Ove

    2005-01-01

    of presumed importance. Binding of S-citalopram, both to the high-affinity-binding site and to the allosteric binding site, was measured in these mutants with the purpose of investigating the connection between the two binding sites. The amino acid substitutions did not introduce large changes in the two...

  3. High resolution infrared synchrotron study of CH2D81Br: ground state constants and analysis of the ν5, ν6 and ν9 fundamentals

    Science.gov (United States)

    Baldacci, A.; Stoppa, P.; Visinoni, R.; Wugt Larsen, R.

    2012-09-01

    The high resolution infrared absorption spectrum of CH2D81Br has been recorded by Fourier transform spectroscopy in the range 550-1075 cm-1, with an unapodized resolution of 0.0025 cm-1, employing a synchrotron radiation source. This spectral region is characterized by the ν6 (593.872 cm-1), ν5 (768.710 cm-1) and ν9 (930.295 cm-1) fundamental bands. The ground state constants up to sextic centrifugal distortion terms have been obtained for the first time by ground-state combination differences from the three bands and subsequently employed for the evaluation of the excited state parameters. Watson's A-reduced Hamiltonian in the Ir representation has been used in the calculations. The ν 6 = 1 level is essentially free from perturbation whereas the ν 5 = 1 and ν 9 = 1 states are mutually interacting through a-type Coriolis coupling. Accurate spectroscopic parameters of the three excited vibrational states and a high-order coupling constant which takes into account the interaction between ν5 and ν9 have been determined.

  4. 高充电效率的恒流太阳能路灯控制器%Solar street lamp controller of high charging efficiency and constant current

    Institute of Scientific and Technical Information of China (English)

    向敏; 冉景海; 屈洪春

    2015-01-01

    针对传统太阳能路灯控制器主要存在充电效率低、恒流精度低的问题,提出基于多路脉冲宽度调制(PWM)恒流技术的新型太阳能路灯控制器的设计方案。该控制器采用复合式DC-DC变换器,通过控制多路 PWM 信号,实现高效地充放电功能。充电过程以改进的扰动观察法实现最大功率点跟踪(MPPT),提高充电效率;放电过程通过实时检测 LED 路灯的实际电流,动态调整相应 PWM 信号的占空比实现高精度恒流驱动 LED 路灯。实物测试结果表明该控制器能有效地改善充电效率和恒流源精度,充电速度提高了4%~7%,恒流源精度高达2.5%。%The traditional solar street lamp controller is constrained with the low charging efficiency and the low precision of the constant current source.A novel solar street lamp controller based on the multi-channel PWM constant current technology is designed.A composite DC-DC converter is adopted to realize charging and discharging efficiently through controlling multi-channel PWM signals.During the charging stage,the efficiency of charging is improved by tracking the maximum power point with the perturbation and observation method.During the discharging stage,the duty cycle of the relevant PWM signals are used to implement the high precision constant current source for driving the LED street lamp,which can be adjusted automatically through real-time detecting of the lamp’s actual current.The test results show that the charging efficiency of the street lamp controller can be improved evidently,and the charging speed increases 4% to 7%.Furthermore,the precision of the constant current source reaches a high level of 2.5%.

  5. High-throughput screening for novel inhibitors of Neisseria gonorrhoeae penicillin-binding protein 2.

    Directory of Open Access Journals (Sweden)

    Alena Fedarovich

    Full Text Available The increasing prevalence of N. gonorrhoeae strains exhibiting decreased susceptibility to third-generation cephalosporins and the recent isolation of two distinct strains with high-level resistance to cefixime or ceftriaxone heralds the possible demise of β-lactam antibiotics as effective treatments for gonorrhea. To identify new compounds that inhibit penicillin-binding proteins (PBPs, which are proven targets for β-lactam antibiotics, we developed a high-throughput assay that uses fluorescence polarization (FP to distinguish the fluorescent penicillin, Bocillin-FL, in free or PBP-bound form. This assay was used to screen a 50,000 compound library for potential inhibitors of N. gonorrhoeae PBP 2, and 32 compounds were identified that exhibited >50% inhibition of Bocillin-FL binding to PBP 2. These included a cephalosporin that provided validation of the assay. After elimination of compounds that failed to exhibit concentration-dependent inhibition, the antimicrobial activity of the remaining 24 was tested. Of these, 7 showed antimicrobial activity against susceptible and penicillin- or cephalosporin-resistant strains of N. gonorrhoeae. In molecular docking simulations using the crystal structure of PBP 2, two of these inhibitors docked into the active site of the enzyme and each mediate interactions with the active site serine nucleophile. This study demonstrates the validity of a FP-based assay to find novel inhibitors of PBPs and paves the way for more comprehensive high-throughput screening against highly resistant strains of N. gonorrhoeae. It also provides a set of lead compounds for optimization of anti-gonococcal agents.

  6. (Krauss) at constant high temperatures

    African Journals Online (AJOL)

    the various physical and chemical factors that may affect freshwater snails. However ... order to assess the effect of temperature on the organism, it is essential to ..... of snails by parasites is of cardinal importance to shed light on the population ...

  7. Research Advances on Polymer Matrix Composites with High Dielectric Constant%高介电聚合物基复合材料的研究进展

    Institute of Scientific and Technical Information of China (English)

    殷卫峰; 苏民社; 颜善银

    2013-01-01

    Application progress of high dielectric constant polymer matrix composites(HDPCs) at embedded capacitors, high energy storage, etc. is reviewed. Dielectric mechanism, research efforts of different types of HDPCs, effect of different factors on HDPCs properties are discussed. In addition, prospect of HDPCs is also presented.%综述了高介电聚合物基复合材料(HDPCs)在埋容器件、高能存储等方面的应用,探讨了HDPCs的高介电机理、不同类型HDPCs的研究状况以及不同因素对HDPCs性能的影响,并展望了HDPCs的前景.

  8. Reduction of the hydraulic retention time at constant high organic loading rate to reach the microbial limits of anaerobic digestion in various reactor systems.

    Science.gov (United States)

    Ziganshin, Ayrat M; Schmidt, Thomas; Lv, Zuopeng; Liebetrau, Jan; Richnow, Hans Hermann; Kleinsteuber, Sabine; Nikolausz, Marcell

    2016-10-01

    The effects of hydraulic retention time (HRT) reduction at constant high organic loading rate on the activity of hydrogen-producing bacteria and methanogens were investigated in reactors digesting thin stillage. Stable isotope fingerprinting was additionally applied to assess methanogenic pathways. Based on hydA gene transcripts, Clostridiales was the most active hydrogen-producing order in continuous stirred tank reactor (CSTR), fixed-bed reactor (FBR) and anaerobic sequencing batch reactor (ASBR), but shorter HRT stimulated the activity of Spirochaetales. Further decreasing HRT diminished Spirochaetales activity in systems with biomass retention. Based on mcrA gene transcripts, Methanoculleus and Methanosarcina were the predominantly active in CSTR and ASBR, whereas Methanosaeta and Methanospirillum activity was more significant in stably performing FBR. Isotope values indicated the predominance of aceticlastic pathway in FBR. Interestingly, an increased activity of Methanosaeta was observed during shortening HRT in CSTR and ASBR despite high organic acids concentrations, what was supported by stable isotope data.

  9. Comparison of the Folding Mechanism of Highly Homologous Proteins in the Lipid-binding Protein Family

    Science.gov (United States)

    The folding mechanism of two closely related proteins in the intracellular lipid binding protein family, human bile acid binding protein (hBABP) and rat bile acid binding protein (rBABP) were examined. These proteins are 77% identical (93% similar) in sequence Both of these singl...

  10. Comparison of the Folding Mechanism of Highly Homologous Proteins in the Lipid-binding Protein Family

    Science.gov (United States)

    The folding mechanism of two closely related proteins in the intracellular lipid binding protein family, human bile acid binding protein (hBABP) and rat bile acid binding protein (rBABP) were examined. These proteins are 77% identical (93% similar) in sequence Both of these singl...

  11. High density of benzodiazepine binding sites in the substantia innominata of the rat

    Energy Technology Data Exchange (ETDEWEB)

    Sarter, M.; Schneider, H.H.

    1988-07-01

    In order to study the neuronal basis of the pharmacological interactions between benzodiazepine receptor ligands and cortical cholinergic turnover, we examined the regional distribution of specific benzodiazepine binding sites using in vitro autoradiography. In the basal forebrain, the substantia innominata contained a high density of (/sup 3/H)lormetazepam (LMZ) binding sites (Bmax = 277 fmol/mg tissue; Kd = 0.55 nM). The label could be displaced by diazepam (IC50 = 100 nM), the benzodiazepine receptor antagonist beta-carboline ZK 93426 (45 nM) and the partial inverse agonist beta-carboline FG 7142 (540 nM). It is hypothesized that the amnesic effects of benzodiazepine receptor agonists are exerted through benzodiazepine receptors which are situated on cholinergic neurons in the substantia innominata and are involved in a tonic inhibition of cortical acetylcholine release. The benzodiazepine receptor antagonist ZK 93426 may exert its nootropic effects via benzodiazepine receptors in the substantia innominata and, consequently, by disinhibiting cortical acetylcholine release.

  12. A high-throughput, nonisotopic, competitive binding assay for kinases using nonselective inhibitor probes (ED-NSIP).

    Science.gov (United States)

    Vainshtein, Inna; Silveria, Scott; Kaul, Poonam; Rouhani, Riaz; Eglen, Richard M; Wang, John

    2002-12-01

    A novel competitive binding assay for protein kinase inhibitors has been developed for high-throughput screening (HTS). Unlike functional kinase assays, which are based on detection of substrate phosphorylation by the enzyme, this novel method directly measures the binding potency of compounds to the kinase ATP binding site through competition with a conjugated binding probe. The binding interaction is coupled to a signal amplification system based on complementation of beta-galactosidase enzyme fragments, a homogeneous, nonisotopic assay technology platform developed by DiscoveRx Corp. In the present study, staurosporine, a potent, nonselective kinase inhibitor, was chemically conjugated to a small fragment of beta-galactosidase (termed ED-SS). This was used as the binding probe to the kinase ATP binding pocket. The binding potencies of several inhibitors with diverse structures were assessed by displacement of ED-SS from the kinase. The assay format was specifically evaluated with GSK3alpha, an enzyme previously screened in a radioactive kinase assay (i.e., measurement of [(33)P]-gamma-ATP incorporation into the kinase peptide substrate). Under optimized assay conditions, nonconjugated staurosporine inhibited ED-SS binding in a concentration-dependent manner with an apparent potency (IC(50)) of 11 nM, which was similar to the IC(50) value determined in a radioactive assay. Furthermore, 9 kinase inhibitors with diverse structures, previously identified from chemical compound library screening, were screened using the competitive binding assay. The potencies in the binding assay were in very good agreement with those obtained previously in the isotopic functional activity assay. The binding assay was adapted for automated HTS using selected compound libraries in a 384-well microtiter plate format. The HTS assay was observed to be highly robust and reproducible (Z' factors > 0.7) with high interassay precision (R(2) > 0.96). Interference of compounds with the beta

  13. Is the empirical approximation Y/G approx constant applicable to high-pressure and high-temperature environments for metals?

    CERN Document Server

    Hua, J S; Hua, T; Hu, S L

    2002-01-01

    Recently, we have found, by means of a shock wave experiment, that an empirical relation Y/G approx 1.9x10 sup - sup 2 (Y is the yield strength and G is the shear modulus) is applicable for describing the strength effect for shocked 93W (93% W with 7% Fe-Ni-Co as binder) in the pressure range up to 150 GPa. This represents an extension of existing knowledge of the empirical approximation Y/G approx constant for potassium obtained at liquid-N sub 2 temperature and in the pressure range below 0.55 GPa. This approximation is advantageous in allowing one to simply and conveniently construct the constitutive equation for shocked metals.

  14. Very High Fuel Economy, Heavy Duty, Constant Speed, Truck Engine Optimized Via Unique Energy Recovery Turbines and Facilitated High Efficiency Continuously Variable Drivetrain

    Energy Technology Data Exchange (ETDEWEB)

    Bahman Habibzadeh

    2010-01-31

    The project began under a corporative agreement between Mack Trucks, Inc and the Department of Energy starting from September 1, 2005. The major objective of the four year project is to demonstrate a 10% efficiency gain by operating a Volvo 13 Litre heavy-duty diesel engine at a constant or narrow speed and coupled to a continuously variable transmission. The simulation work on the Constant Speed Engine started on October 1st. The initial simulations are aimed to give a basic engine model for the VTEC vehicle simulations. Compressor and turbine maps are based upon existing maps and/or qualified, realistic estimations. The reference engine is a MD 13 US07 475 Hp. Phase I was completed in May 2006 which determined that an increase in fuel efficiency for the engine of 10.5% over the OICA cycle, and 8.2% over a road cycle was possible. The net increase in fuel efficiency would be 5% when coupled to a CVT and operated over simulated highway conditions. In Phase II an economic analysis was performed on the engine with turbocompound (TC) and a Continuously Variable Transmission (CVT). The system was analyzed to determine the payback time needed for the added cost of the TC and CVT system. The analysis was performed by considering two different production scenarios of 10,000 and 60,000 units annually. The cost estimate includes the turbocharger, the turbocompound unit, the interstage duct diffuser and installation details, the modifications necessary on the engine and the CVT. Even with the cheapest fuel and the lowest improvement, the pay back time is only slightly more than 12 months. A gear train is necessary between the engine crankshaft and turbocompound unit. This is considered to be relatively straight forward with no design problems.

  15. Ion exchange equilibrium constants

    CERN Document Server

    Marcus, Y

    2013-01-01

    Ion Exchange Equilibrium Constants focuses on the test-compilation of equilibrium constants for ion exchange reactions. The book first underscores the scope of the compilation, equilibrium constants, symbols used, and arrangement of the table. The manuscript then presents the table of equilibrium constants, including polystyrene sulfonate cation exchanger, polyacrylate cation exchanger, polymethacrylate cation exchanger, polysterene phosphate cation exchanger, and zirconium phosphate cation exchanger. The text highlights zirconium oxide anion exchanger, zeolite type 13Y cation exchanger, and

  16. Immobilization of a thermostable alpha-amylase by covalent binding to an alginate matrix increases high temperature usability.

    Science.gov (United States)

    Tee, Boon L; Kaletunç, Gönül

    2009-01-01

    Thermostable alpha-amylase was covalently bound to calcium alginate matrix to be used for starch hydrolysis at liquefaction temperature of 95 degrees C. 1-ethyl-3-(3-dimethylamino-propyl) carbodiimide hydrochloride (EDAC) was used as crosslinker. EDAC reacts with the carboxylate groups on the calcium alginate matrix and the amine groups of the enzyme. Ethylenediamine tetraacetic acid (EDTA) treatment was applied to increase the number of available carboxylate groups on the calcium alginate matrix for EDAC binding. After the immobilization was completed, the beads were treated with 0.1 M calcium chloride solution to reinstate the bead mechanical strength. Enzyme loading efficiency, activity, and reusability of the immobilized alpha-amylase were investigated. Covalently bound thermostable alpha-amylase to calcium alginate produced a total of 53 g of starch degradation/mg of bound protein after seven consecutive starch hydrolysis cycles of 10 min each at 95 degrees C in a stirred batch reactor. The free and covalently bound alpha-amylase had maximum activity at pH 5.5 and 6.0, respectively. The Michaelis-Menten constant (K(m)) of the immobilized enzyme (0.98 mg/mL) was 2.5 times greater than that of the free enzyme (0.40 mg/mL). The maximum reaction rate (V(max)) of immobilized and free enzyme were determined to be 10.4-mg starch degraded/mL min mg bound protein and 25.7-mg starch degraded/mL min mg protein, respectively. The high cumulative activity and seven successive reuses obtained at liquefaction temperature make the covalently bound thermostable alpha-amylase to calcium alginate matrix, a promising candidate for use in industrial starch hydrolysis process.

  17. Autoradiographic imaging and quantification of the high-affinity GHB binding sites in rodent brain using (3)H-HOCPCA.

    Science.gov (United States)

    Klein, A B; Bay, T; Villumsen, I S; Falk-Petersen, C B; Marek, A; Frølund, B; Clausen, R P; Hansen, H D; Knudsen, G M; Wellendorph, P

    2016-11-01

    GHB (γ-hydroxybutyric acid) is a compound endogenous to mammalian brain with high structural resemblance to GABA. GHB possesses nanomolar-micromolar affinity for a unique population of binding sites, but the exact nature of these remains elusive. In this study we utilized the highly selective GHB analogue, 3-hydroxycyclopent-1-enecarboxylic acid (HOCPCA) as a tritiated version ((3)H-HOCPCA) to radioactively label the specific GHB high-affinity binding site and gain further insight into the density, distribution and developmental profile of this protein. We show that, in low nanomolar concentrations, (3)H-HOCPCA displays excellent signal-to-noise ratios using rodent brain autoradiography, which makes it a valuable ligand for anatomical quantification of native GHB binding site levels. Our data confirmed that (3)H-HOCPCA labels only the high-affinity specific GHB binding site, found in high density in cortical and hippocampal regions. The experiments revealed markedly stronger binding at pH 6.0 (Kd 73.8 nM) compared to pH 7.4 (Kd 2312 nM), as previously reported for other GHB radioligands but similar Bmax values. Using (3)H-HOCPCA we analyzed the GHB binding protein profile during mouse brain development. Due to the high sensitivity of this radioligand, we were able to detect low levels of specific binding already at E15 in mouse brain, which increased progressively until adulthood. Collectively, we show that (3)H-HOCPCA is a highly sensitive radioligand, offering advantages over the commonly used radioligand (3)H-NCS-382, and thus a very suitable in vitro tool for qualitative and quantitative autoradiography of the GHB high-affinity site.

  18. Chemotactic response with a constant delay-time mechanism in Ciona spermatozoa revealed by a high time resolution analysis of flagellar motility

    Directory of Open Access Journals (Sweden)

    Daisuke Miyashiro

    2015-01-01

    Full Text Available During their chemotactic swimming toward eggs, sperm cells detect their species-specific chemoattractant and sense concentration gradients by unknown mechanisms. After sensing the attractant, sperm cells commonly demonstrate a series of responses involving different swimming patterns by changing flagellar beats, gradually approaching a swimming path toward the eggs, which is the source of chemoattractants. Shiba et al. observed a rapid increase in intracellular Ca2+ concentrations in Ciona spermatozoa after sensing chemoattractants; however, the biochemical processes occurring inside the sperm cells are unclear. In the present study, we focused on the timing and sensing mechanism of chemical signal detection in Ciona. One of the most crucial problems to be solved is defining the initial epoch of chemotactic responses. We adopted a high rate of video recording (600 Hz for detailed analysis of sperm motion and a novel method for detecting subtle signs of beat forms and moving paths of sperm heads. From these analyses, we estimated a virtual sensing point of the attractant before initiation of motility responses and found that the time delay from sensing to motility responses was almost constant. To evaluate the efficiency of this constant delay model, we performed computer simulation of chemotactic behaviors of Ciona spermatozoa.

  19. Chemotactic response with a constant delay-time mechanism in Ciona spermatozoa revealed by a high time resolution analysis of flagellar motility

    Science.gov (United States)

    Miyashiro, Daisuke; Shiba, Kogiku; Miyashita, Tahahiro; Baba, Shoji A.; Yoshida, Manabu; Kamimura, Shinji

    2015-01-01

    ABSTRACT During their chemotactic swimming toward eggs, sperm cells detect their species-specific chemoattractant and sense concentration gradients by unknown mechanisms. After sensing the attractant, sperm cells commonly demonstrate a series of responses involving different swimming patterns by changing flagellar beats, gradually approaching a swimming path toward the eggs, which is the source of chemoattractants. Shiba et al. observed a rapid increase in intracellular Ca2+ concentrations in Ciona spermatozoa after sensing chemoattractants; however, the biochemical processes occurring inside the sperm cells are unclear. In the present study, we focused on the timing and sensing mechanism of chemical signal detection in Ciona. One of the most crucial problems to be solved is defining the initial epoch of chemotactic responses. We adopted a high rate of video recording (600 Hz) for detailed analysis of sperm motion and a novel method for detecting subtle signs of beat forms and moving paths of sperm heads. From these analyses, we estimated a virtual sensing point of the attractant before initiation of motility responses and found that the time delay from sensing to motility responses was almost constant. To evaluate the efficiency of this constant delay model, we performed computer simulation of chemotactic behaviors of Ciona spermatozoa. PMID:25572419

  20. Chemotactic response with a constant delay-time mechanism in Ciona spermatozoa revealed by a high time resolution analysis of flagellar motility.

    Science.gov (United States)

    Miyashiro, Daisuke; Shiba, Kogiku; Miyashita, Tahahiro; Baba, Shoji A; Yoshida, Manabu; Kamimura, Shinji

    2015-01-08

    During their chemotactic swimming toward eggs, sperm cells detect their species-specific chemoattractant and sense concentration gradients by unknown mechanisms. After sensing the attractant, sperm cells commonly demonstrate a series of responses involving different swimming patterns by changing flagellar beats, gradually approaching a swimming path toward the eggs, which is the source of chemoattractants. Shiba et al. observed a rapid increase in intracellular Ca(2+) concentrations in Ciona spermatozoa after sensing chemoattractants; however, the biochemical processes occurring inside the sperm cells are unclear. In the present study, we focused on the timing and sensing mechanism of chemical signal detection in Ciona. One of the most crucial problems to be solved is defining the initial epoch of chemotactic responses. We adopted a high rate of video recording (600 Hz) for detailed analysis of sperm motion and a novel method for detecting subtle signs of beat forms and moving paths of sperm heads. From these analyses, we estimated a virtual sensing point of the attractant before initiation of motility responses and found that the time delay from sensing to motility responses was almost constant. To evaluate the efficiency of this constant delay model, we performed computer simulation of chemotactic behaviors of Ciona spermatozoa.

  1. Effects of high CO2 on growth and metabolism of Arabidopsis seedlings during growth with a constantly limited supply of nitrogen.

    Science.gov (United States)

    Takatani, Nobuyuki; Ito, Takuro; Kiba, Takatoshi; Mori, Marie; Miyamoto, Tetsuro; Maeda, Shin-Ichi; Omata, Tatsuo

    2014-02-01

    Elevated CO2 has been reported to stimulate plant growth under nitrogen-sufficient conditions, but the effects of CO2 on growth in a constantly nitrogen-limited state, which is relevant to most natural habitats of plants, remain unclear. Here, we maintained Arabidopsis seedlings under such conditions by growing a mutant with reduced nitrate uptake activity on a medium containing nitrate as the sole nitrogen source. Under nitrogen-sufficient conditions (i.e. in the presence of ammonium), growth of shoots and roots of both the wild type (WT) and the mutant was increased approximately 2-fold by elevated CO2. Growth stimulation of shoots and roots by elevated CO2 was observed in the WT growing with nitrate as the sole nitrogen source, but in the mutant grown with nitrate, the high-CO2 conditions stimulated only the growth of roots. In the mutant, elevated CO2 caused well-known symptoms of nitrogen-starved plants, including decreased shoot/root ratio, reduced nitrate content and accumulation of anthocyanin, but also had an increased Chl content in the shoot, which was contradictory to the known effect of nitrogen depletion. A high-CO2-responsive change specific to the mutant was not observed in the levels of the major metabolites, although CO2 responses were observed in the WT and the mutant. These results indicated that elevated CO2 causes nitrogen limitation in the seedlings grown with a constantly limited supply of nitrogen, but the Chl content and the root biomass of the plant increase to enhance the activities of both photosynthesis and nitrogen uptake, while maintaining normal metabolism and response to high CO2.

  2. A complex water network contributes to high-affinity binding in an antibody–antigen interface

    Directory of Open Access Journals (Sweden)

    S.F. Marino

    2016-03-01

    Full Text Available This data article presents an analysis of structural water molecules in the high affinity interaction between a potent tumor growth inhibiting antibody (fragment, J22.9-xi, and the tumor marker antigen CD269 (B cell maturation antigen, BCMA. The 1.89 Å X-ray crystal structure shows exquisite details of the binding interface between the two molecules, which comprises relatively few, mostly hydrophobic, direct contacts but many indirect interactions over solvent waters. These are partly or wholly buried in, and therefore part of, the interface. A partial description of the structure is included in an article on the tumor inhibiting effects of the antibody: “Potent anti-tumor response by targeting B cell maturation antigen (BCMA in a mouse model of multiple myeloma”, Mol. Oncol. 9 (7 (2015 pp. 1348–58.

  3. A universal homogeneous assay for high-throughput determination of binding kinetics.

    Science.gov (United States)

    Schiele, Felix; Ayaz, Pelin; Fernández-Montalván, Amaury

    2015-01-01

    There is an increasing demand for assay technologies that enable accurate, cost-effective, and high-throughput measurements of drug-target association and dissociation rates. Here we introduce a universal homogeneous kinetic probe competition assay (kPCA) that meets these requirements. The time-resolved fluorescence energy transfer (TR-FRET) procedure combines the versatility of radioligand binding assays with the advantages of homogeneous nonradioactive techniques while approaching the time resolution of surface plasmon resonance (SPR) and related biosensors. We show application of kPCA for three important target classes: enzymes, protein-protein interactions, and G protein-coupled receptors (GPCRs). This method is capable of supporting early stages of drug discovery with large amounts of kinetic information.

  4. High sensitivity electron diffraction analysis. A study of divalent cation binding to purple membrane

    Energy Technology Data Exchange (ETDEWEB)

    Mitra, A.K.; Stroud, R.M. (Univ. of California, San Francisco (USA))

    1990-02-01

    A sensitive high-resolution electron diffraction assay for change in structure is described and harnessed to analyze the binding of divalent cations to the purple membrane (PM) of Halobacterium halobium. Low-dose electron diffraction patterns are subject to a matched filter algorithm. to extract accurate values of reflection intensities. This, coupled with a scheme to account for twinning and specimen tilt in the microscope, yields results that are sensitive enough to rapidly quantitate any structure change in PM brought about by site-directed mutagenesis to the level of less than two carbon atoms. Removal of tightly bound divalent cations (mainly Ca2+ and Mg2+) from PM causes a color change to blue and is accompanied by a severely altered photocycle of the protein bacteriohodopsin (bR), a light-driven proton pump. We characterize the structural changes that occur upon association of 3:1 divalent cation to PM, versus membranes rendered purple by addition of excess Na+. High resolution, low dose electron diffraction data obtained from glucose-embedded samples of Pb2+ and Na+ reconstituted PM preparations at room temperature identify several sites with total occupancy of 2.01 +/- 0.05 Pb2+ equivalents. The color transition as a function of ion concentration for Ca2+ or Mg2+ and Pb2+ are strictly comparable. A (Pb2(+)-Na+) PM Fourier difference map in projection was synthesized at 5 A using the averaged data from several nominally untilted patches corrected for twinning and specimen tilt. We find six major sites located on helices 7, 5, 4, 3, 2 in close association with bR. These partially occupied sites (0.55-0.24 Pb2+ equivalents) represent preferential sites of binding for divalent cations and complements our earlier result by x-ray diffraction.

  5. Identification and properties of very high affinity brain membrane-binding sites for a neurotoxic phospholipase from the taipan venom

    Energy Technology Data Exchange (ETDEWEB)

    Lambeau, G.; Barhanin, J.; Schweitz, H.; Qar, J.; Lazdunski, M. (Centre de Biochimie, Nice (France))

    1989-07-05

    Four new monochain phospholipases were purified from the Oxyuranus scutellatus (taipan) venom. Three of them were highly toxic when injected into mice brain. One of these neurotoxic phospholipases, OS2, was iodinated and used in binding experiments to demonstrate the presence of two families of specific binding sites in rat brain synaptic membranes. The affinities were exceptionally high, Kd1 = 1.5 +/- 0.5 pM and Kd2 = 45 +/- 10 pM, and the maximal binding capacities were Bmax 1 = 1 +/- 0.4 and Bmax 2 = 3 +/- 0.5 pmol/mg of protein. Both binding sites were sensitive to proteolysis and demonstrated to be located on proteins of Mr 85,000-88,000 and 36,000-51,000 by cross-linking and photoaffinity labeling techniques. The binding of {sup 125}I-OS2 to synaptic membranes was dependent on Ca2+ ions and enhanced by Zn2+ ions which inhibit phospholipase activity. Competition experiments have shown that, except for beta-bungarotoxin, a number of known toxic snake or bee phospholipases have very high affinities for the newly identified binding sites. A good correlation (r = 0.80) was observed between toxicity and affinity but not between phospholipase activity and affinity.

  6. Cosmological Constant, Fine Structure Constant and Beyond

    CERN Document Server

    Wei, Hao; Li, Hong-Yu; Xue, Dong-Ze

    2016-01-01

    In this work, we consider the cosmological constant model $\\Lambda\\propto\\alpha^{-6}$, which is well motivated from three independent approaches. As is well known, the evidence of varying fine structure constant $\\alpha$ was found in 1998. If $\\Lambda\\propto\\alpha^{-6}$ is right, it means that the cosmological constant $\\Lambda$ should be also varying. In this work, we try to develop a suitable framework to model this varying cosmological constant $\\Lambda\\propto\\alpha^{-6}$, in which we view it from an interacting vacuum energy perspective. We propose two types of models to describe the evolutions of $\\Lambda$ and $\\alpha$. Then, we consider the observational constraints on these models, by using the 293 $\\Delta\\alpha/\\alpha$ data from the absorption systems in the spectra of distant quasars, and the data of type Ia supernovae (SNIa), cosmic microwave background (CMB), baryon acoustic oscillation (BAO). We find that the model parameters can be tightly constrained to the narrow ranges of ${\\cal O}(10^{-5})$ t...

  7. Vibrational Frequencies and Spectroscopic Constants for 1(sup 3)A' HNC and 1(sup 3)A' HOC+ from High-Accuracy Quartic Force Fields

    Science.gov (United States)

    Fortenberry, Ryan C.; Crawford, T. Daniel; Lee, Timothy J.

    2014-01-01

    The spectroscopic constants and vibrational frequencies for the 1(sup 3)A' states of HNC, DNC, HOC+, and DOC+ are computed and discussed in this work. The reliable CcCR quartic force field based on high-level coupled cluster ab initio quantum chemical computations is exclusively utilized to provide the anharmonic potential. Then, second order vibrational perturbation theory and vibrational configuration interaction methods are employed to treat the nuclear Schroedinger equation. Second-order perturbation theory is also employed to provide spectroscopic data for all molecules examined. The relationship between these molecules and the corresponding 1(sup 3)A' HCN and HCO+ isomers is further developed here. These data are applicable to laboratory studies involving formation of HNC and HOC+ as well as astronomical observations of chemically active astrophysical environments.

  8. High thermal stability and low Gilbert damping constant of CoFeB/MgO bilayer with perpendicular magnetic anisotropy by Al capping and rapid thermal annealing

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Ding-Shuo; Lai, Shu-Yu; Lin, Tzu-Ying; Wang, Liang-Wei; Liao, Jung-Wei; Lai, Chih-Huang, E-mail: chlai@mx.nthu.edu.tw [Department of Materials Science and Engineering, National Tsing Hua University, Hsinchu, Taiwan (China); Chien, Cheng-Wei; Wang, Yung-Hung [Electronics and Optoelectronics Research Laboratories, Industrial Technology Research Institute, Chutung, Taiwan (China); Ellsworth, David; Lu, Lei; Wu, Mingzhong [Department of Physics, Colorado State University, Fort Collins, Colorado 80523 (United States)

    2014-04-07

    We demonstrate that the magnetic anisotropy of the CoFeB/MgO bilayer can be manipulated by adding an aluminum capping layer. After rapid thermal annealing, we can achieve large perpendicular magnetic anisotropy of CoFeB with a high thermal stability factor (Δ = 72) while the Gilbert damping constant can be reduced down to only 0.011 simultaneously. The boron and residual oxygen in the bulk CoFeB layer are properly absorbed by the Al capping layer during annealing, leading to the enhanced exchange stiffness and reduced damping. The interfacial Fe-O bonding can be optimized by tuning annealing temperature and thickness of Al, resulting in enhanced perpendicular anisotropy.

  9. Interaction Entropy: A New Paradigm for Highly Efficient and Reliable Computation of Protein-Ligand Binding Free Energy.

    Science.gov (United States)

    Duan, Lili; Liu, Xiao; Zhang, John Z H

    2016-05-04

    Efficient and reliable calculation of protein-ligand binding free energy is a grand challenge in computational biology and is of critical importance in drug design and many other molecular recognition problems. The main challenge lies in the calculation of entropic contribution to protein-ligand binding or interaction systems. In this report, we present a new interaction entropy method which is theoretically rigorous, computationally efficient, and numerically reliable for calculating entropic contribution to free energy in protein-ligand binding and other interaction processes. Drastically different from the widely employed but extremely expensive normal mode method for calculating entropy change in protein-ligand binding, the new method calculates the entropic component (interaction entropy or -TΔS) of the binding free energy directly from molecular dynamics simulation without any extra computational cost. Extensive study of over a dozen randomly selected protein-ligand binding systems demonstrated that this interaction entropy method is both computationally efficient and numerically reliable and is vastly superior to the standard normal mode approach. This interaction entropy paradigm introduces a novel and intuitive conceptual understanding of the entropic effect in protein-ligand binding and other general interaction systems as well as a practical method for highly efficient calculation of this effect.

  10. Crystallographic analysis reveals the structural basis of the high-affinity binding of iophenoxic acid to human serum albumin.

    Science.gov (United States)

    Ryan, Ali J; Chung, Chun-Wa; Curry, Stephen

    2011-04-18

    Iophenoxic acid is an iodinated radiocontrast agent that was withdrawn from clinical use because of its exceptionally long half-life in the body, which was due in part to its high-affinity binding to human serum albumin (HSA). It was replaced by Iopanoic acid, which has an amino rather than a hydroxyl group at position 3 on the iodinated benzyl ring and, as a result, binds to albumin with lower affinity and is excreted more rapidly from the body. To understand how iophenoxic acid binds so tightly to albumin, we wanted to examine the structural basis of its interaction with HSA. We have determined the co-crystal structure of HSA in complex with iophenoxic acid at 2.75 Å resolution, revealing a total of four binding sites, two of which--in drugs sites 1 and 2 on the protein--are likely to be occupied at clinical doses. High-affinity binding of iophenoxic acid occurs at drug site 1. The structure reveals that polar and apolar groups on the compound are involved in its interactions with drug site 1. In particular, the 3-hydroxyl group makes three hydrogen bonds with the side-chains of Tyr 150 and Arg 257. The mode of binding to drug site 2 is similar except for the absence of a binding partner for the hydroxyl group on the benzyl ring of the compound. The HSA-iophenoxic acid structure indicates that high-affinity binding to drug site 1 is likely to be due to extensive desolvation of the compound, coupled with the ability of the binding pocket to provide a full set of salt-bridging or hydrogen bonding partners for its polar groups. Consistent with this interpretation, the structure also suggests that the lower-affinity binding of iopanoic acid arises because replacement of the 3-hydroxyl by an amino group eliminates hydrogen bonding to Arg 257. This finding underscores the importance of polar interactions in high-affinity binding to albumin.

  11. Crystallographic analysis reveals the structural basis of the high-affinity binding of iophenoxic acid to human serum albumin

    Directory of Open Access Journals (Sweden)

    Chung Chun-wa

    2011-04-01

    Full Text Available Abstract Background Iophenoxic acid is an iodinated radiocontrast agent that was withdrawn from clinical use because of its exceptionally long half-life in the body, which was due in part to its high-affinity binding to human serum albumin (HSA. It was replaced by Iopanoic acid, which has an amino rather than a hydroxyl group at position 3 on the iodinated benzyl ring and, as a result, binds to albumin with lower affinity and is excreted more rapidly from the body. To understand how iophenoxic acid binds so tightly to albumin, we wanted to examine the structural basis of its interaction with HSA. Results We have determined the co-crystal structure of HSA in complex with iophenoxic acid at 2.75 Å resolution, revealing a total of four binding sites, two of which - in drugs sites 1 and 2 on the protein - are likely to be occupied at clinical doses. High-affinity binding of iophenoxic acid occurs at drug site 1. The structure reveals that polar and apolar groups on the compound are involved in its interactions with drug site 1. In particular, the 3-hydroxyl group makes three hydrogen bonds with the side-chains of Tyr 150 and Arg 257. The mode of binding to drug site 2 is similar except for the absence of a binding partner for the hydroxyl group on the benzyl ring of the compound. Conclusions The HSA-iophenoxic acid structure indicates that high-affinity binding to drug site 1 is likely to be due to extensive desolvation of the compound, coupled with the ability of the binding pocket to provide a full set of salt-bridging or hydrogen bonding partners for its polar groups. Consistent with this interpretation, the structure also suggests that the lower-affinity binding of iopanoic acid arises because replacement of the 3-hydroxyl by an amino group eliminates hydrogen bonding to Arg 257. This finding underscores the importance of polar interactions in high-affinity binding to albumin.

  12. Design, Synthesis, and in Vitro Pharmacology of New Radiolabeled γ-Hydroxybutyric Acid Analogues Including Photolabile Analogues with Irreversible Binding to the High-Affinity γ-Hydroxybutyric Acid Binding Sites

    DEFF Research Database (Denmark)

    Sabbatini, Paola; Wellendorph, Petrine; Høg, Signe;

    2010-01-01

    γ-Hydroxybutyric acid (GHB) is a psychotropic compound endogenous to the brain. Despite its potential physiological significance, the complete molecular mechanisms of action remain unexplained. To facilitate the isolation and identification of the high-affinity GHB binding site, we herein report...... the design and synthesis of the first 125I-labeled radioligands in the field, one of which contains a photoaffinity label which enables it to bind irreversibly to the high-affinity GHB binding sites....

  13. The binding of pentapeptides to biological and synthetic high affinity heparin.

    Science.gov (United States)

    Flengsrud, Ragnar; Antonsen, Simen Gjelseth

    2015-11-01

    Pentapeptides have been shown to bind the synthetic heparin fondaparinux (Arixtra) as well the biological heparins dalteparin (Fragmin) and salmon heparin. In contrast to heparin binding consensus sequences, the pentapeptides are acidic or neutral, with no arginine or histidine residue. The peptides showed an effect on in vitro heparin anti-factor X activity with a reduction of fondaparinux activity by 65-95%. Heparin binding was further studied by using peptide solid phase chromatography and NMR analysis.

  14. Hyperfine coupling constant for {sup 59}Co estimated from a high-field susceptibility and high-field NMR shift in ferromagnetic Co{sub 2}TiGa and Co{sub 2}VGa

    Energy Technology Data Exchange (ETDEWEB)

    Nishihara, H; Furutani, Y; Wada, T [Faculty of Science and Technology, Ryukoku University, Otsu 520-2194 (Japan); Kanomata, T [Faculty of Engineering, Tohoku Gakuin University, Tagajo 985-8537 (Japan); Kobayashi, K; Ishida, K [Graduate School of Engineering, Tohoku University, Sendai 980-8579 (Japan); Kainuma, R [Institute of Multidisciplinary Research for Advanced Materials, Tohoku University, Sendai 980-8577 (Japan); Koyama, K; Watanabe, K, E-mail: nishihara@rins.ryukoku.ac.j [Institute for Materials Research, Tohoku University, Sendai 980-8577 (Japan)

    2010-01-01

    The hyperfine coupling constant for {sup 59}Co in the ferromagnetic state of a Heusler alloy Co{sub 2}TiGa in the high-field range has been estimated to be 170 kOe per 1 {mu}{sub B} of the moment of cobalt atom from the high-field magnetic susceptibility of 2.3 x 10{sup -6} emu/gOe measured at 5 K in a field range from 20 to 70 kOe and a reported positive high-field NMR shift of +0.83 percent. The value of the hyperfine coupling constant has been found to be even larger in the case of Co{sub 2}VGa. These features suggest that the positive hyperfine field at Co nucleus in the Co-based Heusler alloys is due to conduction electron polarization rather than a transferred hyperfine field.

  15. Selective elimination of high constitutive activity or chemokine binding in the human herpesvirus 8 encoded seven transmembrane oncogene ORF74

    DEFF Research Database (Denmark)

    Rosenkilde, M M; Kledal, T N; Holst, Peter Johannes;

    2000-01-01

    expressing ORF74 under control of the CD2 promoter develop highly vascularized Kaposi's sarcoma-like tumors. Through targeted mutagenesis we here create three distinct phenotypes of ORF74: a receptor with normal, high constitutive signaling through the phospholipase C pathway but deprived of binding...

  16. Constant domain-regulated antibody catalysis.

    Science.gov (United States)

    Sapparapu, Gopal; Planque, Stephanie; Mitsuda, Yukie; McLean, Gary; Nishiyama, Yasuhiro; Paul, Sudhir

    2012-10-19

    Some antibodies contain variable (V) domain catalytic sites. We report the superior amide and peptide bond-hydrolyzing activity of the same heavy and light chain V domains expressed in the IgM constant domain scaffold compared with the IgG scaffold. The superior catalytic activity of recombinant IgM was evident using two substrates, a small model peptide that is hydrolyzed without involvement of high affinity epitope binding, and HIV gp120, which is recognized specifically by noncovalent means prior to the hydrolytic reaction. The catalytic activity was inhibited by an electrophilic phosphonate diester, consistent with a nucleophilic catalytic mechanism. All 13 monoclonal IgMs tested displayed robust hydrolytic activities varying over a 91-fold range, consistent with expression of the catalytic functions at distinct levels by different V domains. The catalytic activity of polyclonal IgM was superior to polyclonal IgG from the same sera, indicating that on average IgMs express the catalytic function at levels greater than IgGs. The findings indicate a favorable effect of the remote IgM constant domain scaffold on the integrity of the V-domain catalytic site and provide a structural basis for conceiving antibody catalysis as a first line immune function expressed at high levels prior to development of mature IgG class antibodies.

  17. Time evolution of the high temperature region formed by laser induced breakdown and of the development of the flame kernel in the constant volume combustion vessel

    Science.gov (United States)

    Hayashi, J.; Nakatsuka, N.; Morimoto, I.; Akamatsu, F.

    2017-02-01

    The lean combustion is one of the key techniques for the advanced internal combustion systems due to the requirement of the higher thermal efficiency. Since the successful ignition must be guaranteed even in the lean combustion, advanced ignition systems have been developed in this decade. Laser ignition is one of the advanced ignition systems which have the profits of the flexibility in the position and the timing of ignition. To develop this ignition system for the actual combustion system, it is required to reveal the underlying physics of the laser ignition. Particularly, the time evolution of high temperature region formed by laser induced breakdown should be discussed. In this study, therefore, the time evolution of the high temperature region formed by the laser induced breakdown and the development of flame kernel were observed by using high-speed imaging. The ignition trials of methane/air lean premixed mixture were carried out in the constant volume combustion vessel to obtain minimum laser pulse energy for ignition (MPE). Results showed that the light emission from plasma formed by laser induced breakdown remained at least in several tens nano-seconds. In addition, there were large differences between the breakdown threshold and the MPE, which meant that the breakdown threshold did not determine the minimum pulse energy for ignition.

  18. Achieving high dielectric constant and low loss property in a dipolar glass polymer containing strongly dipolar and small-sized sulfone groups.

    Science.gov (United States)

    Wei, Junji; Zhang, Zhongbo; Tseng, Jung-Kai; Treufeld, Imre; Liu, Xiaobo; Litt, Morton H; Zhu, Lei

    2015-03-11

    In this report, a dipolar glass polymer, poly(2-(methylsulfonyl)ethyl methacrylate) (PMSEMA), was synthesized by free radical polymerization of the corresponding methacrylate monomer. Due to the large dipole moment (4.25 D) and small size of the side-chain sulfone groups, PMSEMA exhibited a strong γ transition at a temperature as low as -110 °C at 1 Hz, about 220 °C below its glass transition temperature around 109 °C. Because of this strong γ dipole relaxation, the glassy PMSEMA sample exhibited a high dielectric constant of 11.4 and a low dissipation factor (tan δ) of 0.02 at 25 °C and 1 Hz. From an electric displacement-electric field (D-E) loop study, PMSEMA demonstrated a high discharge energy density of 4.54 J/cm(3) at 283 MV/m, nearly 3 times that of an analogue polymer, poly(methyl methacrylate) (PMMA). However, the hysteresis loss was only 1/3-1/2 of that for PMMA. This study suggests that dipolar glass polymers with large dipole moments and small-sized dipolar side groups are promising candidates for high energy density and low loss dielectric applications.

  19. A novel high throughput screening assay for binding affinities of perfluoroalkyl iodide for estrogen receptor alpha and beta isoforms.

    Science.gov (United States)

    Song, Wenting; Zhao, Lixia; Sun, Zhendong; Yang, Xiaoxi; Zhou, Qunfang; Jiang, Guibin

    2017-12-01

    Contaminants of emerging concern are continuously increasing, which makes it important to develop high throughput screening techniques for the evaluation of their potential biological effects, especially endocrine disrupting effects, which would directly influence the population dynamics in environment. A novel competitive binding assay based on enzyme fragmentation complementation technology was established to screen the binding affinities of emerging chemicals for estrogen receptor (ER) α or β isoforms. Exogenous compounds could compete with the fragment (ED-ES) of genetically engineered β-galactosidase enzyme (β-gal) for the binding to ERα or β, thus quantitatively altering the formation of enzymatically active β-gal and the hydrolysis of luminescent substrate. According to the monitoring of luminescence curves and the optimization of ERα or β concentrations, it was found that luminescent signals were sustainably emitted for 9h, and 40nM ERα or β in the system would lead to the most sensitive luminescence response. Using 17β-estrodiol (E2) and genistein as the representative estrogenic hormones, their binding affinities for ERα and β were evaluated. The results were consistent with those determined by traditional methods, which confirmed the reliability of this competitive binding assay based on β-gal. Four polyfluorinated iodine alkanes (PFIs) with specific structural characteristics in iodine substitution and carbon chain length were screened, and the results showed diverse binding affinities and different preferences of these chemicals to ERα or β isoforms. The binding affinities of PFIs for ERα were consistent with the result from MVLN transcriptional reporter assay. Overall, the competitive binding assay presented in this study provided a promising alternative to high throughput screening of emerging chemicals with estrogenic effects, which would be important in explanation of their potential toxicological effects and human exposure risks

  20. Recombinant human nerve growth factor is biologically active and labels novel high-affinity binding sites in rat brain

    Energy Technology Data Exchange (ETDEWEB)

    Altar, C.A.; Burton, L.E.; Bennett, G.L.; Dugich-Djordjevic, M. (Genentech, Inc., South San Francisco, CA (USA))

    1991-01-01

    Iodinated recombinant human nerve growth factor (125I-rhNGF) stimulated neurite formation in PC12 cell cultures with a half-maximal potency of 35-49 pg/ml, compared with 39-52 pg/ml for rhNGF. In quantitative ligand autoradiography, the in vitro equilibrium binding of 125I-rhNGF to brain sections showed a 10-fold regional variation in density and was saturable, reversible, and specifically displaced by up to 74% with rhNGF or murine NGF (muNGF). At equilibrium, 125I-rhNGF bound to these sites with high affinity and low capacity (Bmax less than or equal to 13.2 fmol/mg of protein). Calculation of 125I-rhNGF binding affinity by kinetic methods gave average Kd values of 24 and 31 pM. Computer-generated maps revealed binding in brain regions not identified previously with 125I-muNGF, including hippocampus; dentate gyrus; amygdala; paraventricular thalamus; frontal, parietal, occipital, and cingulate cortices; nucleus accumbens; olfactory tubercle; subiculum; pineal gland; and medial geniculate nucleus. NGF binding sites were distributed in a 2-fold increasing medial-lateral gradient in the caudate-putamen and a 2-fold lateral-medial gradient in the nucleus accumbens. 125I-rhNGF binding sites were also found in most areas labeled by 125I-muNGF, including the interpedunucular nucleus, cerebellum, forebrain cholinergic nuclei, caudoventral caudate-putamen, and trigeminal nerve nucleus. 125I-rhNGF binding sites were absent from areas replete with low-affinity NGF binding sites, including circumventricular organs, myelinated fiber bundles, and choroid plexus. The present analysis provides an anatomical differentiation of high-affinity 125I-rhNGF binding sites and greatly expands the number of brain structures that may respond to endogenous NGF or exogenously administered rhNGF.

  1. High-performance chromatofocusing and size-exclusion chromatography: separation of human uterine estrogen-binding proteins.

    Science.gov (United States)

    Hutchens, T W; Gibbons, W E; Besch, P K

    1984-08-03

    We have employed high-performance chromatofocusing (HPCF) and high-performance size exclusion chromatography (HPSEC) to separate and identify radiolabelled estrogen binding proteins present in human uterine cytosol. Results obtained using these high-performance methods are compared to results of similar analyses by conventional isoelectric focusing procedures and open-column size exclusion chromatography. By HPCF, descending pH gradients (pH 8-4) allow us to discern four to five estrogen binding proteins with elution pH values typically between pH 4.5 and 7.2. However, when using HPCF, significant quantities of estrogen binding proteins are rarely detected between pH 7.2 and 8.0. This observation has been confirmed by open-column chromatofocusing of these proteins on Polybuffer Exchanger 94. In contrast, preparative isoelectric focusing by electrophoresis in polyampholytes reveals substantial quantities of estrogen binding activity eluted between pH 7.5 and 8.0. Several possible explanations for this disparity are discussed. Apparent differences are also observed when the size heterogeneity of estrogen binding proteins is analyzed simultaneously by size exclusion chromatography in open-column (Sephacryl S-300) and high-performance (TSK-3000SW) modes. HPSEC of these estrogen binding proteins on TSK-3000SW columns demonstrates a predominant 80-85 A species, whereas size exclusion chromatography on conventional Sephacryl S-300 columns reveals two to three distinct regions of estrogen binding proteins with Stokes radii of ca. 85, 60 and 30 A (major species). The larger form of receptor, whether a non-specific aggregate or a multisubunit complex, is stable in unfractionated cytosol and becomes more labile only during size exclusion chromatography.

  2. Ribosomal binding site switching: an effective strategy for high-throughput cloning constructions.

    Directory of Open Access Journals (Sweden)

    Yangbo Hu

    Full Text Available Direct cloning of PCR fragments by TA cloning or blunt end ligation are two simple methods which would greatly benefit high-throughput (HTP cloning constructions if the efficiency can be improved. In this study, we have developed a ribosomal binding site (RBS switching strategy for direct cloning of PCR fragments. RBS is an A/G rich region upstream of the translational start codon and is essential for gene expression. Change from A/G to T/C in the RBS blocks its activity and thereby abolishes gene expression. Based on this property, we introduced an inactive RBS upstream of a selectable marker gene, and designed a fragment insertion site within this inactive RBS. Forward and reverse insertions of specifically tailed fragments will respectively form an active and inactive RBS, thus all background from vector self-ligation and fragment reverse insertions will be eliminated due to the non-expression of the marker gene. The effectiveness of our strategy for TA cloning and blunt end ligation are confirmed. Application of this strategy to gene over-expression, a bacterial two-hybrid system, a bacterial one-hybrid system, and promoter bank construction are also verified. The advantages of this simple procedure, together with its low cost and high efficiency, makes our strategy extremely useful in HTP cloning constructions.

  3. Drug binding to the inactivated state is necessary but not sufficient for high-affinity binding to human ether-à-go-go-related gene channels.

    Science.gov (United States)

    Perrin, Mark J; Kuchel, Philip W; Campbell, Terence J; Vandenberg, Jamie I

    2008-11-01

    Drug block of the human ether-à-go-go-related gene K(+) channel (hERG) is the most common cause of acquired long QT syndrome, a disorder of cardiac repolarization that may result in ventricular tachycardia and sudden cardiac death. We investigated the open versus inactivated state dependence of drug block by using hERG mutants N588K and N588E, which shift the voltage dependence of inactivation compared with wild-type but in which the mutated residue is remote from the drug-binding pocket in the channel pore. Four high-affinity drugs (cisapride, dofetilide, terfenadine, and astemizole) demonstrated lower affinity for the inactivation-deficient N588K mutant hERG channel compared with N588E and wild-type hERG. Three of four low-affinity drugs (erythromycin, perhexiline, and quinidine) demonstrated no preference for N588E over N588K channels, whereas dl-sotalol was an example of a low-affinity state-dependent blocker. All five state-dependent blockers showed an even lower affinity for S620T mutant hERG (no inactivation) compared with N588K mutant hERG (greatly reduced inactivation). Computer modeling indicates that the reduced affinity for S620T compared with N588K and wild-type channels can be explained by the relative kinetics of drug block and unblock compared with the kinetics of inactivation and recovery from inactivation. We were also able to calculate, for the first time, the relative affinities for the inactivated versus the open state, which for the drugs tested here ranged from 4- to 70-fold. Our results show that preferential binding to the inactivated state is necessary but not sufficient for high-affinity binding to hERG channels.

  4. Humanized mAb H22 binds the human high affinity Fc receptor for IgG (FcgammaRI), blocks phagocytosis, and modulates receptor expression.

    Science.gov (United States)

    Wallace, P K; Keler, T; Coleman, K; Fisher, J; Vitale, L; Graziano, R F; Guyre, P M; Fanger, M W

    1997-10-01

    About 10-15% of patients with immune thrombocytopenic purpura (ITP) cannot be controlled by corticosteroid therapy and splenectomy. For these patients treatment with high-dose IVIgG induces partial or complete responses. The clinical benefits of IVIgG could be due to blockade of Fc receptors for IgG (FcgammaR), because several model systems clearly show that functional FcgammaR are essential for establishment of ITP and related diseases. However, the specific contributions of the three individual classes of FcgammaR remain to be more completely defined. Recently monoclonal antibody (mAb) H22, which recognizes an epitope on FcgammaRI (CD64) outside the ligand binding domain, was humanized by grafting its complementarity determining regions onto human IgG1 constant domains. Because FcgammaRI has a high affinity for human IgG1 antibodies, we predicted mAb H22 would also bind to FcgammaRI through its Fc domain and block FcgammaRI-mediated phagocytosis. These studies demonstrate that mAb H22 blocked phagocytosis of opsonized red blood cells 1000 times more effectively than an irrelevant IgG. Moreover, cross-linking FcgammaRI with mAb H22 rapidly down-modulated FcgammaRI expression on monocytes without affecting other surface antigens. We conclude that because mAb H22 is a humanized mAb that blocks the FcgammaRI ligand binding domain and down-modulates FcgammaRI expression, it is a particularly good candidate for evaluating the role of FcgammaRI in patients with ITP.

  5. High sensitivity electron diffraction analysis. A study of divalent cation binding to purple membrane.

    Science.gov (United States)

    Mitra, A K; Stroud, R M

    1990-02-01

    A sensitive high-resolution electron diffraction assay for change in structure is described and harnessed to analyze the binding of divalent cations to the purple membrane (PM) of Halobacterium halobium. Low-dose electron diffraction patterns are subject to a matched filter algorithm (Spencer, S. A., and A. A. Kossiakoff. 1980. J. Appl. Crystallogr. 13:563-571). to extract accurate values of reflection intensities. This, coupled with a scheme to account for twinning and specimen tilt in the microscope, yields results that are sensitive enough to rapidly quantitate any structure change in PM brought about by site-directed mutagenesis to the level of less than two carbon atoms. Removal of tightly bound divalent cations (mainly Ca2+ and Mg2+) from PM causes a color change to blue and is accompanied by a severely altered photocycle of the protein bacteriohodopsin (bR), a light-driven proton pump. We characterize the structural changes that occur upon association of 3:1 divalent cation to PM, versus membranes rendered purple by addition of excess Na+. High resolution, low dose electron diffraction data obtained from glucose-embedded samples of Pb2+ and Na+ reconstituted PM preparations at room temperature identify several sites with total occupancy of 2.01 +/- 0.05 Pb2+ equivalents. The color transition as a function of ion concentration for Ca2+ or Mg2+ and Pb2+ are strictly comparable. A (Pb2(+)-Na+) PM Fourier difference map in projection was synthesized at 5 A using the averaged data from several nominally untilted patches corrected for twinning and specimen tilt. We find six major sites located on helices 7, 5, 4, 3, 2 (nomenclature of Engelman et al. 1980. Proc. Natl. Acad. Sci. USA. 77:2023-2027) in close association with bR. These partially occupied sites (0.55-0.24 Pb2+ equivalents) represent preferential sites of binding for divalent cations and complements our earlier result by x-ray diffraction (Katre et al. 1986. Biophys. J. 50:277-284).

  6. The prion protein binds thiamine.

    Science.gov (United States)

    Perez-Pineiro, Rolando; Bjorndahl, Trent C; Berjanskii, Mark V; Hau, David; Li, Li; Huang, Alan; Lee, Rose; Gibbs, Ebrima; Ladner, Carol; Dong, Ying Wei; Abera, Ashenafi; Cashman, Neil R; Wishart, David S

    2011-11-01

    Although highly conserved throughout evolution, the exact biological function of the prion protein is still unclear. In an effort to identify the potential biological functions of the prion protein we conducted a small-molecule screening assay using the Syrian hamster prion protein [shPrP(90-232)]. The screen was performed using a library of 149 water-soluble metabolites that are known to pass through the blood-brain barrier. Using a combination of 1D NMR, fluorescence quenching and surface plasmon resonance we identified thiamine (vitamin B1) as a specific prion ligand with a binding constant of ~60 μM. Subsequent studies showed that this interaction is evolutionarily conserved, with similar binding constants being seen for mouse, hamster and human prions. Various protein construct lengths, both with and without the unstructured N-terminal region in the presence and absence of copper, were examined. This indicates that the N-terminus has no influence on the protein's ability to interact with thiamine. In addition to thiamine, the more biologically abundant forms of vitamin B1 (thiamine monophosphate and thiamine diphosphate) were also found to bind the prion protein with similar affinity. Heteronuclear NMR experiments were used to determine thiamine's interaction site, which is located between helix 1 and the preceding loop. These data, in conjunction with computer-aided docking and molecular dynamics, were used to model the thiamine-binding pharmacophore and a comparison with other thiamine binding proteins was performed to reveal the common features of interaction.

  7. High-spatial-resolution three-dimensional MR cholangiography using a high-sampling-efficiency technique (SPACE) at 3T: comparison with the conventional constant flip angle sequence in healthy volunteers.

    Science.gov (United States)

    Arizono, Shigeki; Isoda, Hiroyoshi; Maetani, Yoji S; Hirokawa, Yuusuke; Shimada, Kotaro; Nakamoto, Yuji; Togashi, Kaori

    2008-09-01

    To evaluate the image quality of high-spatial-resolution three-dimensional magnetic resonance cholangiography (MRC) with a high-sampling-efficiency technique (sampling perfection with application optimized contrasts using different flip angle evolutions [SPACE]) in comparison with a conventional constant flip angle (FA) sequence at 3T. Eighteen volunteers were examined on a 3T MR unit using MRC imaging performed with three different free-breathing three-dimensional T2-weighted turbo spin-echo (TSE) sequences: 1) SPACE (spatial resolution, 1.1x1.0x0.84 mm), 2) constant FA (1.1x1.0x0.84 mm), and 3) SPACE at a higher resolution (SPACE HR; 1.0x0.9x0.644 mm). A five-point scale was used to compare overall image quality and visualization of the third branches of the bile duct (B2, B6, and B8). Depictions of cystic duct insertion and the highest order of bile duct visible were also compared. MRC with SPACE and SPACE HR sequences produced significantly better overall image quality than the constant FA sequence. In all analyses of duct visibility, SPACE and SPACE HR sequences showed higher scores than the constant FA sequence. High-resolution three-dimensional MRC with SPACE at 3T allows high-quality imaging of the biliary tract, and has the ability to depict nondilated intrahepatic bile ducts (IHBD) in healthy volunteers. Copyright (c) 2008 Wiley-Liss, Inc.

  8. A novel high efficiency CMOS RF/DC power harvester based on constant on/off time buck controller for 60GHz frequency band

    Directory of Open Access Journals (Sweden)

    Ninić Marko

    2017-01-01

    Full Text Available A novel 60 GHz RF/DC power harvesting system is presented. The system consists of RF to DC rectifier and a DC/DC Buck converter based on constant ON/OFF time (COOT control. The rectifier has a structure of voltage doubler, but employs diodes that have lower parasitics compared to those of the standard MOSFET diodes, resulting in improved power conversion efficiency. The peak efficiency of the rectifier obtained with the extracted parasitics for the output power of 1 mW is about 25%. In order to keep the output voltage of the system to 1.2 V, the COOT control in the Buck converter is used. COOT control has much better efficiency at low output powers compared to the PWM systems. For correct operation of the COOT control, auxiliary sub-blocks: a low power high-speed comparator, a hysteresis comparator, and a high-speed voltage reference are designed and presented. The maximum switching frequency in the Buck converter is about 100MHz and the whole control system has very low static power consumption. The efficiency of the overall system for the output power of 1mW is about 21%. The system is designed in 65 nm CMOS technology.

  9. Highly charged W+13, Ir+16, and Pt+17 ions as promising optical clock candidates for probing variations of the fine-structure constant

    Science.gov (United States)

    Nandy, D. K.; Sahoo, B. K.

    2016-09-01

    Transitions among the first three low-lying states in the highly charged W+13, Ir+16, and Pt+17 ions are found to be strongly forbidden with wavelengths in the optical regime. By determining their energy levels, lifetimes, and other spectroscopic properties that are decisive quantities for estimating dominant systematics due to stray electromagnetic interactions in an experiment, we demonstrate that it can be possible to measure frequencies of the lowest forbidden transitions below a 10-19 precision level in the above ions, and hence, they seem to be suitable for frequency standards. We employ a sophisticated relativistic coupled cluster method to carry out calculations of these properties of the above states involving 4 f - and 5 s -core orbitals. We also found, by estimating their relativistic sensitivity coefficients, that these clock transitions can be highly sensitive to the tiny drift in the fine-structure constant αe. Consequently, a clock based on one of these ions, particularly Pt+17, could be used for corroborating the hypothesis of temporal and spatial variation in αe.

  10. Overcoming silicon limitations: new 3D-AFM carbon tips with constantly high-resolution for sub-28nm node semiconductor requirements

    Science.gov (United States)

    Foucher, Johann; Schmidt, Sebastian W.; Penzkofer, Christian; Irmer, Bernd

    2012-03-01

    The demands on atomic force microscopy (AFM) as a reference technique for precisely determining surface properties and structural designs of multiple patterns in the semiconductor industry are steadily increasing. With the aim to meet ITRS requirements and simultaneously improve the accuracy of AFM-based critical dimension (CD) measurements at constant resolution, the AFM tip more and more becomes a factor crucially determining the AFM performance. In this context, AFM tip limitations are given by lack of sharpness with too large tip radii/diameter, insufficient wear resistance, and high total cost, which does not conform to production environment needs. One technical approach to overcome these tip limitations is provided by electron beam induced processing (EBIP), which allows for manufacturing AFM tips of desired sharpness, shape, and mechanical stability. Here, we present T-shape-like 3D-AFM tips made of bulk amorphous, high density diamond-like carbon (HDC/DLC), and compare their performance and wear resistance to standard silicon tips. We show the advantages of this approach for the semiconductor industry, in particular on AFM3D technology in order to answer to sub-28 nm nodes requirements, and present tips with 15 nm diameter at 10 nm vertical edge height.

  11. Highly effective recognition of carbohydrates by phenanthroline-based receptors: alpha- versus beta-anomer binding preference.

    Science.gov (United States)

    Mazik, Monika; Hartmann, Andrè; Jones, Peter G

    2009-09-14

    (1)H NMR spectroscopic titrations in competitive and non-competitive media, as well as binding studies in two-phase systems, such as phase transfer of sugars from aqueous into organic solvents and dissolution of solid carbohydrates in apolar media revealed both highly effective recognition of neutral carbohydrates and interesting binding preferences of an acyclic phenanthroline-based receptor 1. Compared to the previously described acyclic receptors, compound 1 displays significantly higher binding affinities, the rare capability to extract sugars from water into non-polar organic solutions and alpha- versus beta-anomer binding preference in the recognition of glycosides, which differs from those observed for other receptor systems. X-ray crystallographic investigations revealed the presence of water molecules in the binding pocket of 1 that are engaged in the formation of hydrogen-bonding motifs similar to those suggested by molecular modelling for the sugar OH groups in the receptor-sugar complexes. The molecular modelling calculations, synthesis, crystal structure and binding properties of 1 are described and compared with those of the previously described receptors.

  12. Effect of Proteolysis with Alkaline Protease Following High Hydrostatic Pressure Treatment on IgE Binding of Buckwheat Protein.

    Science.gov (United States)

    Lee, Chaeyoon; Lee, Wonhui; Han, Youngshin; Oh, Sangsuk

    2017-03-01

    Buckwheat is a popular food material in many Asian countries and it contains major allergenic proteins. This study was performed to analyze the effects of hydrolysis with alkaline protease following high hydrostatic pressure (HHP) treatment on the IgE binding of buckwheat protein. Extracted buckwheat protein was treated with HHP at 600 MPa for 30 min and hydrolyzed with alkaline protease for 240 min. IgE binding was examined using an enzyme-linked immunosorbent assay (ELISA) with serum samples from 14 patients who were allergic to buckwheat. Depending on the serum samples, HHP treatment of buckwheat protein without enzymatic hydrolysis decreased the IgE binding by 8.9% to 73.2% or increased by 31% to 78%. The IgE binding of buckwheat protein hydrolyzed with alkaline protease decreased by 73.8% to 100%. The IgE binding of buckwheat protein hydrolyzed with alkaline protease following HHP treatment decreased by 83.8% to 100%. This suggested that hydrolysis with alkaline protease following HHP treatment could be applied to reduce the IgE binding of buckwheat protein. © 2017 Institute of Food Technologists®.

  13. Spatial Variations of Fundamental Constants

    CERN Document Server

    Barrow, John D; Barrow, John D.; Toole, Chris O'

    1999-01-01

    We show that observational limits on the possible time variation of constants of Nature are significantly affected by allowing for both space and time variation. Bekenstein's generalisation of Maxwell's equations to allow for cosmological variation of $alpha$ is investigated in a universe containing spherically symmetric inhomogeneities. The time variation of $alpha$ is determined by the local matter density and hence limits obtained in high-density geophysical enviroments are far more constraining than those obtained at high redshift. This new feature is expected to be a property of a wide class of theories for the variation of constants.

  14. Limiting values of the one-bond Csbnd H spin-spin coupling constants of the imidazole ring of histidine at high-pH

    Science.gov (United States)

    Vila, Jorge A.; Scheraga, Harold A.

    2017-04-01

    Assessment of the relative amounts of the forms of the imidazole ring of Histidine (His), namely the protonated (H+) and the tautomeric Nε2-H and Nδ1-H forms, respectively, is a challenging task in NMR spectroscopy. Indeed, their determination by direct observation of the 15N and 13C chemical shifts or the one-bond Csbnd H, 1JCH, Spin-Spin Coupling Constants (SSCC) requires knowledge of the "canonical" limiting values of these forms in which each one is present to the extent of 100%. In particular, at high-pH, an accurate determination of these "canonical" limiting values, at which the tautomeric forms of His coexist, is an elusive problem in NMR spectroscopy. Among different NMR-based approaches to treat this problem, we focus here on the computation, at the DFT level of theory, of the high-pH limiting value for the 1JCH SSCC of the imidazole ring of His. Solvent effects were considered by using the polarizable continuum model approach. The results of this computation suggest, first, that the value of 1JCε1H = 205 ± 1.0 Hz should be adopted as the canonical high-pH limiting value for this SSCC; second, the variation of 1JCε1H SSCC during tautomeric changes is minor, i.e., within ±1 Hz; and, finally, the value of 1JCδ2H SSCC upon tautomeric changes is large (15 Hz) indicating that, at high-pH or for non-protonated His at any pH, the tautomeric fractions of the imidazole ring of His can be predicted accurately as a function of the observed value of 1JCδ2H SSCC.

  15. Generalized Pickands constants

    NARCIS (Netherlands)

    Debicki, K.G.

    2001-01-01

    Pickands constants play an important role in the exact asymptotic of extreme values for Gaussian stochastic processes. By the {it generalized Pickands constant ${cal H_{eta$ we mean the limit begin{eqnarray* {cal H_{eta= lim_{T to inftyfrac{ {cal H_{eta(T){T, end{eqnarray* where ${cal H_{eta(T)= Exp

  16. Trimeric gp120-specific bovine monoclonal antibodies require cysteine and aromatic residues in CDRH3 for high affinity binding to HIV Env

    Science.gov (United States)

    Center, Rob J.; Bebbington, Jonathan; Cuthbertson, Jack; Khoury, Georges; Lichtfuss, Marit; Rawlin, Grant; Purcell, Damian

    2017-01-01

    ABSTRACT We isolated HIV-1 Envelope (Env)-specific memory B cells from a cow that had developed high titer polyclonal immunoglobulin G (IgG) with broad neutralizing activity after a long duration vaccination with HIV-1AD8 Env gp140 trimers. We cloned the bovine IgG matched heavy (H) and light (L) chain variable (V) genes from these memory B cells and constructed IgG monoclonal antibodies (mAbs) with either a human constant (C)-region/bovine V-region chimeric or fully bovine C and V regions. Among 42 selected Ig+ memory B cells, two mAbs (6A and 8C) showed high affinity binding to gp140 Env. Characterization of both the fully bovine and human chimeric isoforms of these two mAbs revealed them as highly type-specific and capable of binding only to soluble AD8 uncleaved gp140 trimers and covalently stabilized AD8 SOSIP gp140 cleaved trimers, but not monomeric gp120. Genomic sequence analysis of the V genes showed the third heavy complementarity-determining region (CDRH3) of 6A mAb was 21 amino acids in length while 8C CDRH3 was 14 amino acids long. The entire V heavy (VH) region was 27% and 25% diverged for 6A and 8C, respectively, from the best matched germline V genes available, and the CDRH3 regions of 6A and 8C were 47.62% and 78.57% somatically mutated, respectively, suggesting a high level of somatic hypermutation compared with CDRH3 of other species. Alanine mutagenesis of the VH genes of 6A and 8C, showed that CDRH3 cysteine and tryptophan amino acids were crucial for antigen binding. Therefore, these bovine vaccine-induced anti-HIV antibodies shared some of the notable structural features of elite human broadly neutralizing antibodies, such as CDRH3 size and somatic mutation during affinity-maturation. However, while the 6A and 8C mAbs inhibited soluble CD4 binding to gp140 Env, they did not recapitulate the neutralizing activity of the polyclonal antibodies against HIV infection. PMID:27996375

  17. High-Mannose Specific Lectin and Its Recombinants from a Carrageenophyta Kappaphycus alvarezii Represent a Potent Anti-HIV Activity Through High-Affinity Binding to the Viral Envelope Glycoprotein gp120.

    Science.gov (United States)

    Hirayama, Makoto; Shibata, Hiromi; Imamura, Koji; Sakaguchi, Takemasa; Hori, Kanji

    2016-04-01

    We previously reported that a high-mannose binding lectin KAA-2 from the red alga Kappaphycus alvarezii, which is an economically important species and widely cultivated as a source of carrageenans, had a potent anti-influenza virus activity. In this study, the full-length sequences of two KAA isoforms, KAA-1 and KAA-2, were elucidated by a combination of peptide mapping and cDNA cloning. They consisted of four internal tandem-repeated domains, which are conserved in high-mannose specific lectins from lower organisms, including a cyanobacterium Oscillatoria agardhii and a red alga Eucheuma serra. Using an Escherichia coli expression system, an active recombinant form of KAA-1 (His-tagged rKAA-1) was successfully generated in the yield of 115 mg per a litter of culture. In a detailed oligosaccharide binding analysis by a centrifugal ultrafiltration-HPLC method with 27 pyridylaminated oligosaccharides, His-tagged rKAA-1 and rKAA-1 specifically bound to high-mannose N-glycans with an exposed α1-3 mannose in the D2 arm as the native lectin did. Predicted from oligosaccharide-binding specificity, a surface plasmon resonance analysis revealed that the recombinants exhibit strong interaction with gp120, a heavily glycosylated envelope glycoprotein of HIV with high association constants (1.48-1.61 × 10(9) M(-1)). Native KAAs and the recombinants inhibited the HIV-1 entry at IC50s of low nanomolar levels (7.3-12.9 nM). Thus, the recombinant proteins would be useful as antiviral reagents targeting the viral surface glycoproteins with high-mannose N-glycans, and the cultivated alga K. alvarezii could also be a good source of not only carrageenans but also this functional lectin(s).

  18. Human insulin production from a novel mini-proinsulin which has high receptor-binding activity.

    Science.gov (United States)

    Chang, S G; Kim, D Y; Choi, K D; Shin, J M; Shin, H C

    1998-02-01

    To increase the folding efficiency of the insulin precursor and the production yield of insulin, we have designed a mini-proinsulin (M2PI) having the central C-peptide region replaced with a sequence forming a reverse turn. The mini-proinsulin was fused at the N-terminus to a 21-residue fusion partner containing a His10 tag for affinity purification. The gene for the fusion protein was inserted downstream of the T7 promoter of the expression plasmid pET-3a, and the fusion proteins were produced as inclusion bodies in the Escherichia coli cytoplasm at levels up to 25% of the total cell protein. The protein was sulphonated, cleaved by CNBr and the M2PI mini-proinsulin was purified using ion-exchange chromatography. The refolding yield of M2PI was 20-40% better than that of proinsulin studied at the same molar concentrations, indicating that the short turn-forming sequence is more effective in the refolding process than the much longer C-peptide. Native human insulin was successfully generated by subsequent enzymic conversion of mini-proinsulin. The mini-proinsulin exhibited high receptor-binding activity, about 50% as potent as insulin, suggesting that this single-chained mini-proinsulin may provide a foundation in understanding the receptor-bound structure of insulin as well as the role of C-peptide in the folding and activity of proinsulin.

  19. van Hove singularities and tight-binding model in high-temperature superconductor H3Se

    Science.gov (United States)

    Cui, Huijuan; Li, Menglei; Zheng, Fawei; Zhang, Ping

    2017-08-01

    Ever since high-Tc superconductivity was found in H3S at 203 K under 200 GPa pressure, researches on hydrogen-rich family including H3S, H3P, and H3Te have been intensively carried out. Among those compounds, H3Se is a promising candidate of the same space group with H3S. In this work, we have thoroughly studied the electronic and phonon structures of H3Se using density functional theory (DFT) calculations. It is demonstrated that the electronic bands of H3Se contain van Hove singularities near the Fermi energy, which play a significant role in the superconductivity of this material. Besides, an accurate tight-binding model is proposed based on the Wannier function interpolations which can capture the main features of the electronic structures of H3Se. Moreover, we have also discussed the phonon dispersions and vibration modes for a better understanding of the phonon-electron interaction in H3Se.

  20. Spanning high-dimensional expression space using ribosome-binding site combinatorics.

    Science.gov (United States)

    Zelcbuch, Lior; Antonovsky, Niv; Bar-Even, Arren; Levin-Karp, Ayelet; Barenholz, Uri; Dayagi, Michal; Liebermeister, Wolfram; Flamholz, Avi; Noor, Elad; Amram, Shira; Brandis, Alexander; Bareia, Tasneem; Yofe, Ido; Jubran, Halim; Milo, Ron

    2013-05-01

    Protein levels are a dominant factor shaping natural and synthetic biological systems. Although proper functioning of metabolic pathways relies on precise control of enzyme levels, the experimental ability to balance the levels of many genes in parallel is a major outstanding challenge. Here, we introduce a rapid and modular method to span the expression space of several proteins in parallel. By combinatorially pairing genes with a compact set of ribosome-binding sites, we modulate protein abundance by several orders of magnitude. We demonstrate our strategy by using a synthetic operon containing fluorescent proteins to span a 3D color space. Using the same approach, we modulate a recombinant carotenoid biosynthesis pathway in Escherichia coli to reveal a diversity of phenotypes, each characterized by a distinct carotenoid accumulation profile. In a single combinatorial assembly, we achieve a yield of the industrially valuable compound astaxanthin 4-fold higher than previously reported. The methodology presented here provides an efficient tool for exploring a high-dimensional expression space to locate desirable phenotypes.

  1. Insecticidal 3-benzamido-N-phenylbenzamides specifically bind with high affinity to a novel allosteric site in housefly GABA receptors.

    Science.gov (United States)

    Ozoe, Yoshihisa; Kita, Tomo; Ozoe, Fumiyo; Nakao, Toshifumi; Sato, Kazuyuki; Hirase, Kangetsu

    2013-11-01

    γ-Aminobutyric acid (GABA) receptors (GABARs) are an important target for existing insecticides such as fiproles. These insecticides act as noncompetitive antagonists (channel blockers) for insect GABARs by binding to a site within the intrinsic channel of the GABAR. Recently, a novel class of insecticides, 3-benzamido-N-phenylbenzamides (BPBs), was shown to inhibit GABARs by binding to a site distinct from the site for fiproles. We examined the binding site of BPBs in the adult housefly by means of radioligand-binding and electrophysiological experiments. 3-Benzamido-N-(2,6-dimethyl-4-perfluoroisopropylphenyl)-2-fluorobenzamide (BPB 1) (the N-demethyl BPB) was a partial, but potent, inhibitor of [(3)H]4'-ethynyl-4-n-propylbicycloorthobenzoate (GABA channel blocker) binding to housefly head membranes, whereas the 3-(N-methyl)benzamido congener (the N-methyl BPB) had low or little activity. A total of 15 BPB analogs were tested for their abilities to inhibit [(3)H]BPB 1 binding to the head membranes. The N-demethyl analogs, known to be highly effective insecticides, potently inhibited the [(3)H]BPB 1 binding, but the N-methyl analogs did not even though they, too, are considered highly effective. [(3)H]BPB 1 equally bound to the head membranes from wild-type and dieldrin-resistant (rdl mutant) houseflies. GABA allosterically inhibited [(3)H]BPB 1 binding. By contrast, channel blocker-type antagonists enhanced [(3)H]BPB 1 binding to housefly head membranes by increasing the affinity of BPB 1. Antiparasitic macrolides, such as ivermectin B1a, were potent inhibitors of [(3)H]BPB 1 binding. BPB 1 inhibited GABA-induced currents in housefly GABARs expressed in Xenopus oocytes, whereas it failed to inhibit l-glutamate-induced currents in inhibitory l-glutamate receptors. Overall, these findings indicate that BPBs act at a novel allosteric site that is different from the site for channel blocker-type antagonists and that is probably overlapped with the site for macrolides

  2. Mutational analysis of the high-affinity zinc binding site validates a refined human dopamine transporter homology model.

    Directory of Open Access Journals (Sweden)

    Thomas Stockner

    Full Text Available The high-resolution crystal structure of the leucine transporter (LeuT is frequently used as a template for homology models of the dopamine transporter (DAT. Although similar in structure, DAT differs considerably from LeuT in a number of ways: (i when compared to LeuT, DAT has very long intracellular amino and carboxyl termini; (ii LeuT and DAT share a rather low overall sequence identity (22% and (iii the extracellular loop 2 (EL2 of DAT is substantially longer than that of LeuT. Extracellular zinc binds to DAT and restricts the transporter's movement through the conformational cycle, thereby resulting in a decrease in substrate uptake. Residue H293 in EL2 praticipates in zinc binding and must be modelled correctly to allow for a full understanding of its effects. We exploited the high-affinity zinc binding site endogenously present in DAT to create a model of the complete transmemberane domain of DAT. The zinc binding site provided a DAT-specific molecular ruler for calibration of the model. Our DAT model places EL2 at the transporter lipid interface in the vicinity of the zinc binding site. Based on the model, D206 was predicted to represent a fourth co-ordinating residue, in addition to the three previously described zinc binding residues H193, H375 and E396. This prediction was confirmed by mutagenesis: substitution of D206 by lysine and cysteine affected the inhibitory potency of zinc and the maximum inhibition exerted by zinc, respectively. Conversely, the structural changes observed in the model allowed for rationalizing the zinc-dependent regulation of DAT: upon binding, zinc stabilizes the outward-facing state, because its first coordination shell can only be completed in this conformation. Thus, the model provides a validated solution to the long extracellular loop and may be useful to address other aspects of the transport cycle.

  3. Mutational analysis of the high-affinity zinc binding site validates a refined human dopamine transporter homology model.

    Science.gov (United States)

    Stockner, Thomas; Montgomery, Therese R; Kudlacek, Oliver; Weissensteiner, Rene; Ecker, Gerhard F; Freissmuth, Michael; Sitte, Harald H

    2013-01-01

    The high-resolution crystal structure of the leucine transporter (LeuT) is frequently used as a template for homology models of the dopamine transporter (DAT). Although similar in structure, DAT differs considerably from LeuT in a number of ways: (i) when compared to LeuT, DAT has very long intracellular amino and carboxyl termini; (ii) LeuT and DAT share a rather low overall sequence identity (22%) and (iii) the extracellular loop 2 (EL2) of DAT is substantially longer than that of LeuT. Extracellular zinc binds to DAT and restricts the transporter's movement through the conformational cycle, thereby resulting in a decrease in substrate uptake. Residue H293 in EL2 praticipates in zinc binding and must be modelled correctly to allow for a full understanding of its effects. We exploited the high-affinity zinc binding site endogenously present in DAT to create a model of the complete transmemberane domain of DAT. The zinc binding site provided a DAT-specific molecular ruler for calibration of the model. Our DAT model places EL2 at the transporter lipid interface in the vicinity of the zinc binding site. Based on the model, D206 was predicted to represent a fourth co-ordinating residue, in addition to the three previously described zinc binding residues H193, H375 and E396. This prediction was confirmed by mutagenesis: substitution of D206 by lysine and cysteine affected the inhibitory potency of zinc and the maximum inhibition exerted by zinc, respectively. Conversely, the structural changes observed in the model allowed for rationalizing the zinc-dependent regulation of DAT: upon binding, zinc stabilizes the outward-facing state, because its first coordination shell can only be completed in this conformation. Thus, the model provides a validated solution to the long extracellular loop and may be useful to address other aspects of the transport cycle.

  4. The ion exchange properties and equilibrium constants of Li+, Na+ and K+ on zirconium phosphate highly dispersed on a cellulose acetate fibers surface

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    Borgo Claudemir Adriano

    2004-01-01

    Full Text Available Highly dispersed zirconium phosphate was prepared by reacting celullose acetate/ZrO2 (ZrO2 = 11 wt%, 1.0 mmol g-1 of zirconium atom per gram of the material with phosphoric acid. High power decoupling magic angle spinning (HPDEC-MAS 31P NMR and X-ray photoelectron spectroscopy data indicated that HPO4(2- is the species present on the membrane surface. The specific concentration of acidic centers, determined by ammonia gas adsorption, is 0.60 mmol g-1. The ion exchange capacities for Li+, Na+ and K+ ions were determined from ion exchange isotherms at 298 K and showed the following values (in mmol g-1: Li+= 0.05, Na+= 0.38 and K+= 0.57. Due to the strong cooperative effect, the H+/Na+ and H+/K+ ion exchange is of non ideal nature. These ion exchange equilibria were treated with the use of models of fixed tridentate centers, which consider the surface of the sorbent as polyfunctional sorption centers. Both the observed ion exchange capacities with respect to the alkaline metal ions and the equilibrium constants are discussed by taking into consideration the sequence of the ionic hydration radii for Li+, Na+ and K+. The matrix affinity for the ions decreases with increasing the cations hydration radii from K+ to Li+. The high values of the separation factors S Na+/Li+ and S K+/Li+ (up to several hundreds support the application of this material for the quantitative separation of Na+ and K+ from Li+ from a mixture containing these three ions.

  5. Organogel-assisted topochemical synthesis of multivalent glyco-polymer for high-affinity lectin binding.

    Science.gov (United States)

    Krishnan, Baiju P; Raghu, Sreedevi; Mukherjee, Somnath; Sureshan, Kana M

    2016-12-01

    An organogelator, 2,4-undeca-diynyl-4',6'-O-benzylidene-β-d-galactopyranoside, which aligns its diacetylene upon gelation, has been synthesized. UV irradiation of its gel resulted in topochemical polymerization of the gelator forming polydiacetylene (PDA). We have used this gel-state reaction for the synthesis of surface-immobilized multi-valent glycoclusters, which showed 1000-fold enhanced binding, compared to monomers, with various galactose-binding lectins.

  6. Recognition of anesthetic barbiturates by a protein binding site: a high resolution structural analysis.

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    Simon Oakley

    Full Text Available Barbiturates potentiate GABA actions at the GABA(A receptor and act as central nervous system depressants that can induce effects ranging from sedation to general anesthesia. No structural information has been available about how barbiturates are recognized by their protein targets. For this reason, we tested whether these drugs were able to bind specifically to horse spleen apoferritin, a model protein that has previously been shown to bind many anesthetic agents with affinities that are closely correlated with anesthetic potency. Thiopental, pentobarbital, and phenobarbital were all found to bind to apoferritin with affinities ranging from 10-500 µM, approximately matching the concentrations required to produce anesthetic and GABAergic responses. X-ray crystal structures were determined for the complexes of apoferritin with thiopental and pentobarbital at resolutions of 1.9 and 2.0 Å, respectively. These structures reveal that the barbiturates bind to a cavity in the apoferritin shell that also binds haloalkanes, halogenated ethers, and propofol. Unlike these other general anesthetics, however, which rely entirely upon van der Waals interactions and the hydrophobic effect for recognition, the barbiturates are recognized in the apoferritin site using a mixture of both polar and nonpolar interactions. These results suggest that any protein binding site that is able to recognize and respond to the chemically and structurally diverse set of compounds used as general anesthetics is likely to include a versatile mixture of both polar and hydrophobic elements.

  7. High-throughput 1,536-well fluorescence polarization assays for α(1-acid glycoprotein and human serum albumin binding.

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    Adam Yasgar

    Full Text Available Two major plasma proteins in humans are primarily responsible for drug binding, the α(1-acid-glycoprotein (AGP and human serum albumin (HSA. The availability of at least a semiquantitative high-throughput assay for assessment of protein binding is expected to aid in bridging the current gap between high-throughput screening and early lead discovery, where cell-based and biochemical assays are deployed routinely to test up to several million compounds rapidly, as opposed to the late-stage candidate drug profiling methods which test at most dozens of compounds at a time. Here, we describe the miniaturization of a pair of assays based on the binding- and displacement-induced changes in fluorescence polarization (FP of fluorescent small molecule probes known to specifically target the drug-binding sites of these two proteins. A robust and reproducible assay performance was achieved in ≤4 µL assay volume in 1,536-well format. The assays were tested against a validation set of 10 known protein binders, and the results compared favorably with data obtained using protein-coated beads with high-performance liquid chromatography analysis. The miniaturized assays were taken to a high-throughput level in a screen of the LOPAC(1280 collection of 1,280 pharmacologically active compounds. The adaptation of the AGP and HSA FP assays to a 1,536-well format should allow their use in early-stage profiling of large-size compound sets.

  8. A human β-III-spectrin spinocerebellar ataxia type 5 mutation causes high-affinity F-actin binding

    Science.gov (United States)

    Avery, Adam W.; Crain, Jonathan; Thomas, David D.; Hays, Thomas S.

    2016-01-01

    Spinocerebellar ataxia type 5 (SCA5) is a human neurodegenerative disease that stems from mutations in the SPTBN2 gene encoding the protein β-III-spectrin. Here we investigated the molecular consequence of a SCA5 missense mutation that results in a L253P substitution in the actin-binding domain (ABD) of β-III-spectrin. We report that the L253P substitution in the isolated β-III-spectrin ABD causes strikingly high F-actin binding affinity (Kd = 75.5 nM) compared to the weak F-actin binding affinity of the wild-type ABD (Kd = 75.8 μM). The mutation also causes decreased thermal stability (Tm = 44.6 °C vs 59.5 °C). Structural analyses indicate that leucine 253 is in a loop at the interface of the tandem calponin homology (CH) domains comprising the ABD. Leucine 253 is predicted to form hydrophobic contacts that bridge the CH domains. The decreased stability of the mutant indicates that these bridging interactions are probably disrupted, suggesting that the high F-actin binding affinity of the mutant is due to opening of the CH domain interface. These results support a fundamental role for leucine 253 in regulating opening of the CH domain interface and binding of the ABD to F-actin. This study indicates that high-affinity actin binding of L253P β-III-spectrin is a likely driver of neurodegeneration. PMID:26883385

  9. Cyr61/CCN1 displays high-affinity binding to the somatomedin B(1-44 domain of vitronectin.

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    Ivo M B Francischetti

    Full Text Available BACKGROUND: Cyr61 is a member of the CCN (Cyr61, connective tissue growth, NOV family of extracellular-associated (matricellular proteins that present four distinct functional modules, namely insulin-like growth factor binding protein (IGFBP, von Willebrand factor type C (vWF, thrombospondin type 1 (TSP, and C-terminal growth factor cysteine knot (CT domain. While heparin sulphate proteoglycans reportedly mediate the interaction of Cyr61 with the matrix and cell surface, the role of other extracellular associated proteins has not been revealed. METHODS AND FINDINGS: In this report, surface plasmon resonance (SPR experiments and solid-phase binding assays demonstrate that recombinant Cyr61 interacts with immobilized monomeric or multimeric vitronectin (VTNC with K(D in the nanomolar range. Notably, the binding site for Cyr61 was identified as the somatomedin B domain (SMTB(1-44 of VTNC, which mediates its interaction with PAI-1, uPAR, and integrin alphav beta3. Accordingly, PAI-1 outcompetes Cyr61 for binding to immobilized SMTB(1-44, and Cyr61 attenuates uPAR-mediated U937 adhesion to VTNC. In contrast, isothermal titration calorimetry shows that Cyr61 does not display high-affinity binding for SMTB(1-44 in solution. Nevertheless, competitive ELISA revealed that multimeric VTNC, heat-modified monomeric VTNC, or SMTB(1-44 at high concentrations attenuate Cyr61 binding to immobilized VTNC, while monomeric VTNC was ineffective. Therefore, immobilization of VTNC exposes cryptic epitopes that recognize Cyr61 with high affinity, as reported for a number of antibodies, beta-endorphin, and other molecules. CONCLUSIONS: The finding that Cyr61 interacts with the SMTB(1-44 domain suggests that VTNC represent a point of anchorage for CCN family members to the matrix. Results are discussed in the context of the role of CCN and VTNC in matrix biology and angiogenesis.

  10. Neuronal entry and high neurotoxicity of botulinum neurotoxin A require its N-terminal binding sub-domain.

    Science.gov (United States)

    Wang, Jiafu; Meng, Jianghui; Nugent, Marc; Tang, Minhong; Dolly, J Oliver

    2017-03-15

    Botulinum neurotoxins (BoNTs) are the most toxic proteins known, due to inhibiting the neuronal release of acetylcholine and causing flaccid paralysis. Most BoNT serotypes target neurons by binding to synaptic vesicle proteins and gangliosides via a C-terminal binding sub-domain (HCC). However, the role of their conserved N-terminal sub-domain (HCN) has not been established. Herein, we created a mutant form of recombinant BoNT/A lacking HCN (rAΔHCN) and showed that the lethality of this mutant is reduced 3.3 × 10(4)-fold compared to wild-type BoNT/A. Accordingly, low concentrations of rAΔHCN failed to bind either synaptic vesicle protein 2C or neurons, unlike the high-affinity neuronal binding obtained with (125)I-BoNT/A (Kd = 0.46 nM). At a higher concentration, rAΔHCN did bind to cultured sensory neurons and cluster on the surface, even after 24 h exposure. In contrast, BoNT/A became internalised and its light chain appeared associated with the plasmalemma, and partially co-localised with vesicle-associated membrane protein 2 in some vesicular compartments. We further found that a point mutation (W985L) within HCN reduced the toxicity over 10-fold, while this mutant maintained the same level of binding to neurons as wild type BoNT/A, suggesting that HCN makes additional contributions to productive internalization/translocation steps beyond binding to neurons.

  11. In vitro study of the binding between chlorpyrfos and sex hormones using headspace solid-phase microextraction combined with high-performance liquid chromatography: A new aspect of pesticides and breast cancer risk.

    Science.gov (United States)

    Farhadi, K; Tahmasebi, R; Biparva, P; Maleki, R

    2015-08-01

    Endocrine-disrupting chemicals are compounds that alter the normal functioning of the endocrine system. Organophosphorus insecticides, as chlorpyrifos (CPS), receive an increasing consideration as potential endocrine disrupters. Physiological estrogens, including estrone (E1), 17β-estradiol (E2), and diethylstilbestrol (DES) fluctuate with life stage, suggesting specific roles for them in biological and disease processes. There has been great interest in whether certain organophosphorus pesticides can affect the risk of breast cancer. An understanding of the interaction processes is the key to describe the fate of CPS in biological media. The objectives of this study were to evaluate total, bound, and freely dissolved amount of CPS in the presence of three estrogenic sex hormones (ESHs). In vitro experiments were conducted utilizing a headspace solid phase microextraction (HS-SPME) combined with high-performance liquid chromatography (HPLC) method. The obtained Scatchard plot based on the proposed SPME-HPLC method was employed to determine CPS-ESHs binding constant and the number of binding sites as well as binding percentage of each hormone to CPS. The number of binding sites per studied hormone molecule was 1.10, 1, and 0.81 for E1, E2, and DES, respectively. The obtained results confirmed that CPS bound to one class of binding sites on sex hormones.

  12. Sensitive and direct detection of receptor binding specificity of highly pathogenic avian influenza A virus in clinical samples.

    Directory of Open Access Journals (Sweden)

    Tadanobu Takahashi

    Full Text Available Influenza A virus (IAV recognizes two types of N-acetylneuraminic acid (Neu5Ac by galactose (Gal linkages, Neu5Acα2,3Gal and Neu5Acα2,6Gal. Avian IAV preferentially binds to Neu5Acα2,3Gal linkage, while human IAV preferentially binds to Neu5Acα2,6Gal linkage, as a virus receptor. Shift in receptor binding specificity of avian IAV from Neu5Acα2,3Gal linkage to Neu5Acα2,6Gal linkage is generally believed to be a critical factor for its transmission ability among humans. Surveillance of this shift of highly pathogenic H5N1 avian IAV (HPAI is thought to be a very important for prediction and prevention of a catastrophic pandemic of HPAI among humans. In this study, we demonstrated that receptor binding specificity of IAV bound to sialo-glycoconjugates was sensitively detected by quantifying the HA gene with real-time reverse-transcription-PCR. The new assay enabled direct detection of receptor binding specificity of HPAIs in chicken clinical samples including trachea and cloaca swabs in only less than 4 h.

  13. The Structure of a High-Affinity Kainate Receptor: GluK4 Ligand-Binding Domain Crystallized with Kainate.

    Science.gov (United States)

    Kristensen, Ole; Kristensen, Lise Baadsgaard; Møllerud, Stine; Frydenvang, Karla; Pickering, Darryl S; Kastrup, Jette Sandholm

    2016-09-01

    Ionotropic glutamate receptors play a key role in fast neurotransmission in the CNS and have been linked to several neurological diseases and disorders. One subfamily is the kainate receptors, which are grouped into low-affinity (GluK1-3) and high-affinity (GluK4-5) receptors based on their affinity for kainate. Although structures of the ligand-binding domain (LBD) of all low-affinity kainate receptors have been reported, no structures of the high-affinity receptor subunits are available. Here, we present the X-ray structure of GluK4-LBD with kainate at 2.05 Å resolution, together with thermofluor and radiolabel binding affinity data. Whereas binding-site residues in GluK4 are most similar to the AMPA receptor subfamily, the domain closure and D1-D2 interlobe contacts induced by kainate are similar to the low-affinity kainate receptor GluK1. These observations provide a likely explanation for the high binding affinity of kainate at GluK4-LBD.

  14. Viral reverse transcriptases show selective high affinity binding to DNA-DNA primer-templates that resemble the polypurine tract.

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    Gauri R Nair

    Full Text Available Previous results using a SELEX (Systematic Evolution of Ligands by Exponential Enrichment-based approach that selected DNA primer-template duplexes binding with high affinity to HIV reverse transcriptase (RT showed that primers mimicking the 3' end, and in particular the six nt terminal G tract, of the RNA polypurine tract (PPT; HIV PPT: 5'-AAAAGAAAAGGGGGG-3' were preferentially selected. In this report, two viral (Moloney murine leukemia virus (MuLV and avian myeloblastosis virus (AMV and one retrotransposon (Ty3 RTs were used for selection. Like HIV RT, both viral RTs selected duplexes with primer strands mimicking the G tract at the PPT 3' end (AMV PPT: 5'-AGGGAGGGGGA-3'; MuLV PPT: 5'-AGAAAAAGGGGGG-3'. In contrast, Ty3, whose PPT lacks a G tract (5'-GAGAGAGAGGAA-3' showed no selective binding to any duplex sequences. Experiments were also conducted with DNA duplexes (termed DNA PPTs mimicking the RNA PPT-DNA duplex of each virus and a control duplex with a random DNA sequence. Retroviral RTs bound with high affinity to all viral DNA PPT constructs, with HIV and MuLV RTs showing comparable binding to the counterpart DNA PPT duplexes and reduced affinity to the AMV DNA PPT. AMV RT showed similar behavior with a modest preference for its own DNA PPT. Ty3 RT showed no preferential binding for its own or any other DNA PPT and viral RTs bound the Ty3 DNA PPT with relatively low affinity. In contrast, binding affinity of HIV RT to duplexes containing the HIV RNA PPT was less dependent on the G tract, which is known to be pivotal for efficient extension. We hypothesize that the G tract on the RNA PPT helps shift the binding orientation of RT to the 3' end of the PPT where extension can occur.

  15. Combining RAFT polymerization and thiol-ene click reaction for core-shell structured polymer@BaTiO3 nanodielectrics with high dielectric constant, low dielectric loss, and high energy storage capability.

    Science.gov (United States)

    Yang, Ke; Huang, Xingyi; Zhu, Ming; Xie, Liyuan; Tanaka, Toshikatsu; Jiang, Pingkai

    2014-02-12

    Nanodielectric materials with high dielectric constant, low dielectric loss, and high energy storage capability are highly desirable in modern electric and electronics industries. It has been proved that the preparation of core-shell structured dielectric polymer nanocomposites via "grafting from" method is an effective approach to these materials. However, by using this approach, the deep understanding of the structure-dielectric property relationship of the core-shell structured nanodielectrics has been limited because of the lack of detailed information (e.g., molecular weight, grafting density) about the macromolecules grafted onto the nanoparticle surfaces. In this work, by the combination of reversible addition-fragmentation chain transfer (RAFT) polymerization and thiol-ene click reaction, two types of core-shell structured polymer@BaTiO3 (polymer@BT) nanocomposites with high dielectric constant and low dielectric loss were successfully prepared via a "grafting to" method. Compared with the "grafting from" method, this "grafting to" method has two merits: the molecular weight of the polymer chains in the shell layer can be easily controlled and the grafting density can be tailored by changing the molecular weight of the grafting polymer. Moreover, a clear insight into the relationship among the dielectric properties and energy storage capability of the core-shell structured polymer@BT nanocomposites, the molecular weight of the polymer chains, and the grafting density of the core-shell structured nanoparticles was achieved. The study provides new insights into the design and preparation of nanodielectric materials with desirable dielectric properties.

  16. Discovery of a cobalt complex with high MEK1 binding affinity.

    Science.gov (United States)

    Li, Hongyue; Zhou, Tongliang; Liu, Hui; Xu, Fengrong; Niu, Yan; Wang, Chao; Liang, Lei; Xu, Ping

    2017-05-15

    A series of Schiff base ligands (L(1)-L(5)) and their cobalt(II) complexes (1-5) were designed and synthesized for MEK1 binding experiment. The biological evaluation results showed that Bis(N,N'-disalicylidene)-3,4-phenylenediamine-cobalt(II) 1 and Bis(N,N'-disalicylidene)-1,2-cyclohexanediamine-cobalt(II) 2 are much more effective than the parent Schiff bases (L(1) and L(2)). Importantly, 2 exhibited MEK1 binding affinity with IC5071nM, which is so far the best result for metal complexes and more potent than U0126 (7.02μM) and AZD6244 (2.20μM). Docking study was used to elucidate the binding modes of complex 2 with MEK1. Thus cobalt(II) complex 2 may be further developed as a novel MEK1 inhibitor. Copyright © 2017 Elsevier Ltd. All rights reserved.

  17. Transcription factor binding sites are highly enriched within microRNA precursor sequences

    Directory of Open Access Journals (Sweden)

    Piriyapongsa Jittima

    2011-12-01

    Full Text Available Abstract Background Transcription factors are thought to regulate the transcription of microRNA genes in a manner similar to that of protein-coding genes; that is, by binding to conventional transcription factor binding site DNA sequences located in or near promoter regions that lie upstream of the microRNA genes. However, in the course of analyzing the genomics of human microRNA genes, we noticed that annotated transcription factor binding sites commonly lie within 70- to 110-nt long microRNA small hairpin precursor sequences. Results We report that about 45% of all human small hairpin microRNA (pre-miR sequences contain at least one predicted transcription factor binding site motif that is conserved across human, mouse and rat, and this rises to over 75% if one excludes primate-specific pre-miRs. The association is robust and has extremely strong statistical significance; it affects both intergenic and intronic pre-miRs and both isolated and clustered microRNA genes. We also confirmed and extended this finding using a separate analysis that examined all human pre-miR sequences regardless of conservation across species. Conclusions The transcription factor binding sites localized within small hairpin microRNA precursor sequences may possibly regulate their transcription. Transcription factors may also possibly bind directly to nascent primary microRNA gene transcripts or small hairpin microRNA precursors and regulate their processing. Reviewers This article was reviewed by Guillaume Bourque (nominated by Jerzy Jurka, Dmitri Pervouchine (nominated by Mikhail Gelfand, and Yuriy Gusev.

  18. Low density and high affinity of platelet [3H]paroxetine binding in women with bulimia nervosa.

    Science.gov (United States)

    Ekman, Agneta; Sundblad-Elverfors, Charlotta; Landén, Mikael; Eriksson, Tomas; Eriksson, Elias

    2006-06-15

    Impaired serotonin transmission has been suggested to be implicated in the pathophysiology of bulimia nervosa. As an indirect measure of brain serotonergic activity, the binding of tritiated ligands to platelet serotonin transporters has been studied in bulimia nervosa as well as in other putatively serotonin-related psychiatric disorders. In this study, the density and affinity of platelet serotonin transporters were assessed in 20 women meeting the DSM-IV criteria for bulimia nervosa and in 14 controls without previous or ongoing eating disorder using [(3)H]paroxetine as a ligand. In comparison to controls, women with bulimia nervosa had a significantly reduced number of platelet binding sites (B(max) = 721 +/- 313 vs. 1145 +/- 293 fmol/mg protein) and an increase in the affinity for the ligand demonstrated by a lower dissociaton constant (K(d) = 33 +/- 10 vs. 44 +/- 10 pM). A significant correlation between B(max) and K(d) values was found in patients but not in controls. Our results support the notion that bulimia nervosa is associated with a reduction in platelet serotonin transporter density. In addition, our study is the first to report that this reduced transporter density in women with bulimia nervosa is accompanied by an increase in the affinity of the transporter for the ligand.

  19. SPECTROSCOPIC CONSTANTS AND VIBRATIONAL FREQUENCIES FOR l-C{sub 3}H{sup +} AND ISOTOPOLOGUES FROM HIGHLY ACCURATE QUARTIC FORCE FIELDS: THE DETECTION OF l-C{sub 3}H{sup +} IN THE HORSEHEAD NEBULA PDR QUESTIONED

    Energy Technology Data Exchange (ETDEWEB)

    Huang Xinchuan [SETI Institute, 189 Bernardo Avenue, Suite 100, Mountain View, CA 94043 (United States); Fortenberry, Ryan C.; Lee, Timothy J., E-mail: Timothy.J.Lee@nasa.gov [NASA Ames Research Center, Moffett Field, CA 94035-1000 (United States)

    2013-05-10

    Very recently, molecular rotational transitions observed in the photon-dominated region of the Horsehead nebula have been attributed to l-C{sub 3}H{sup +}. In an effort to corroborate this finding, we employed state-of-the-art and proven high-accuracy quantum chemical techniques to compute spectroscopic constants for this cation and its isotopologues. Even though the B rotational constant from the fit of the observed spectrum and our computations agree to within 20 MHz, a typical level of accuracy, the D rotational constant differs by more than 40%, while the H rotational constant differs by three orders of magnitude. With the likely errors in the rotational transition energies resulting from this difference in D on the order of 1 MHz for the lowest observed transition (J = 4 {yields} 3) and growing as J increases, the assignment of the observed rotational lines from the Horsehead nebula to l-C{sub 3}H{sup +} is questionable.

  20. Variation of Fundamental Constants

    Science.gov (United States)

    Flambaum, V. V.

    2006-11-01

    Theories unifying gravity with other interactions suggest temporal and spatial variation of the fundamental ``constants'' in expanding Universe. The spatial variation can explain a fine tuning of the fundamental constants which allows humans (and any life) to appear. We appeared in the area of the Universe where the values of the fundamental constants are consistent with our existence. We present a review of recent works devoted to the variation of the fine structure constant α, strong interaction and fundamental masses. There are some hints for the variation in quasar absorption spectra. Big Bang nucleosynthesis, and Oklo natural nuclear reactor data. A very promising method to search for the variation of the fundamental constants consists in comparison of different atomic clocks. Huge enhancement of the variation effects happens in transition between accidentally degenerate atomic and molecular energy levels. A new idea is to build a ``nuclear'' clock based on the ultraviolet transition between very low excited state and ground state in Thorium nucleus. This may allow to improve sensitivity to the variation up to 10 orders of magnitude! Huge enhancement of the variation effects is also possible in cold atomic and molecular collisions near Feshbach resonance.

  1. Conversion of a putative Agrobacterium sugar-binding protein into a FRET sensor with high selectivity for sucrose.

    Science.gov (United States)

    Lager, Ida; Looger, Loren L; Hilpert, Melanie; Lalonde, Sylvie; Frommer, Wolf B

    2006-10-13

    Glucose is the main sugar transport form in animals, whereas plants use sucrose to supply non-photosynthetic organs with carbon skeletons and energy. Many aspects of sucrose transport, metabolism, and signaling are not well understood, including the route of sucrose efflux from leaf mesophyll cells and transport across vacuolar membranes. Tools that can detect sucrose with high spatial and temporal resolution in intact organs may help elucidate the players involved. Here, FRET sensors were generated by fusing putative sucrose-binding proteins to green fluorescent protein variants. Plant-associated bacteria such as Rhizobium and Agrobacterium can use sucrose as a nutrient source; sugar-binding proteins were, thus, used as scaffolds for developing sucrose nanosensors. Among a set of putative sucrose-binding protein genes cloned in between eCFP and eYFP and tested for sugar-dependent FRET changes, an Agrobacterium sugar-binding protein bound sucrose with 4 mum affinity. This FLIPsuc-4mu protein also recognized other sugars including maltose, trehalose, and turanose and, with lower efficiency, glucose and palatinose. Homology modeling enabled the prediction of binding pocket mutations to modulate the relative affinity of FLIPsuc-4mu for sucrose, maltose, and glucose. Mutant nanosensors showed up to 50- and 11-fold increases in specificity for sucrose over maltose and glucose, respectively, and the sucrose binding affinity was simultaneously decreased to allow detection in the physiological range. In addition, the signal-to-noise ratio of the sucrose nanosensor was improved by linker engineering. This novel reagent complements FLIPs for glucose, maltose, ribose, glutamate, and phosphate and will be used for analysis of sucrose-derived carbon flux in bacterial, fungal, plant, and animal cells.

  2. High-Throughput Testing of Antibody-Dependent Binding Inhibition of Placental Malaria Parasites

    DEFF Research Database (Denmark)

    Nielsen, Morten A; Salanti, Ali

    2015-01-01

    The particular virulence of Plasmodium falciparum manifests in diverse severe malaria syndromes as cerebral malaria, severe anemia and placental malaria. The cause of both the severity and the diversity of infection outcome, is the ability of the infected erythrocyte (IE) to bind a range......-throughput assay used in the preclinical and clinical development of a VAR2CSA based vaccine against placental malaria....

  3. High-affinity small molecule-phospholipid complex formation: binding of siramesine to phosphatidicacid

    DEFF Research Database (Denmark)

    Khandelia, Himanshu

    2008-01-01

    , comparable to the affinities for the binding of small molecule ligands to proteins, was measured for phosphatidic acid (PA, mole fraction of XPA ) 0.2 in phosphatidylcholine vesicles), yielding a molecular partition coefficient of 240 ( 80 × 106. An MD simulation on the siramesine:PA interaction...

  4. Peptide Binding to HLA Class I Molecules: Homogenous, High-Throughput Screening, and Affinity Assays

    DEFF Research Database (Denmark)

    Harndahl, Mikkel; Justesen, Sune Frederik Lamdahl; Lamberth, Kasper;

    2009-01-01

    present a homogenous, proximity-based assay for detection of peptide binding to HLA class I molecules. It uses a conformation-dependent anti-HLA class I antibody, W6/32, as one tag and a biotinylated recombinant HLA class I molecule as the other tag, and a proximity-based signal is generated through...

  5. Development of a Novel Nonradiometric Assay for Nucleic Acid Binding to TDP-43 Suitable for High-Throughput Screening Using AlphaScreen® Technology

    OpenAIRE

    Cassel, Joel A.; Blass, Benjamin E.; Reitz, Allen B.; Pawlyk, Aaron C.

    2010-01-01

    TAR DNA binding protein 43 (TDP-43) is a nucleic acid binding protein that is associated with the pathology of cystic fibrosis and neurodegenerative diseases such as amyotrophic lateral sclerosis and frontotemporal lobar dementia. We have developed a robust, quantitative, nonradiometric high-throughput assay measuring oligonucleotide binding to TDP-43 using AlphaScreen® technology. Biotinylated single-stranded TAR DNA (bt-TAR-32) and 6 TG repeats (bt-TG6) bound with high affinity to TDP-43, w...

  6. Elastic constants of calcite

    Science.gov (United States)

    Peselnick, L.; Robie, R.A.

    1962-01-01

    The recent measurements of the elastic constants of calcite by Reddy and Subrahmanyam (1960) disagree with the values obtained independently by Voigt (1910) and Bhimasenachar (1945). The present authors, using an ultrasonic pulse technique at 3 Mc and 25??C, determined the elastic constants of calcite using the exact equations governing the wave velocities in the single crystal. The results are C11=13.7, C33=8.11, C44=3.50, C12=4.82, C13=5.68, and C14=-2.00, in units of 1011 dyncm2. Independent checks of several of the elastic constants were made employing other directions and polarizations of the wave velocities. With the exception of C13, these values substantially agree with the data of Voigt and Bhimasenachar. ?? 1962 The American Institute of Physics.

  7. Algorithm for structure constants

    CERN Document Server

    Paiva, F M

    2011-01-01

    In a $n$-dimensional Lie algebra, random numerical values are assigned by computer to $n(n-1)$ especially selected structure constants. An algorithm is then created, which calculates without ambiguity the remaining constants, obeying the Jacobi conditions. Differently from others, this algorithm is suitable even for poor personal computer. ------------- En $n$-dimensia algebro de Lie, hazardaj numeraj valoroj estas asignitaj per komputilo al $n(n-1)$ speciale elektitaj konstantoj de strukturo. Tiam algoritmo estas kreita, kalkulante senambigue la ceterajn konstantojn, obeante kondicxojn de Jacobi. Malsimile al aliaj algoritmoj, tiu cxi tauxgas ecx por malpotenca komputilo.

  8. Radiographic constant exposure technique

    DEFF Research Database (Denmark)

    Domanus, Joseph Czeslaw

    1985-01-01

    The constant exposure technique has been applied to assess various industrial radiographic systems. Different X-ray films and radiographic papers of two producers were compared. Special attention was given to fast film and paper used with fluorometallic screens. Radiographic image quality...... was tested by the use of ISO wire IQI's and ASTM penetrameters used on Al and Fe test plates. Relative speed and reduction of kilovoltage obtained with the constant exposure technique were calculated. The advantages of fast radiographic systems are pointed out...

  9. The aliquot constant

    CERN Document Server

    Bosma, Wieb

    2009-01-01

    The average value of log s(n)/n taken over the first N even integers is shown to converge to a constant lambda when N tends to infinity; moreover, the value of this constant is approximated and proven to be less than 0. Here s(n) sums the divisors of n less than n. Thus the geometric mean of s(n)/n, the growth factor of the function s, in the long run tends to be less than 1. This could be interpreted as probabilistic evidence that aliquot sequences tend to remain bounded.

  10. In vivo Target Residence Time and Kinetic Selectivity: The Association Rate Constant as Determinant.

    Science.gov (United States)

    de Witte, Wilhelmus E A; Danhof, Meindert; van der Graaf, Piet H; de Lange, Elizabeth C M

    2016-10-01

    It is generally accepted that, in conjunction with pharmacokinetics, the first-order rate constant of target dissociation is a major determinant of the time course and duration of in vivo target occupancy. Here we show that the second-order rate constant of target association can be equally important. On the basis of the commonly used mathematical models for drug-target binding, it is shown that a high target association rate constant can increase the (local) concentration of the drug, which decreases the rate of decline of target occupancy. The increased drug concentration can also lead to increased off-target binding and decreased selectivity. Therefore, the kinetics of both target association and dissociation need to be taken into account in the selection of drug candidates with optimal pharmacodynamic properties. Copyright © 2016 Elsevier Ltd. All rights reserved.

  11. dPeak: high resolution identification of transcription factor binding sites from PET and SET ChIP-Seq data.

    Directory of Open Access Journals (Sweden)

    Dongjun Chung

    Full Text Available Chromatin immunoprecipitation followed by high throughput sequencing (ChIP-Seq has been successfully used for genome-wide profiling of transcription factor binding sites, histone modifications, and nucleosome occupancy in many model organisms and humans. Because the compact genomes of prokaryotes harbor many binding sites separated by only few base pairs, applications of ChIP-Seq in this domain have not reached their full potential. Applications in prokaryotic genomes are further hampered by the fact that well studied data analysis methods for ChIP-Seq do not result in a resolution required for deciphering the locations of nearby binding events. We generated single-end tag (SET and paired-end tag (PET ChIP-Seq data for σ⁷⁰ factor in Escherichia coli (E. coli. Direct comparison of these datasets revealed that although PET assay enables higher resolution identification of binding events, standard ChIP-Seq analysis methods are not equipped to utilize PET-specific features of the data. To address this problem, we developed dPeak as a high resolution binding site identification (deconvolution algorithm. dPeak implements a probabilistic model that accurately describes ChIP-Seq data generation process for both the SET and PET assays. For SET data, dPeak outperforms or performs comparably to the state-of-the-art high-resolution ChIP-Seq peak deconvolution algorithms such as PICS, GPS, and GEM. When coupled with PET data, dPeak significantly outperforms SET-based analysis with any of the current state-of-the-art methods. Experimental validations of a subset of dPeak predictions from σ⁷⁰ PET ChIP-Seq data indicate that dPeak can estimate locations of binding events with as high as 2 to 21 bp resolution. Applications of dPeak to σ⁷⁰ ChIP-Seq data in E. coli under aerobic and anaerobic conditions reveal closely located promoters that are differentially occupied and further illustrate the importance of high resolution analysis of ChIP-Seq data.

  12. No difference in high-magnification morphology and hyaluronic acid binding in the selection of euploid spermatozoa with intact DNA

    Institute of Scientific and Technical Information of China (English)

    Suchada Mongkolchaipak; Teraporn Vutyavanich

    2013-01-01

    In this study,we compared conventional sperm selection with high-magnification morphology based on the motile sperm organellar morphology examination (MSOME) criteria,and hyaluronic acid (HA) binding for sperm chromosome aneuploidy and DNA fragmentation rates.Semen from 50 severe male factor cases was processed through density gradient centrifugation,and subjected to sperm selection by using the conventional method (control),high magnification at x 6650 or HA binding.Aneuploidy was detected by fluorescence in situ hybridization with probes for chromosomes 13,18,21,X and Y,and DNA fragmentation by the terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL) method.Spermatozoa selected under high-magnification had a lower DNA fragmentation rate (2.6% vs.1.7%; P=0.032),with no significant difference in aneuploidy rate (0.8% vs 0.7%; P=0.583),than those selected by the HA binding method.Spermatozoa selected by both methods had much lower aneuploidy and DNA fragmentation rate than the controls (7% aneuploidy and 26.8% DNA fragmentation rates,respectively).In the high-magnification group,the aneuploidy rate was lower when the best spermatozoa were selected than when only the second-best spermatozoa were available for selection,but the DNA fragmentation rate was not different.In conclusion,sperm selection under high magnification was more effective than under HA binding in selecting spermatozoa with low DNA fragmentation rate,but the small difference (0.9%) might not be clinically meaningful.Both methods were better than the conventional method of sperm selection.

  13. CD14 mediates binding of high doses of LPS but is dispensable for TNF-α production.

    Science.gov (United States)

    Borzęcka, Kinga; Płóciennikowska, Agnieszka; Björkelund, Hanna; Sobota, Andrzej; Kwiatkowska, Katarzyna

    2013-01-01

    Activation of macrophages with lipopolysaccharide (LPS) involves a sequential engagement of serum LPS-binding protein (LBP), plasma membrane CD14, and TLR4/MD-2 signaling complex. We analyzed participation of CD14 in TNF-α production stimulated with 1-1000 ng/mL of smooth or rough LPS (sLPS or rLPS) and in sLPS binding to RAW264 and J744 cells. CD14 was indispensable for TNF-α generation induced by a low concentration, 1 ng/mL, of sLPS and rLPS. At higher doses of both LPS forms (100-1000 ng/mL), TNF-α release required CD14 to much lower extent. Among the two forms of LPS, rLPS-induced TNF-α production was less CD14-dependent and could proceed in the absence of serum as an LBP source. On the other hand, the involvement of CD14 was crucial for the binding of 1000 ng/mL of sLPS judging from an inhibitory effect of the anti-CD14 antibody. The binding of sLPS was also strongly inhibited by dextran sulfate, a competitive ligand of scavenger receptors (SR). In the presence of dextran sulfate, sLPS-induced production of TNF-α was upregulated about 1.6-fold. The data indicate that CD14 together with SR participates in the binding of high doses of sLPS. However, CD14 contribution to TNF α production induced by high concentrations of sLPS and rLPS can be limited.

  14. High level theoretical study of binding and of the potential energy surface in benzene-hydride system

    Energy Technology Data Exchange (ETDEWEB)

    Coletti, Cecilia, E-mail: ccoletti@unich.it [Dipartimento di Scienze del Farmaco, Universita ' G. d' Annunzio' Chieti-Pescara, Via dei Vestini 31, 66100 Chieti (Italy); Re, Nazzareno [Dipartimento di Scienze del Farmaco, Universita ' G. d' Annunzio' Chieti-Pescara, Via dei Vestini 31, 66100 Chieti (Italy)

    2012-04-04

    Graphical abstract: In-plane minimum geometries for benzene-H{sup -} non-covalent adducts: linear adduct (left) with the hydride ion hydrogen bonded to one aromatic hydrogen; bifurcated adduct (right), with the hydride ion hydrogen bonded to two adjacent aromatic hydrogens. Highlights: Black-Right-Pointing-Pointer Theoretical study on covalent and non-covalent binding in benzene-hydride. Black-Right-Pointing-Pointer Two non-covalent stable adducts were characterized in the in-plane geometry. Black-Right-Pointing-Pointer Significant sections of the potential energy surface were determined. Black-Right-Pointing-Pointer Formation of a very stable C{sub 6}H{sub 7}{sup -} anion upon covalent binding to carbon. - Abstract: High level ab initio calculations were performed on the interaction of the hydride anion with benzene, a system of potential interest for modelling the interactions occurring in hydrogen rich planetary atmospheres. We investigated both non-covalent and covalent binding, exploring the complete basis set limit using highly correlated MP2 and CCSD(T) levels of theory. Two non-covalent minima on the potential energy surface have been characterized, and found to correspond to moderately strong hydrogen bonding interactions. To gain further insight on the nature of binding, the total interaction energy was decomposed into its physically meaningful components and selected sections of the potential energy surface were calculated. Moreover, we found that H{sup -} can easily covalently bind to one of the carbon atoms of benzene to form a stable C{sub 6}H{sub 7}{sup -} anion, a global minimum on the potential energy surface, characterized by a puckered geometry, with a carbon atom bending out of the benzene plane. A slightly less stable planar C{sub 6}H{sub 7}{sup -} structure was also identified, corresponding to the transition state for the flipping motion of the puckered species.

  15. A Peptide Mimetic of 5-Acetylneuraminic Acid-Galactose Binds with High Avidity to Siglecs and NKG2D.

    Directory of Open Access Journals (Sweden)

    Laura L Eggink

    Full Text Available We previously identified several peptide sequences that mimicked the terminal sugars of complex glycans. Using plant lectins as analogs of lectin-type cell-surface receptors, a tetravalent form of a peptide with the sequence NPSHPLSG, designated svH1C, bound with high avidity to lectins specific for glycans with terminal 5-acetylneuraminic acid (Neu5Ac-galactose (Gal/N-acetylgalactosamine (GalNAc sequences. In this report, we show by circular dichroism and NMR spectra that svH1C lacks an ordered structure and thus interacts with binding sites from a flexible conformation. The peptide binds with high avidity to several recombinant human siglec receptors that bind preferentially to Neu5Ac(α2,3Gal, Neu5Ac(α2,6GalNAc or Neu5Ac(α2,8Neu5Ac ligands. In addition, the peptide bound the receptor NKG2D, which contains a lectin-like domain that binds Neu5Ac(α2,3Gal. The peptide bound to these receptors with a KD in the range of 0.6 to 1 μM. Binding to these receptors was inhibited by the glycoprotein fetuin, which contains multiple glycans that terminate in Neu5Ac(α2,3Gal or Neu5Ac(α2,6Gal, and by sialyllactose. Binding of svH1C was not detected with CLEC9a, CLEC10a or DC-SIGN, which are lectin-type receptors specific for other sugars. Incubation of neuraminidase-treated human peripheral blood mononuclear cells with svH1C resulted in binding of the peptide to a subset of the CD14+ monocyte population. Tyrosine phosphorylation of siglecs decreased dramatically when peripheral blood mononuclear cells were treated with 100 nM svH1C. Subcutaneous, alternate-day injections of svH1C into mice induced several-fold increases in populations of several types of immune cells in the peritoneal cavity. These results support the conclusion that svH1C mimics Neu5Ac-containing sequences and interacts with cell-surface receptors with avidities sufficient to induce biological responses at low concentrations. The attenuation of inhibitory receptors suggests that svH1C

  16. Ab initio design of Ca-decorated organic frameworks for high capacity molecular hydrogen storage with enhanced binding

    Science.gov (United States)

    Sun, Y. Y.; Lee, Kyuho; Kim, Yong-Hyun; Zhang, S. B.

    2009-07-01

    Ab initio calculations show that Ca can decorate organic linkers of metal-organic framework, MOF-5, with a binding energy of 1.25 eV. The Ca-decorated MOF-5 can store molecular hydrogen (H2) in both high gravimetric (4.6 wt %) and high volumetric (36 g/l) capacities. Even higher capacities (5.7 wt % and 45 g/l) can be obtained in a rationally designed covalent organic framework system, COF-α, with decorated Ca. Both density functional theory and second-order Møller-Plesset perturbation calculations show that the H2 binding in these systems is significantly stronger than the van der Waals interactions, which is required for H2 storage at near ambient conditions.

  17. Compassion is a constant.

    Science.gov (United States)

    Scott, Tricia

    2015-11-01

    Compassion is a powerful word that describes an intense feeling of commiseration and a desire to help those struck by misfortune. Most people know intuitively how and when to offer compassion to relieve another person's suffering. In health care, compassion is a constant; it cannot be rationed because emergency nurses have limited time or resources to manage increasing demands.

  18. Norfloxacin binds to human fecal material.

    Science.gov (United States)

    Edlund, C; Lindqvist, L; Nord, C E

    1988-01-01

    Earlier studies have reported very high (120 to 2,700 mg/kg) concentrations of norfloxacin in feces after therapeutic doses. MICs for fecal microorganisms are with few exceptions far below these levels. Nevertheless, clinical investigations show that the main part of the aerobic gram-positive and the anaerobic microflora remains unaffected after norfloxacin administration. In this study, the binding of [14C]norfloxacin to fecal material was analyzed. The binding of a group of nonlabeled quinolones to feces and the interactions between Enterococcus faecium, Bacteroides fragilis, and norfloxacin were also investigated. The results showed that norfloxacin has the ability to bind to feces. The specific binding was reversible, saturated after 90 min of incubation at 37 degrees C, and increased linearly with fecal concentration. Scatchard plots and nonlinear regression computer analyses revealed two different binding classes. The primary specific binding had a dissociation constant (KD) of 1.0 microM and a maximal binding capacity (Bmax) of 0.12 mumol/g of feces. The KD and Bmax of the secondary, more unspecific binding were 450 microM and 11.8 mumol/g of feces, respectively. The binding of unlabeled ciprofloxacin, enoxacin, ofloxacin, pefloxacin, and norfloxacin to feces was comparable to that of [14C]norfloxacin. The results of norfloxacin binding to suspensions of B. fragilis suggested that the main part of the binding is to the bacterial fraction of feces. In the presence of 8.0 g (dry weight) of B. fragilis per liter, the MBC of norfloxacin for E. faecium increased from 8 to 256 micrograms/ml. The finding of the present study indicated that binding of norfloxacin to feces may explain the paradox of high fecal concentrations of norfloxacin versus the actual effect on the normal gastrointestinal microflora. PMID:2854456

  19. The 1% concordance Hubble constant

    Energy Technology Data Exchange (ETDEWEB)

    Bennett, C. L.; Larson, D.; Weiland, J. L. [Johns Hopkins University, 3400 North Charles Street, Baltimore, MD 21218 (United States); Hinshaw, G., E-mail: cbennett@jhu.edu [Department of Physics and Astronomy, University of British Columbia, Vancouver, BC V6T 1Z1 (Canada)

    2014-10-20

    The determination of the Hubble constant has been a central goal in observational astrophysics for nearly a hundred years. Extraordinary progress has occurred in recent years on two fronts: the cosmic distance ladder measurements at low redshift and cosmic microwave background (CMB) measurements at high redshift. The CMB is used to predict the current expansion rate through a best-fit cosmological model. Complementary progress has been made with baryon acoustic oscillation (BAO) measurements at relatively low redshifts. While BAO data do not independently determine a Hubble constant, they are important for constraints on possible solutions and checks on cosmic consistency. A precise determination of the Hubble constant is of great value, but it is more important to compare the high and low redshift measurements to test our cosmological model. Significant tension would suggest either uncertainties not accounted for in the experimental estimates or the discovery of new physics beyond the standard model of cosmology. In this paper we examine in detail the tension between the CMB, BAO, and cosmic distance ladder data sets. We find that these measurements are consistent within reasonable statistical expectations and we combine them to determine a best-fit Hubble constant of 69.6 ± 0.7 km s{sup –1} Mpc{sup –1}. This value is based upon WMAP9+SPT+ACT+6dFGS+BOSS/DR11+H {sub 0}/Riess; we explore alternate data combinations in the text. The combined data constrain the Hubble constant to 1%, with no compelling evidence for new physics.

  20. High resolution crystal structures of unliganded and liganded human liver ACBP reveal a new mode of binding for the acyl-CoA ligand

    DEFF Research Database (Denmark)

    Taskinen, Jukka P; van Aalten, Daan M; Knudsen, Jens;

    2007-01-01

    The acyl-CoA binding protein (ACBP) is essential for the fatty acid metabolism, membrane structure, membrane fusion, and ceramide synthesis. Here high resolution crystal structures of human cytosolic liver ACBP, unliganded and liganded with a physiological ligand, myristoyl-CoA are described....... The binding of the acyl-CoA molecule induces only few structural differences near the binding pocket. The crystal form of the liganded ACBP, which has two ACBP molecules in the asymmetric unit, shows that in human ACBP the same acyl-CoA binding pocket is present as previously described for the bovine...... and Plasmodium falciparum ACBP and the mode of binding of the 3'-phosphate-AMP moiety is conserved. Unexpectedly, in one of the acyl-CoA binding pockets the acyl moiety is bound in a reversed mode as compared with the bovine and P. falciparum structures. In this binding mode, the myristoyl-CoA molecule is fully...

  1. Efficient and cost-effective experimental determination of kinetic constants and data: the success of a Bayesian systematic approach to drug transport, receptor binding, continuous culture and cell transport kinetics.

    Science.gov (United States)

    Murphy, Emma F; Gilmour, Steven G; Crabbe, M James C

    2004-01-02

    Details about the parameters of kinetic systems are crucial for progress in both medical and industrial research, including drug development, clinical diagnosis and biotechnology applications. Such details must be collected by a series of kinetic experiments and investigations. The correct design of the experiment is essential to collecting data suitable for analysis, modelling and deriving the correct information. We have developed a systematic and iterative Bayesian method and sets of rules for the design of enzyme kinetic experiments. Our method selects the optimum design to collect data suitable for accurate modelling and analysis and minimises the error in the parameters estimated. The rules select features of the design such as the substrate range and the number of measurements. We show here that this method can be directly applied to the study of other important kinetic systems, including drug transport, receptor binding, microbial culture and cell transport kinetics. It is possible to reduce the errors in the estimated parameters and, most importantly, increase the efficiency and cost-effectiveness by reducing the necessary amount of experiments and data points measured.

  2. VHS domains of ESCRT-0 cooperate in high-avidity binding to polyubiquitinated cargo

    Energy Technology Data Exchange (ETDEWEB)

    Ren, Xuefeng; Hurley, James H. (NIH)

    2010-03-30

    VHS (Vps27, Hrs, and STAM) domains occur in ESCRT-0 subunits Hrs and STAM, GGA adapters, and other trafficking proteins. The structure of the STAM VHS domain-ubiquitin complex was solved at 2.6 {angstrom} resolution, revealing that determinants for ubiquitin recognition are conserved in nearly all VHS domains. VHS domains from all classes of VHS-domain containing proteins in yeast and humans, including both subunits of ESCRT-0, bound ubiquitin in vitro. ESCRTs have been implicated in the sorting of Lys63-linked polyubiquitinated cargo. Intact human ESCRT-0 binds Lys63-linked tetraubiquitin 50-fold more tightly than monoubiquitin, though only 2-fold more tightly than Lys48-linked tetraubiquitin. The gain in affinity is attributed to the cooperation of flexibly connected VHS and UIM motifs of ESCRT-0 in avid binding to the polyubiquitin chain. Mutational analysis of all the five ubiquitin-binding sites in yeast ESCRT-0 shows that cooperation between them is required for the sorting of the Lys63-linked polyubiquitinated cargo Cps1 to the vacuole.

  3. Radiosynthesis and Evaluation of [(11)C]3-Hydroxycyclopent-1-enecarboxylic Acid as Potential PET Ligand for the High-Affinity γ-Hydroxybutyric Acid Binding Sites

    DEFF Research Database (Denmark)

    Jensen, Claus H; Hansen, Hanne D; Bay, Tina

    2017-01-01

    the (11)C-labeling and subsequent evaluation of [(11)C]HOCPCA in a domestic pig, as a PET-radioligand for visualization of the high-affinity GHB binding sites in the live pig brain. To investigate the regional binding of HOCPCA in pig brain prior to in vivo PET studies, in vitro quantitative......γ-Hydroxybutyric acid (GHB) is an endogenous neuroactive substance and proposed neurotransmitter with affinity for both low- and high-affinity binding sites. A radioligand with high and specific affinity toward the high-affinity GHB binding site would be a unique tool toward a more complete...... understanding of this population of binding sites. With its high specific affinity and monocarboxylate transporter (MCT1) mediated transport across the blood-brain barrier in pharmacological doses, 3-hydroxycyclopent-1-enecarboxylic acid (HOCPCA) seems like a suitable PET radiotracer candidate. Here, we report...

  4. Positive allosteric modulation of the GHB high-affinity binding site by the GABAA receptor modulator monastrol and the flavonoid catechin

    DEFF Research Database (Denmark)

    Eghorn, Laura Friis; Høstgaard-Jensen, Kirsten; Kongstad, Kenneth Thermann

    2014-01-01

    conformational changes in the binding site, demonstrating a positive allosteric modulation of radioligand binding. Surprisingly, binding of [3H]GHB and the GHB high-affinity site-specific radioligands [125I]BnOPh-GHB and [3H]HOCPCA was either decreased or only weakly increased, indicating that the observed......γ-Hydroxybutyric acid (GHB) is a metabolite of γ-aminobutyric acid (GABA) and a proposed neurotransmitter in the mammalian brain. We recently identified α4βδ GABAA receptors as possible high-affinity GHB targets. GABAA receptors are highly sensitive to allosteric modulation. Thus to investigate...... whether GHB high-affinity binding sites are also sensitive to allosteric modulation, we screened both known GABAA receptor ligands and a library of natural compounds in the rat cortical membrane GHB specific high-affinity [3H]NCS-382 binding assay. Two hits were identified: Monastrol, a positive...

  5. Role of phospholipids in drug-LDL bindings as studied by high-performance frontal analysis/capillary electrophoresis.

    Science.gov (United States)

    Kuroda, Yukihiro; Watanabe, Yoshinori; Shibukawa, Akimasa; Nakagawa, Terumichi

    2003-01-15

    The binding study between basic drugs ((S)-verapamil (VER) and (S)-propranolol (PRO)) and phospholipid liposomes was performed by using high-performance frontal analysis/capillary electrophoresis (HPFA/CE) in order to investigate the effect of oxidative modification of low-density lipoprotein (LDL) upon drug-binding affinity from molecule-based viewpoint. 1-Palmitoyl-2-oleoyl-phosphatidylcholine (POPC, 16:0, 18:1), 1-palmitoyl-2-linoleoyl-phosphatidylcholine (PLPC, 16:0, 18:2), dilauloyl-phosphatidylcholine (DLaPC, 12:0, 12:0), 1-palmitoyl-2-oleoyl-phosphatidyl-glycerol (POPG, 16:0, 18:1), and 1-palmitoyl-sn-glycero-3-phosphocholine (monoPPC, 16:0) were used to prepare the model liposomes. At physiological pH (pH 7.4), the model liposome prepared from POPG+POPC had negative net charges, while the total net charge of the other model liposomes (POPC liposome, PLPC liposome, DLaPC liposome, and monoPPC+POPC liposome) was zero. The drug and the model liposome mixed solutions were subjected to HPFA/CE, and the total binding affinities (nK) were calculated. The nK values of VER and PRO to POPG+POPC liposome were more than six and 10 times higher than those of other liposomes, respectively. On the other hand, the nK values of the model drugs to POPC liposome, PLPC liposome, DLaPC liposome and monoPPC+POPC liposome showed small differences less than twice. These results indicate that the electrostatic interaction plays an important effect on drug-liposome binding, and suggest that the increase in the negative charge of LDL phospholipids gives more significant effect on the drug-binding affinity of the basic drugs than the acyl-chain structure.

  6. Bioinformatic analysis of CaBP/calneuron proteins reveals a family of highly conserved vertebrate Ca2+-binding proteins

    Directory of Open Access Journals (Sweden)

    Burgoyne Robert D

    2010-04-01

    Full Text Available Abstract Background Ca2+-binding proteins are important for the transduction of Ca2+ signals into physiological outcomes. As in calmodulin many of the Ca2+-binding proteins bind Ca2+ through EF-hand motifs. Amongst the large number of EF-hand containing Ca2+-binding proteins are a subfamily expressed in neurons and retinal photoreceptors known as the CaBPs and the related calneuron proteins. These were suggested to be vertebrate specific but exactly which family members are expressed outside of mammalian species had not been examined. Findings We have carried out a bioinformatic analysis to determine when members of this family arose and the conserved aspects of the protein family. Sequences of human members of the family obtained from GenBank were used in Blast searches to identify corresponding proteins encoded in other species using searches of non-redundant proteins, genome sequences and mRNA sequences. Sequences were aligned and compared using ClustalW. Some families of Ca2+-binding proteins are known to show a progressive expansion in gene number as organisms increase in complexity. In contrast, the results for CaBPs and calneurons showed that a full complement of CaBPs and calneurons are present in the teleost fish Danio rerio and possibly in cartilaginous fish. These findings suggest that the entire family of genes may have arisen at the same time during vertebrate evolution. Certain members of the family (for example the short form of CaBP1 and calneuron 1 are highly conserved suggesting essential functional roles. Conclusions The findings support the designation of the calneurons as a distinct sub-family. While the gene number for CaBPs/calneurons does not increase, a distinctive evolutionary change in these proteins in vertebrates has been an increase in the number of splice variants present in mammals.

  7. Unusually strong binding to the DNA minor groove by a highly twisted benzimidazole diphenylether: induced fit and bound water.

    Science.gov (United States)

    Tanious, Farial A; Laine, William; Peixoto, Paul; Bailly, Christian; Goodwin, Kristie D; Lewis, Mark A; Long, Eric C; Georgiadis, Millie M; Tidwell, Richard R; Wilson, W David

    2007-06-12

    RT29 is a dicationic diamidine derivative that does not obey the classical "rules" for shape and functional group placement that are expected to result in strong binding and specific recognition of the DNA minor groove. The compound contains a benzimidazole diphenyl ether core that is flanked by the amidine cations. The diphenyl ether is highly twisted and gives the entire compound too much curvature to fit well to the shape of the minor groove. DNase I footprinting, fluorescence intercalator displacement studies, and circular dichroism spectra, however, indicate that the compound is an AT specific minor groove binding agent. Even more surprisingly, quantitative biosensor-surface plasmon resonance and isothermal titration calorimetric results indicate that the compound binds with exceptional strength to certain AT sequences in DNA with a large negative enthalpy of binding. Crystallographic results for the DNA complex of RT29 compared to calculated results for the free compound show that the compound undergoes significant conformational changes to enhance its minor groove interactions. In addition, a water molecule is incorporated directly into the complex to complete the compound-DNA interface, and it forms an essential link between the compound and base pair edges at the floor of the minor groove. The calculated DeltaCp value for complex formation is substantially less than the experimentally observed value, which supports the idea of water being an intrinsic part of the complex with a major contribution to the DeltaCp value. Both the induced fit conformational changes of the compound and the bound water are essential for strong binding to DNA by RT29.

  8. High Dielectric Constant and Low-loss Dielectric Ceramics of Sr5LnTi3Ta7O30 (Ln= La, Nd, Sm and Y)

    Institute of Scientific and Technical Information of China (English)

    TU Wenmao; CHEN Pu; PENG Xiyang; LIU Ting

    2008-01-01

    Polycrystalline Sr5LnTi3Ta7O30 (Ln=La, Nd, Sm and Y) ceramics were prepared as single-phase materials through conventional solid-state ceramics method. The structure was characterized by X-ray diffraction method and scanning electron microscopy (SEM). The dielectric properties were measured from room temperature to 400 ℃. All compounds are paraelectric phases adopting the filled tetragonal tungsten bronze (TB) structure at room temperature. At 1 MHz their dielectric constant (e) varied from 109 to 139,dielectric loss changed from 0.003 3 to 0.005 8, and the temperature coefficients of the dielectric constant (τε)moved from -710 to -880×10-6 ℃-1.

  9. Development of Pasteuria penetrans in Meloidogyne javanica females as affected by constantly high vs fluctuating temperature in an in-vivo system

    Institute of Scientific and Technical Information of China (English)

    DARBAN D.A.; GOWEN S.R.; PEMBROKE B.; MAHAR A.N.

    2005-01-01

    Growth room and glasshouse experiment was conducted to investigate the effect of constant and fluctuating temperatures on the development of Pasteuria penetrans a hyperparasite of root-knot nematodes. Tomato plants (Lycopersicon esculentum Mill) were inoculated with Meloidogynejavanica second-stage juveniles attached with endospores ofP. penetrans and were grown in growth room at 26-29 ℃ and in glasshouse at 20-32 ℃. The tomato plants were sampled from the growth room after 600 degree-days based on 17 ℃/d, accumulating each day above a base temperature of 10 ℃ and from the glasshouse after 36 calendar days. Temperature affected the development ofP. penetrans directly. The rate of development at constant temperature in growth room was faster than that in the glasshouse at fluctuating temperatures.

  10. Non-Relativistic Spacetimes with Cosmological Constant

    OpenAIRE

    Aldrovandi, R.; Barbosa, A. L.; Crispino, L.C.B.; Pereira, J. G.

    1998-01-01

    Recent data on supernovae favor high values of the cosmological constant. Spacetimes with a cosmological constant have non-relativistic kinematics quite different from Galilean kinematics. De Sitter spacetimes, vacuum solutions of Einstein's equations with a cosmological constant, reduce in the non-relativistic limit to Newton-Hooke spacetimes, which are non-metric homogeneous spacetimes with non-vanishing curvature. The whole non-relativistic kinematics would then be modified, with possible ...

  11. Variation of fundamental constants

    CERN Document Server

    Flambaum, V V

    2006-01-01

    We present a review of recent works devoted to the variation of the fine structure constant alpha, strong interaction and fundamental masses. There are some hints for the variation in quasar absorption spectra, Big Bang nucleosynthesis, and Oklo natural nuclear reactor data. A very promising method to search for the variation of the fundamental constants consists in comparison of different atomic clocks. Huge enhancement of the variation effects happens in transition between accidentally degenerate atomic and molecular energy levels. A new idea is to build a ``nuclear'' clock based on the ultraviolet transition between very low excited state and ground state in Thorium nucleus. This may allow to improve sensitivity to the variation up to 10 orders of magnitude! Huge enhancement of the variation effects is also possible in cold atomic and molecular collisions near Feschbach resonance.

  12. String Scale Cosmological Constant

    OpenAIRE

    Chalmers, Gordon

    2006-01-01

    The cosmological constant is an unexplained until now phenomena of nature that requires an explanation through string effects. The apparent discrepancy between theory and experiment is enourmous and has already been explained several times by the author including mechanisms. In this work the string theory theory of abolished string modes is documented and given perturbatively to all loop orders. The holographic underpinning is also exposed. The matching with the data of the LIGO and D0 experi...

  13. The Hubble Constant.

    Science.gov (United States)

    Jackson, Neal

    2015-01-01

    I review the current state of determinations of the Hubble constant, which gives the length scale of the Universe by relating the expansion velocity of objects to their distance. There are two broad categories of measurements. The first uses individual astrophysical objects which have some property that allows their intrinsic luminosity or size to be determined, or allows the determination of their distance by geometric means. The second category comprises the use of all-sky cosmic microwave background, or correlations between large samples of galaxies, to determine information about the geometry of the Universe and hence the Hubble constant, typically in a combination with other cosmological parameters. Many, but not all, object-based measurements give H0 values of around 72-74 km s(-1) Mpc(-1), with typical errors of 2-3 km s(-1) Mpc(-1). This is in mild discrepancy with CMB-based measurements, in particular those from the Planck satellite, which give values of 67-68 km s(-1) Mpc(-1) and typical errors of 1-2 km s(-1) Mpc(-1). The size of the remaining systematics indicate that accuracy rather than precision is the remaining problem in a good determination of the Hubble constant. Whether a discrepancy exists, and whether new physics is needed to resolve it, depends on details of the systematics of the object-based methods, and also on the assumptions about other cosmological parameters and which datasets are combined in the case of the all-sky methods.

  14. Universe of constant

    Science.gov (United States)

    Yongquan, Han

    2016-10-01

    The ideal gas state equation is not applicable to ordinary gas, it should be applied to the Electromagnetic ``gas'' that is applied to the radiation, the radiation should be the ultimate state of matter changes or initial state, the universe is filled with radiation. That is, the ideal gas equation of state is suitable for the Singular point and the universe. Maybe someone consider that, there is no vessel can accommodate radiation, it is because the Ordinary container is too small to accommodate, if the radius of your container is the distance that Light through an hour, would you still think it can't accommodates radiation? Modern scientific determinate that the radius of the universe now is about 1027 m, assuming that the universe is a sphere whose volume is approximately: V = 4.19 × 1081 cubic meters, the temperature radiation of the universe (cosmic microwave background radiation temperature of the universe, should be the closest the average temperature of the universe) T = 3.15k, radiation pressure P = 5 × 10-6 N / m 2, according to the law of ideal gas state equation, PV / T = constant = 6 × 1075, the value of this constant is the universe, The singular point should also equal to the constant Author: hanyongquan

  15. Structural and Dynamic Basis for Low-Affinity, High-Selectivity Binding of L-Glutamine by the Glutamine Riboswitch

    Directory of Open Access Journals (Sweden)

    Aiming Ren

    2015-12-01

    Full Text Available Naturally occurring L-glutamine riboswitches occur in cyanobacteria and marine metagenomes, where they reside upstream of genes involved in nitrogen metabolism. By combining X-ray, NMR, and MD, we characterized an L-glutamine-dependent conformational transition in the Synechococcus elongatus glutamine riboswitch from tuning fork to L-shaped alignment of stem segments. This transition generates an open ligand-binding pocket with L-glutamine selectivity enforced by Mg2+-mediated intermolecular interactions. The transition also stabilizes the P1 helix through a long-range “linchpin” Watson-Crick G-C pair-capping interaction, while melting a short helix below P1 potentially capable of modulating downstream readout. NMR data establish that the ligand-free glutamine riboswitch in Mg2+ solution exists in a slow equilibrium between flexible tuning fork and a minor conformation, similar, but not identical, to the L-shaped bound conformation. We propose that an open ligand-binding pocket combined with a high conformational penalty for forming the ligand-bound state provide mechanisms for reducing binding affinity while retaining high selectivity.

  16. Structural and Dynamic Basis for Low-Affinity, High-Selectivity Binding of L-Glutamine by the Glutamine Riboswitch.

    Science.gov (United States)

    Ren, Aiming; Xue, Yi; Peselis, Alla; Serganov, Alexander; Al-Hashimi, Hashim M; Patel, Dinshaw J

    2015-12-01

    Naturally occurring L-glutamine riboswitches occur in cyanobacteria and marine metagenomes, where they reside upstream of genes involved in nitrogen metabolism. By combining X-ray, NMR, and MD, we characterized an L-glutamine-dependent conformational transition in the Synechococcus elongatus glutamine riboswitch from tuning fork to L-shaped alignment of stem segments. This transition generates an open ligand-binding pocket with L-glutamine selectivity enforced by Mg(2+)-mediated intermolecular interactions. The transition also stabilizes the P1 helix through a long-range "linchpin" Watson-Crick G-C pair-capping interaction, while melting a short helix below P1 potentially capable of modulating downstream readout. NMR data establish that the ligand-free glutamine riboswitch in Mg(2+) solution exists in a slow equilibrium between flexible tuning fork and a minor conformation, similar, but not identical, to the L-shaped bound conformation. We propose that an open ligand-binding pocket combined with a high conformational penalty for forming the ligand-bound state provide mechanisms for reducing binding affinity while retaining high selectivity.

  17. The Mycobacterium tuberculosis high-affinity iron importer, IrtA, contains an FAD-binding domain.

    Science.gov (United States)

    Ryndak, Michelle B; Wang, Shuishu; Smith, Issar; Rodriguez, G Marcela

    2010-02-01

    Iron is an essential nutrient not freely available to microorganisms infecting mammals. To overcome iron deficiency, bacteria have evolved various strategies including the synthesis and secretion of high-affinity iron chelators known as siderophores. The siderophores produced and secreted by Mycobacterium tuberculosis, exomycobactins, compete for iron with host iron-binding proteins and, together with the iron-regulated ABC transporter IrtAB, are required for the survival of M. tuberculosis in iron deficient conditions and for normal replication in macrophages and in mice. This study further characterizes the role of IrtAB in M. tuberculosis iron acquisition. Our results demonstrate a role for IrtAB in iron import and show that the amino terminus domain of IrtA is a flavin-adenine dinucleotide-binding domain essential for iron acquisition. These results suggest a model in which the amino terminus of IrtA functions to couple iron transport and assimilation.

  18. The occurrence and production of avidin: a new conception of the high-affinity biotin-binding protein.

    Science.gov (United States)

    Elo, H A; Korpela, J

    1984-01-01

    The production of avidin, a high-affinity biotin-binding egg-white protein, is not restricted to the avian, amphibian and reptilian oviducts. In the acute phase of inflammation, avidin is synthesized and secreted by various injured tissues in the domestic fowl, both male and female. Also in other avian species and a lizard, injured tissues produce an avidin-like biotin-binding factor. The non-oviductal production of avidin in domestic fowl has a great variety of inducers, for example acute inflammation caused by mechanical or thermal tissue injury, septic bacterial infection and (toxic) drugs, and even retrovirus-induced cell transformation. In culture, chicken embryo fibroblasts and yolk sac macrophages synthesize and secrete avidin. Besides the albumen, avidin may act as an antibacterial protein also in the tissues.

  19. Assessing high affinity binding to HLA-DQ2.5 by a novel peptide library based approach

    DEFF Research Database (Denmark)

    Jüse, Ulrike; Arntzen, Magnus; Højrup, Peter

    2011-01-01

    Here we report on a novel peptide library based method for HLA class II binding motif identification. The approach is based on water soluble HLA class II molecules and soluble dedicated peptide libraries. A high number of different synthetic peptides are competing to interact with a limited amount...... to HLA are then isolated by size exclusion chromatography and sequenced by tandem mass spectrometry online coupled to liquid chromatography. The MS/MS data are subsequently searched against a library defined database using a search engine such as Mascot, followed by manual inspection of the results. We...... used two dodecamer and two decamer peptide libraries and HLA-DQ2.5 to test possibilities and limits of this method. The selected sequences which we identified in the fraction eluted from HLA-DQ2.5 showed a higher average of their predicted binding affinity values compared to the original peptide...

  20. High-throughput identification of telomere-binding ligands based on the fluorescence regulation of DNA-copper nanoparticles.

    Science.gov (United States)

    Yang, Luzhu; Wang, Yanjun; Li, Baoxin; Jin, Yan

    2017-01-15

    Formation of the G-quadruplex in the human telomeric DNA is an effective way to inhibit telomerase activity. Therefore, screening ligands of G-quadruplex has potential applications in the treatment of cancer by inhibit telomerase activity. Although several techniques have been explored for screening of telomeric G-quadruplexes ligands, high-throughput screening method for fast screening telomere-binding ligands from the large compound library is still urgently needed. Herein, a label-free fluorescence strategy has been proposed for high-throughput screening telomere-binding ligands by using DNA-copper nanoparticles (DNA-CuNPs) as a signal probe. In the absence of ligands, human telomeric DNA (GDNA) hybridized with its complementary DNA (cDNA) to form double stranded DNA (dsDNA) which can act as an efficient template for the formation of DNA-CuNPs, leading to the high fluorescence of DNA-CuNPs. In the presence of ligands, GDNA folded into G-quadruplex. Single-strdanded cDNA does not support the formation of DNA-CuNP, resulting in low fluorescence of DNA-CuNPs. Therefore, telomere-binding ligands can be high-throughput screened by monitoring the change in the fluorescence of DNA-CuNPs. Thirteen traditional chinese medicines were screened. Circular dichroism (CD) measurements demonstrated that the selected ligands could induce single-stranded telomeric DNA to form G-quadruplex. The telomere repeat amplification protocol (TRAP) assay demonstrated that the selected ligands can effectively inhibit telomerase activity. Therefore, it offers a cost-effective, label-free and reliable high-throughput way to identify G-quadruplex ligands, which holds great potential in discovering telomerase-targeted anticancer drugs. Copyright © 2016 Elsevier B.V. All rights reserved.

  1. High levels of mannose-binding lectin are associated with lower pulse wave velocity in uraemic patients

    DEFF Research Database (Denmark)

    Hornum, Mads; Bay, Jakob T; Clausen, Peter

    2014-01-01

    -concentration below or above the median concentration. These groups were comparable regarding age, BMI, and duration of ESRD. PWV was significantly lower in the group with high MBL levels compared to the group with low MBL levels and trends toward better AIX and higher insulin sensitivity (ISI) was also seen......BACKGROUND: Uraemia is associated with a highly increased risk of cardiovascular disease. Mannose-binding lectin (MBL) has been shown to be involved in cardiovascular pathophysiology and a protective effect of MBL is suggested. The purpose of the present study was to evaluate a potential impact...... in the group with high MBL levels. No difference was seen in FMD and NID. CONCLUSIONS: High levels of MBL are associated with lower PWV and the use of antihypertensive drugs in a cohort of patients with ESRD awaiting kidney transplantation suggesting a beneficial role of high levels of MBL on arterial...

  2. Cryptography in constant parallel time

    CERN Document Server

    Applebaum, Benny

    2013-01-01

    Locally computable (NC0) functions are 'simple' functions for which every bit of the output can be computed by reading a small number of bits of their input. The study of locally computable cryptography attempts to construct cryptographic functions that achieve this strong notion of simplicity and simultaneously provide a high level of security. Such constructions are highly parallelizable and they can be realized by Boolean circuits of constant depth.This book establishes, for the first time, the possibility of local implementations for many basic cryptographic primitives such as one-way func

  3. Constant-bandwidth constant-temperature hot-wire anemometer.

    Science.gov (United States)

    Ligeza, P

    2007-07-01

    A constant-temperature anemometer (CTA) enables the measurement of fast-changing velocity fluctuations. In the classical solution of CTA, the transmission band is a function of flow velocity. This is a minor drawback when the mean flow velocity does not significantly change, though it might lead to dynamic errors when flow velocity varies over a considerable range. A modification is outlined, whereby an adaptive controller is incorporated in the CTA system such that the anemometer's transmission band remains constant in the function of flow velocity. For that purpose, a second feedback loop is provided, and the output signal from the anemometer will regulate the controller's parameters such that the transmission bandwidth remains constant. The mathematical model of a CTA that has been developed and model testing data allow a through evaluation of the proposed solution. A modified anemometer can be used in measurements of high-frequency variable flows in a wide range of velocities. The proposed modification allows the minimization of dynamic measurement errors.

  4. Influence of sulfhydryl sites on metal binding by bacteria

    Science.gov (United States)

    Nell, Ryan M.; Fein, Jeremy B.

    2017-02-01

    The role of sulfhydryl sites within bacterial cell envelopes is still unknown, but the sites may control the fate and bioavailability of metals. Organic sulfhydryl compounds are important complexing ligands in aqueous systems and they can influence metal speciation in natural waters. Though representing only approximately 5-10% of the total available binding sites on bacterial surfaces, sulfhydryl sites exhibit high binding affinities for some metals. Due to the potential importance of bacterial sulfhydryl sites in natural systems, metal-bacterial sulfhydryl site binding constants must be determined in order to construct accurate models of the fate and distribution of metals in these systems. To date, only Cd-sulfhydryl binding has been quantified. In this study, the thermodynamic stabilities of Mn-, Co-, Ni-, Zn-, Sr- and Pb-sulfhydryl bacterial cell envelope complexes were determined for the bacterial species Shewanella oneidensis MR-1. Metal adsorption experiments were conducted as a function of both pH, ranging from 5.0 to 7.0, and metal loading, from 0.5 to 40.0 μmol/g (wet weight) bacteria, in batch experiments in order to determine if metal-sulfhydryl binding occurs. Initially, the data were used to calculate the value of the stability constants for the important metal-sulfhydryl bacterial complexes for each metal-loading condition studied, assuming a single binding reaction for the dominant metal-binding site type under the pH conditions of the experiments. For most of the metals that we studied, these calculated stability constant values increased significantly with decreasing metal loading, strongly suggesting that our initial assumption was not valid and that more than one type of binding occurs at the assumed binding site. We then modeled each dataset with two distinct site types with identical acidity constants: one site with a high metal-site stability constant value, which we take to represent metal-sulfhydryl binding and which dominates under low

  5. Comparative studies of vertebrate scavenger receptor class B type 1: a high-density lipoprotein binding protein

    Directory of Open Access Journals (Sweden)

    Holmes RS

    2012-06-01

    Full Text Available Roger S Holmes,1,2 Laura A Cox11Department of Genetics and Southwest National Primate Research Center, Texas Biomedical Research Institute, San Antonio, TX, USA; 2School of Biomolecular and Physical Sciences, Griffith University, Nathan, Queensland, AustraliaAbstract: Scavenger receptor class B type 1 protein (SCARB1 plays an essential role in cholesterol homeostasis and functions in binding high density lipoprotein cholesterol (HDL in liver and other tissues of the body. SCARB1 also functions in lymphocyte homeostasis and in the uptake of hepatitis C virus (HCV by the liver. A genetic deficiency of this protein results in autoimmune disorders and significant changes in blood cholesterol phenotype. Comparative SCARB1 amino acid sequences and structures and SCARB1 gene locations were examined using data from several vertebrate genome projects. Vertebrate SCARB1 sequences shared 50%–99% identity as compared with 28%–31% sequence identities with other CD36-like superfamily members, ie, SCARB2 and SCARB3 (also called CD36. At least eight N-glycosylation sites were conserved among most of the vertebrate SCARB1 proteins examined. Sequence alignments, key amino acid residues, and conserved predicted secondary structures were also studied, including: cytoplasmic, transmembrane, and exoplasmic sequences; conserved N-terminal and C-terminal transmembrane glycines which participate in oligomer formation; conserved cystine disulfides and a free SH residue which participates in lipid transport; carboxyl terminal PDZ-binding domain sequences (Ala507-Arg/Lys508-Leu509; and 30 conserved proline and 18 conserved glycine residues, which may contribute to short loop formation within the exoplasmic HDL-binding sequence. Vertebrate SCARB1 genes usually contained 12 coding exons. The human SCARB1 gene contained CpG islands, micro RNA binding sites, and several transcription factor binding sites (including PPARG which may contribute to the high level (13.7 times

  6. The Hubble Constant

    Directory of Open Access Journals (Sweden)

    Neal Jackson

    2015-09-01

    Full Text Available I review the current state of determinations of the Hubble constant, which gives the length scale of the Universe by relating the expansion velocity of objects to their distance. There are two broad categories of measurements. The first uses individual astrophysical objects which have some property that allows their intrinsic luminosity or size to be determined, or allows the determination of their distance by geometric means. The second category comprises the use of all-sky cosmic microwave background, or correlations between large samples of galaxies, to determine information about the geometry of the Universe and hence the Hubble constant, typically in a combination with other cosmological parameters. Many, but not all, object-based measurements give H_0 values of around 72–74 km s^–1 Mpc^–1, with typical errors of 2–3 km s^–1 Mpc^–1. This is in mild discrepancy with CMB-based measurements, in particular those from the Planck satellite, which give values of 67–68 km s^–1 Mpc^–1 and typical errors of 1–2 km s^–1 Mpc^–1. The size of the remaining systematics indicate that accuracy rather than precision is the remaining problem in a good determination of the Hubble constant. Whether a discrepancy exists, and whether new physics is needed to resolve it, depends on details of the systematics of the object-based methods, and also on the assumptions about other cosmological parameters and which datasets are combined in the case of the all-sky methods.

  7. Soluble T cell receptor Vβ domains engineered for high-affinity binding to staphylococcal or streptococcal superantigens.

    Science.gov (United States)

    Sharma, Preeti; Wang, Ningyan; Kranz, David M

    2014-01-28

    Staphylococcus aureus and group A Streptococcus secrete a collection of toxins called superantigens (SAgs), so-called because they stimulate a large fraction of an individual's T cells. One consequence of this hyperactivity is massive cytokine release leading to severe tissue inflammation and, in some cases, systemic organ failure and death. The molecular basis of action involves the binding of the SAg to both a T cell receptor (TCR) on a T cell and a class II product of the major histocompatibility complex (MHC) on an antigen presenting cell. This cross-linking leads to aggregation of the TCR complex and signaling. A common feature of SAgs is that they bind with relatively low affinity to the variable region (V) of the beta chain of the TCR. Despite this low affinity binding, SAgs are very potent, as each T cell requires only a small fraction of their receptors to be bound in order to trigger cytokine release. To develop high-affinity agents that could neutralize the activity of SAgs, and facilitate the development of detection assays, soluble forms of the Vβ regions have been engineered to affinities that are up to 3 million-fold higher for the SAg. Over the past decade, six different Vβ regions against SAgs from S. aureus (SEA, SEB, SEC3, TSST-1) or S. pyogenes (SpeA and SpeC) have been engineered for high-affinity using yeast display and directed evolution. Here we review the engineering of these high-affinity Vβ proteins, structural features of the six different SAgs and the Vβ proteins, and the specific properties of the engineered Vβ regions that confer high-affinity and specificity for their SAg ligands.

  8. Soluble T Cell Receptor Vβ Domains Engineered for High-Affinity Binding to Staphylococcal or Streptococcal Superantigens

    Directory of Open Access Journals (Sweden)

    Preeti Sharma

    2014-01-01

    Full Text Available Staphylococcus aureus and group A Streptococcus secrete a collection of toxins called superantigens (SAgs, so-called because they stimulate a large fraction of an individual’s T cells. One consequence of this hyperactivity is massive cytokine release leading to severe tissue inflammation and, in some cases, systemic organ failure and death. The molecular basis of action involves the binding of the SAg to both a T cell receptor (TCR on a T cell and a class II product of the major histocompatibility complex (MHC on an antigen presenting cell. This cross-linking leads to aggregation of the TCR complex and signaling. A common feature of SAgs is that they bind with relatively low affinity to the variable region (V of the beta chain of the TCR. Despite this low affinity binding, SAgs are very potent, as each T cell requires only a small fraction of their receptors to be bound in order to trigger cytokine release. To develop high-affinity agents that could neutralize the activity of SAgs, and facilitate the development of detection assays, soluble forms of the Vβ regions have been engineered to affinities that are up to 3 million-fold higher for the SAg. Over the past decade, six different Vβ regions against SAgs from S. aureus (SEA, SEB, SEC3, TSST-1 or S. pyogenes (SpeA and SpeC have been engineered for high-affinity using yeast display and directed evolution. Here we review the engineering of these high-affinity Vβ proteins, structural features of the six different SAgs and the Vβ proteins, and the specific properties of the engineered Vβ regions that confer high-affinity and specificity for their SAg ligands.

  9. Phosphate-binding pocket in Dicer-2 PAZ domain for high-fidelity siRNA production.

    Science.gov (United States)

    Kandasamy, Suresh K; Fukunaga, Ryuya

    2016-12-06

    The enzyme Dicer produces small silencing RNAs such as micro-RNAs (miRNAs) and small interfering RNAs (siRNAs). In Drosophila, Dicer-1 produces ∼22-24-nt miRNAs from pre-miRNAs, whereas Dicer-2 makes 21-nt siRNAs from long double-stranded RNAs (dsRNAs). How Dicer-2 precisely makes 21-nt siRNAs with a remarkably high fidelity is unknown. Here we report that recognition of the 5'-monophosphate of a long dsRNA substrate by a phosphate-binding pocket in the Dicer-2 PAZ (Piwi, Argonaute, and Zwille/Pinhead) domain is crucial for the length fidelity, but not the efficiency, in 21-nt siRNA production. Loss of the length fidelity, meaning increased length heterogeneity of siRNAs, caused by point mutations in the phosphate-binding pocket of the Dicer-2 PAZ domain decreased RNA silencing activity in vivo, showing the importance of the high fidelity to make 21-nt siRNAs. We propose that the 5'-monophosphate of a long dsRNA substrate is anchored by the phosphate-binding pocket in the Dicer-2 PAZ domain and the distance between the pocket and the RNA cleavage active site in the RNaseIII domain corresponds to the 21-nt pitch in the A-form duplex of a long dsRNA substrate, resulting in high-fidelity 21-nt siRNA production. This study sheds light on the molecular mechanism by which Dicer-2 produces 21-nt siRNAs with a remarkably high fidelity for efficient RNA silencing.

  10. Replacement of the Bryostatin A- and B-Pyran Rings With Phenyl Rings Leads to Loss of High Affinity Binding With PKC.

    Science.gov (United States)

    Petersen, Mark E; Kedei, Noemi; Lewin, Nancy E; Blumberg, Peter M; Keck, Gary E

    2016-10-19

    We describe a convergent synthesis of a bryostatin analogue in which the natural A- and B-ring pyrans have been replaced by phenyl rings. The new analogue exhibited PMA like behavior in cell assays, but failed to maintain high affinity binding for PKC, despite retaining an unaltered C-ring 'binding domain'.

  11. Isolation and partial characterization of gypsy moth BTR-270, an anionic brush border membrane glycoconjugate that binds Bacillus thuringiensis Cry1A toxins with high affinity

    Science.gov (United States)

    Algimantas P. Valaitis; Jeremy L. Jenkins; Mi Kyong Lee; Donald H. Dean; Karen J. Garner

    2001-01-01

    BTR-270, a gypsy moth (Lymantria dispar) brush border membrane molecule that binds Bacillus thuringiensis (Bt) Cry1A toxins with high affinity, was purified by preparative gel electrophoresis. Rabbit antibodies specific for the Bt toxin-binding molecule were raised. Attempts to label BTR-270 by protein-directed techniques were...

  12. A bambusuril macrocycle that binds anions in water with high affinity and selectivity.

    Science.gov (United States)

    Yawer, Mirza Arfan; Havel, Vaclav; Sindelar, Vladimir

    2015-01-02

    Synthetic receptors that function in water are important for the qualitative and quantitative detection of anions, which may act as pollutants in the environment or play important roles in biological processes. Neutral receptors are particularly appealing because they are often more selective than positively charged receptors; however, their affinity towards anions in pure water is only in range of 1-10(3)  L mol(-1) . The anion-templated synthesis of a water-soluble bambusuril derivative is shown to be an outstanding receptor for various inorganic anions in pure water, with association constants of up to 10(7)  L mol(-1) . Furthermore, the macrocycle discriminates between anions with unprecedented selectivity (up to 500 000-fold). We anticipate that the combination of remarkable affinity and selectivity of this macrocycle will enable the efficient detection and isolation of diverse anions in aqueous solutions, which is not possible with current supramolecular systems.

  13. Fragile X mental retardation protein recognition of G quadruplex structure per se is sufficient for high affinity binding to RNA.

    Science.gov (United States)

    Bole, Medhavi; Menon, Lakshmi; Mihailescu, Mihaela-Rita

    2008-12-01

    Fragile X syndrome, the most common form of inherited mental retardation is caused by the expansion of a CGG trinucleotide repeat in the fragile X mental retardation 1 (fmr1) gene. The abnormal expansion of the CGG repeat causes hypermethylation and subsequent silencing of the fmr1 gene, resulting in the loss of the fragile X mental retardation protein (FMRP). FMRP has been shown to use its arginine-glycine-glycine rich region (RGG box) to bind to messenger RNAs that form G quadruplex structures. Several studies reported that the G quadruplex RNA recognition alone is not sufficient for FMRP RGG box binding and that an additional stem and/or a G quadruplex-stem junction region may also be important in recognition. In this study we have used biophysical methods such as fluorescence, UV, CD and NMR spectroscopy to demonstrate that the recognition of the RNA G quadruplex structure per se, in the absence of a stem region, is sufficient for the FMRP high affinity and specific binding. These findings indicate that the presence of a stem structure in some of the FMRP G quadruplex forming mRNAs is not a requirement for protein recognition as previously believed, but rather for the proper formation of the correct RNA G quadruplex structure recognized by FMRP.

  14. High trait aggression in men is associated with low 5-HT levels, as indexed by 5-HT4 receptor binding.

    Science.gov (United States)

    da Cunha-Bang, Sofi; Mc Mahon, Brenda; Fisher, Patrick MacDonald; Jensen, Peter Steen; Svarer, Claus; Knudsen, Gitte Moos

    2016-04-01

    Impulsive aggression has commonly been associated with a dysfunction of the serotonin (5-HT) system: many, but not all, studies point to an inverse relationship between 5-HT and aggression. As cerebral 5-HT4 receptor (5-HT4R) binding has recently been recognized as a proxy for stable brain levels of 5-HT, we here test the hypothesis in healthy men and women that brain 5-HT levels, as indexed by cerebral 5-HT4R, are inversely correlated with trait aggression and impulsivity. Sixty-one individuals (47 men) underwent positron emission tomography scanning with the radioligand [(11)C]SB207145 for quantification of brain 5-HT4R binding. The Buss-Perry Aggression Questionnaire (BPAQ) and the Barratt Impulsiveness Scale were used for assessment of trait aggression and trait impulsivity. Among male subjects, there was a positive correlation between global 5-HT4R and BPAQ total score (P = 0.037) as well as BPAQ physical aggression (P = 0.025). No main effect of global 5-HT4R on trait aggression or impulsivity was found in the mixed gender sample, but there was evidence for sex interaction effects in the relationship between global 5-HT4R and BPAQ physical aggression. In conclusion we found that low cerebral 5-HT levels, as indexed by 5-HT4R binding were associated with high trait aggression in males, but not in females.

  15. High affinity RGD-binding sites at the plasma membrane of Arabidopsis thaliana links the cell wall.

    Science.gov (United States)

    Canut, H; Carrasco, A; Galaud, J P; Cassan, C; Bouyssou, H; Vita, N; Ferrara, P; Pont-Lezica, R

    1998-10-01

    The heptapeptide Tyr-Gly-Arg-Gly-Asp-Ser-Pro containing the sequence Arg-Gly-Asp (RGD--the essential structure recognised by animal cells in substrate adhesion molecules) was tested on epidermal cells of onion and cultured cells of Arabidopsis upon plasmolysis. Dramatic changes were observed on both types of cells following treatment: on onion cells, Hechtian strands linking the cell wall to the membrane were lost, while Arabidopsis cells changed from concave to convex plasmolysis. A control heptapeptide Tyr-Gly-Asp-Gly-Arg-Ser-Pro had no effect on the shape of plasmolysed cells. Protoplasts isolated from Arabidopsis cells agglutinate in the presence of ProNectinF, a genetically engineered protein of 72 kDa containing 13 RGD sequences: several protoplasts may adhere to a single molecule of ProNectinF. The addition of the RGD-heptapeptide disrupted the adhesion between the protoplasts. Purified plasma membrane from Arabidopsis cells exhibits specific binding sites for the iodinated RGD-heptapeptide. The binding is saturable, reversible, and two types of high affinity sites (Kd1 approximately 1 nM, and Kd2 approximately 40 nM) can be discerned. Competitive inhibition by several structurally related peptides and proteins noted the specific requirement for the RGD sequence. Thus, the RGD-binding activity of Arabidopsis fulfils the adhesion features of integrins, i.e. peptide specificity, subcellular location, and involvement in plasma membrane-cell wall attachments.

  16. High-affinity DNA binding sites for H-NS provide a molecular basis for selective silencing within proteobacterial genomes.

    Science.gov (United States)

    Lang, Benjamin; Blot, Nicolas; Bouffartigues, Emeline; Buckle, Malcolm; Geertz, Marcel; Gualerzi, Claudio O; Mavathur, Ramesh; Muskhelishvili, Georgi; Pon, Cynthia L; Rimsky, Sylvie; Stella, Stefano; Babu, M Madan; Travers, Andrew

    2007-01-01

    The global transcriptional regulator H-NS selectively silences bacterial genes associated with pathogenicity and responses to environmental insults. Although there is ample evidence that H-NS binds preferentially to DNA containing curved regions, we show here that a major basis for this selectivity is the presence of a conserved sequence motif in H-NS target transcriptons. We further show that there is a strong tendency for the H-NS binding sites to be clustered, both within operons and in genes contained in the pathogenicity-associated islands. In accordance with previously published findings, we show that these motifs occur in AT-rich regions of DNA. On the basis of these observations, we propose that H-NS silences extensive regions of the bacterial chromosome by binding first to nucleating high-affinity sites and then spreading along AT-rich DNA. This spreading would be reinforced by the frequent occurrence of the motif in such regions. Our findings suggest that such an organization enables the silencing of extensive regions of the genetic material, thereby providing a coherent framework that unifies studies on the H-NS protein and a concrete molecular basis for the genetic control of H-NS transcriptional silencing.

  17. Glycosylphosphatidylinositol-anchored high-density lipoprotein-binding protein 1 and the intravascular processing of triglyceride-rich lipoproteins.

    Science.gov (United States)

    Adeyo, O; Goulbourne, C N; Bensadoun, A; Beigneux, A P; Fong, L G; Young, S G

    2012-12-01

    Lipoprotein lipase (LPL) is produced by parenchymal cells, mainly adipocytes and myocytes, but is involved in hydrolysing triglycerides in plasma lipoproteins at the capillary lumen. For decades, the mechanism by which LPL reaches its site of action in capillaries was unclear, but this mystery was recently solved. Glycosylphosphatidylinositol-anchored high-density lipoprotein-binding protein 1 (GPIHBP1), a glycosylphosphatidylinositol-anchored protein of capillary endothelial cells, 'picks up' LPL from the interstitial spaces and shuttles it across endothelial cells to the capillary lumen. When GPIHBP1 is absent, LPL is mislocalized to the interstitial spaces, leading to severe hypertriglyceridaemia. Some cases of hypertriglyceridaemia in humans are caused by GPIHBP1 mutations that interfere with the ability of GPIHBP1 to bind to LPL, and some are caused by LPL mutations that impair the ability of LPL to bind to GPIHBP1. Here, we review recent progress in understanding the role of GPIHBP1 in health and disease and discuss some of the remaining unresolved issues regarding the processing of triglyceride-rich lipoproteins. © 2012 The Association for the Publication of the Journal of Internal Medicine.

  18. High cadmium-binding ability of a novel Colocasia esculenta metallothionein increases cadmium tolerance in Escherichia coli and tobacco.

    Science.gov (United States)

    Kim, Yeon-Ok; Patel, Darshan H; Lee, Dae-Seok; Song, Younho; Bae, Hyeun-Jong

    2011-01-01

    Experimental evidence in vivo as to the functional roles and binding properties to cadmium (Cd) of type-2 plants metallothionein (MT) has been limited thus far. We investigated the biological role of metallothionein from Colocasia esculenta (CeMT2b) in Escherichia coli and tobacco, and developed a new model for the relationship between Cd tolerance and Cd-binding ability. Heterologous expression of CeMT2b in Escherichia coli greatly enhanced Cd tolerance and accumulated Cd content as compared to control cells. The molecular weight of CeMT2b increased with Cd, and CeMT2b bound up to 5.96±1 molar ratio (Cd/protein). Under Cd stress, transgenic tobacco plants displayed much better seedling growth and high Cd accumulation than the wild type. The presence of an extra CXC motif in CeMT2b contributed to the enhanced Cd-tolerance. The present study provides the first insight into the ability of type-2 plant MT to bind physiological Cd.

  19. The conserved lymphokine element-0 in the IL5 promoter binds to a high mobility group-1 protein.

    Science.gov (United States)

    Marrugo, J; Marsh, D G; Ghosh, B

    1996-10-01

    The conserved lymphokine elements-0 (CLE0) in the IL5 promoter is essential for the expression of IL-5. Here, we report the cloning and expression of a cDNA encoding a novel CLE0-binding protein, CLEBP-1 from a mouse Th2 clone, D10.G4.1. Interestingly, it was found that the CLEBP1 cDNA sequence was almost identical to the sequences of known high mobility group-1 (HMG1) cDNAs. When expressed as a recombinant fusion protein in Escherichia coli, CLEBP-1 was shown to bind to the IL5-CLE0 element in electrophoretic mobility-shift assays (EMSA) and southwestern blot analysis. The CLEBP-1 fusion protein cross-reacts with and-HMG-1/2 in Western blot analysis. It also binds to the CLE0 elements of IL4, GMCSF and GCSF genes. CLEBP-1 and closely related HMG-1 and HMG-2 proteins may play key roles in facilitating the expression of the lymphokine genes that contain CLE0 elements.

  20. Ice-binding site of snow mold fungus antifreeze protein deviates from structural regularity and high conservation.

    Science.gov (United States)

    Kondo, Hidemasa; Hanada, Yuichi; Sugimoto, Hiroshi; Hoshino, Tamotsu; Garnham, Christopher P; Davies, Peter L; Tsuda, Sakae

    2012-06-12

    Antifreeze proteins (AFPs) are found in organisms ranging from fish to bacteria, where they serve different functions to facilitate survival of their host. AFPs that protect freeze-intolerant fish and insects from internal ice growth bind to ice using a regular array of well-conserved residues/motifs. Less is known about the role of AFPs in freeze-tolerant species, which might be to beneficially alter the structure of ice in or around the host. Here we report the 0.95-Å high-resolution crystal structure of a 223-residue secreted AFP from the snow mold fungus Typhula ishikariensis. Its main structural element is an irregular β-helix with six loops of 18 or more residues that lies alongside an α-helix. β-Helices have independently evolved as AFPs on several occasions and seem ideally structured to bind to several planes of ice, including the basal plane. A novelty of the β-helical fold is the nonsequential arrangement of loops that places the N- and C termini inside the solenoid of β-helical coils. The ice-binding site (IBS), which could not be predicted from sequence or structure, was located by site-directed mutagenesis to the flattest surface of the protein. It is remarkable for its lack of regularity and its poor conservation in homologs from psychrophilic diatoms and bacteria and other fungi.

  1. ZipA binds to FtsZ with high affinity and enhances the stability of FtsZ protofilaments.

    Directory of Open Access Journals (Sweden)

    Anuradha Kuchibhatla

    Full Text Available A bacterial membrane protein ZipA that tethers FtsZ to the membrane is known to promote FtsZ assembly. In this study, the binding of ZipA to FtsZ was monitored using fluorescence spectroscopy. ZipA was found to bind to FtsZ with high affinities at three different (6.0, 6.8 and 8.0 pHs, albeit the binding affinity decreased with increasing pH. Further, thick bundles of FtsZ protofilaments were observed in the presence of ZipA under the pH conditions used in this study indicating that ZipA can promote FtsZ assembly and stabilize FtsZ polymers under unfavorable conditions. Bis-ANS, a hydrophobic probe, decreased the interaction of FtsZ and ZipA indicating that the interaction between FtsZ and ZipA is hydrophobic in nature. ZipA prevented the dilution induced disassembly of FtsZ polymers suggesting that it stabilizes FtsZ protofilaments. Fluorescein isothiocyanate-labeled ZipA was found to be uniformly distributed along the length of the FtsZ protofilaments indicating that ZipA stabilizes FtsZ protofilaments by cross-linking them.

  2. Subtractive Cell-SELEX Selection of DNA Aptamers Binding Specifically and Selectively to Hepatocellular Carcinoma Cells with High Metastatic Potential

    Directory of Open Access Journals (Sweden)

    Hao Chen

    2016-01-01

    Full Text Available Relapse and metastasis are two key risk factors of hepatocellular carcinoma (HCC prognosis; thus, it is emergent to develop an early and accurate detection method for prognostic evaluation of HCC after surgery. In this study, we sought to acquire oligonucleotide DNA aptamers that specifically bind to HCC cells with high metastatic potential. Two HCC cell lines derived from the same genetic background but with different metastatic potential were employed: MHCC97L (low metastatic properties as subtractive targets and HCCLM9 (high metastatic properties as screening targets. To mimic a fluid combining environment, initial DNA aptamers library was firstly labelled with magnetic nanoparticles using biotin-streptavidin system and then applied for aptamers selection. Through 10-round selection with subtractive Cell-SELEX, six aptamers, LY-1, LY-13, LY-46, LY-32, LY-27/45, and LY-7/43, display high affinity to HCCLM9 cells and do not bind to MHCC97L cells, as well as other tumor cell lines, including breast cancer, lung cancer, colon adenocarcinoma, gastric cancer, and cervical cancer, suggesting high specificity for HCCLM9 cells. Thus, the aptamers generated here will provide solid basis for identifying new diagnostic targets to detect HCC metastasis and also may provide valuable clues for developing new targeted therapeutics.

  3. 2-O-[2-(Methylthio)ethyl]-Modified Oligonucleotide: An Analog of 2-O-[2-(Methoxy)ethyl]-Modified Oligonucleotide with Improved Protein Binding Properties and High Binding Affinity to Target RNA

    Energy Technology Data Exchange (ETDEWEB)

    Prakash, T.P.; Manoharan, M.; Fraser, A.S.; Kawasaki, A.M.; Lesnik, E.; Sioufi, N.; Leeds, J.M.; Teplova, M.; Egli, M.

    2010-03-08

    A novel 2'-modification, 2'-O-[2-(methylthio)ethyl] or 2'-O-MTE, has been incorporated into oligonucleotides and evaluated for properties relevant to antisense activity. The results were compared with the previously characterized 2'-O-[2-(methoxy)ethyl] 2'-O-MOE modification. As expected, the 2'-O-MTE modified oligonucleotides exhibited improved binding to human serum albumin compared to the 2'-O-MOE modified oligonucleotides. The 2'-O-MTE oligonucleotides maintained high binding affinity to target RNA. Nuclease digestion of 2'-O-MTE oligonucleotides showed that they have limited resistance to exonuclease degradation. We analyzed the crystal structure of a decamer DNA duplex containing the 2'-O-MTE modifcation. Analysis of the crystal structure provides insight into the improved RNA binding affinity, protein binding affinity and limited resistance of 2'-O-MTE modified oligonucleotides to exonuclease degradation.

  4. High yield purification of nanobodies from the periplasm of E. coli as fusions with the maltose binding protein.

    Science.gov (United States)

    Salema, Valencio; Fernández, Luis Ángel

    2013-09-01

    Nanobodies (Nbs) are single domain antibodies based on the variable domains of heavy chain only antibodies (HCAbs) found in camelids, also referred to as VHHs. Their small size (ca. 12-15kDa), superior biophysical and antigen binding properties have made Nbs very attractive molecules for multiple biotechnological applications, including human therapy. The most widely used system for the purification of Nbs is their expression in the periplasm of Escherichia coli with a C-terminal hexa-histidine (His6) tag followed by immobilized metal affinity chromatography (IMAC). However, significant variability in the expression levels of different Nbs are routinely observed and a single affinity chromatography step is often not sufficient to obtain Nbs of high purity. Here, we report an alternative method for expression and purification of Nbs from the periplasm of E. coli based on their fusion to maltose binding protein (MBP) in the N-terminus and His6 tag in the C-terminus (MBP-NbHis6). Soluble MBP-NbHis6 fusions were consistently expressed at high levels (⩾12mg/L of induced culture in shake flasks) in the periplasm of E. coli HM140, a strain deficient in several periplasmic proteases. Highly pure MBP-NbHis6 fusions and free NbHis6 (after site specific proteolysis of the fusions), were recovered by amylose and metal affinity chromatography steps. The monomeric nature of the purified NbHis6 was determined by gel filtration chromatography. Lastly, we demonstrated by ELISA that both monomeric NbHis6 and MBP-NbHis6 fusions retained antigen binding activity and specificity, thus facilitating their direct use in antigen recognition assays.

  5. Inhibition of Enterococcus faecium adherence to collagen by antibodies against high-affinity binding subdomains of Acm.

    Science.gov (United States)

    Nallapareddy, Sreedhar R; Sillanpää, Jouko; Ganesh, Vannakambadi K; Höök, Magnus; Murray, Barbara E

    2007-06-01

    Strains of Enterococcus faecium express a cell wall-anchored protein, Acm, which mediates adherence to collagen. Here, we (i) identify the minimal and high-affinity binding subsegments of Acm and (ii) show that anti-Acm immunoglobulin Gs (IgGs) purified against these subsegments reduced E. faecium TX2535 strain collagen adherence up to 73 and 50%, respectively, significantly more than the total IgGs against the full-length Acm A domain (28%) (P Acm adherence with functional subsegment-specific antibodies raises the possibility of their use as therapeutic or prophylactic agents.

  6. A dualistic conformational response to substrate binding in the human serotonin transporter reveals a high affinity state for serotonin

    DEFF Research Database (Denmark)

    Bjerregaard, Henriette; Severinsen, Kasper; Said, Saida

    2015-01-01

    Serotonergic neurotransmission is modulated by the membrane-embedded serotonin transporter (SERT). SERT mediates the reuptake of serotonin into the presynaptic neurons. Conformational changes in SERT occur upon binding of ions and substrate and are crucial for translocation of serotonin across...... that were sensitized to detect a more outward-facing conformation of SERT. We found a novel high affinity outward-facing conformational state of the human SERT induced by serotonin. The ionic requirements for this new conformational response to serotonin mirror the ionic requirements for translocation...

  7. Magnetic core test stand for energy loss and permeability measurements at a high constant magnetization rate and test results for nanocrystalline and ferrite materials.

    Science.gov (United States)

    Burdt, Russell; Curry, Randy D

    2008-09-01

    A test stand was developed to measure the energy losses and unsaturated permeability of toroidal magnetic cores, relevant to applications of magnetic switching requiring a constant magnetization rate of the order of 1-10 T/micros. These applications in pulsed power include linear induction accelerators, pulse transformers, and discharge switches. The test stand consists of a coaxial transmission line pulse charged up to 100 kV that is discharged into a magnetic core load. Suitable diagnostics measure the voltage across and the current through a winding on the magnetic core load, from which the energy losses and unsaturated permeability are calculated. The development of the test stand is discussed, and test results for ferrite CN20 and the nanocrystalline material Finemet FT-1HS are compared to demonstrate the unique properties of a nanocrystalline material. The experimental data are compared with published data in a similar parameter space to demonstrate the efficacy of the experimental methods.

  8. New Quasar Studies Keep Fundamental Physical Constant Constant

    Science.gov (United States)

    2004-03-01

    atoms together and the way light interacts with atoms. But are these fundamental physical constants really constant? Are those numbers always the same, everywhere in the Universe and at all times? This is not as naive a question as it may seem. Contemporary theories of fundamental interactions, such as the Grand Unification Theory or super-string theories that treat gravity and quantum mechanics in a consistent way, not only predict a dependence of fundamental physical constants with energy - particle physics experiments have shown the fine structure constant to grow to a value of about 1/128 at high collision energies - but allow for their cosmological time and space variations. A time dependence of the fundamental constants could also easily arise if, besides the three space dimensions, there exist more hidden dimensions. Already in 1955, the Russian physicist Lev Landau considered the possibility of a time dependence of alpha. In the late 1960s, George Gamow in the United States suggested that the charge of the electron, and therefore also alpha, may vary. It is clear however that such changes, if any, cannot be large or they would already have been detected in comparatively simple experiments. Tracking these possible changes thus requires the most sophisticated and precise techniques. Looking back in time In fact, quite strong constraints are already known to exist for the possible variation of the fine structure constant alpha. One such constraint is of geological nature. It is based on measures taken in the ancient natural fission reactor located near Oklo (Gabon, West Africa) and which was active roughly 2,000 million years ago. By studying the distribution of a given set of elements - isotopes of the rare earths, for example of samarium - which were produced by the fission of uranium, one can estimate whether the physical process happened at a faster or slower pace than we would expect it nowadays. Thus we can measure a possible change of the value of the

  9. High Throughput Screening of High-Affinity Ligands for Proteins with Anion-Binding Sites using Desorption Electrospray Ionization (DESI) Mass Spectrometry

    Science.gov (United States)

    Lu, Xin; Ning, Baoming; He, Dacheng; Huang, Lingyun; Yue, Xiangjun; Zhang, Qiming; Huang, Haiwei; Liu, Yang; He, Lan; Ouyang, Jin

    2014-03-01

    A high throughput screening system involving a linear ion trap (LTQ) analyzer, a house-made platform and a desorption electrospray ionization (DESI) source was established to screen ligands with a high affinity for proteins with anion-binding sites. The complexes were analyzed after incubation, ultrafiltration, washing, and displacement. A new anionic region inhibited dissociation (ARID) mechanism that was suitable for a protein with anion-binding site was proposed. We utilized the differences in detectable dissociation of protein-ligand complexes, combined with displacement experiments, to distinguish free ligands displaced from anion-binding sites from liberated ligands dissociated from nonspecific interactions. The method was validated by α1-acid glycoprotein (AGP) and (R), (S)-amlodipine. Site-specific enantioselectivity shown in our experiments was consistent with earlier studies. Obtaining all of the qualitative information of 15*3 samples in 2.3 min indicates that the analysis process is no longer the time-limiting step in the initial stage of drug discovery. Quantitative information verified that our method was at least a semiquantitative method.

  10. The Hubble Constant

    Directory of Open Access Journals (Sweden)

    Jackson Neal

    2007-09-01

    Full Text Available I review the current state of determinations of the Hubble constant, which gives the length scale of the Universe by relating the expansion velocity of objects to their distance. In the last 20 years, much progress has been made and estimates now range between 60 and 75 km s^-1 Mpc^-1, with most now between 70 and 75 km s^-1 Mpc^-1, a huge improvement over the factor-of-2 uncertainty which used to prevail. Further improvements which gave a generally agreed margin of error of a few percent rather than the current 10% would be vital input to much other interesting cosmology. There are several programmes which are likely to lead us to this point in the next 10 years.

  11. When constants are important

    Energy Technology Data Exchange (ETDEWEB)

    Beiu, V.

    1997-04-01

    In this paper the authors discuss several complexity aspects pertaining to neural networks, commonly known as the curse of dimensionality. The focus will be on: (1) size complexity and depth-size tradeoffs; (2) complexity of learning; and (3) precision and limited interconnectivity. Results have been obtained for each of these problems when dealt with separately, but few things are known as to the links among them. They start by presenting known results and try to establish connections between them. These show that they are facing very difficult problems--exponential growth in either space (i.e. precision and size) and/or time (i.e., learning and depth)--when resorting to neural networks for solving general problems. The paper will present a solution for lowering some constants, by playing on the depth-size tradeoff.

  12. Constant Proportion Portfolio Insurance

    DEFF Research Database (Denmark)

    Jessen, Cathrine

    2014-01-01

    Portfolio insurance, as practiced in 1987, consisted of trading between an underlying stock portfolio and cash, using option theory to place a floor on the value of the position, as if it included a protective put. Constant Proportion Portfolio Insurance (CPPI) is an option-free variation...... on the theme, originally proposed by Fischer Black. In CPPI, a financial institution guarantees a floor value for the “insured” portfolio and adjusts the stock/bond mix to produce a leveraged exposure to the risky assets, which depends on how far the portfolio value is above the floor. Plain-vanilla portfolio...... insurance largely died with the crash of 1987, but CPPI is still going strong. In the frictionless markets of finance theory, the issuer’s strategy to hedge its liability under the contract is clear, but in the real world with transactions costs and stochastic jump risk, the optimal strategy is less obvious...

  13. Decay constants in geochronology

    Institute of Scientific and Technical Information of China (English)

    IgorM.Villa; PaulR.Renne

    2005-01-01

    Geologic time is fundamental to the Earth Sciences, and progress in many disciplines depends critically on our ability to measure time with increasing accuracy and precision. Isotopic geochronology makes use of the decay of radioactive nuclides as a help to quantify the histories of rock, minerals, and other materials. Both accuracy and precision of radioisotopic ages are, at present, limited by those of radioactive decay constants. Modem mass spectrometers can measure isotope ratios with a precision of 10-4 or better. On the other hand, the uncertainties associated with direct half-life determinations are, in most cases, still at the percent level. The present short note briefly summarizes progress and problems that have been encountered during the Working Group's activity.

  14. High Affinity Heme Binding to a Heme Regulatory Motif on the Nuclear Receptor Rev-erbβ Leads to Its Degradation and Indirectly Regulates Its Interaction with Nuclear Receptor Corepressor.

    Science.gov (United States)

    Carter, Eric L; Gupta, Nirupama; Ragsdale, Stephen W

    2016-01-29

    Rev-erbα and Rev-erbβ are heme-binding nuclear receptors (NR) that repress the transcription of genes involved in regulating metabolism, inflammation, and the circadian clock. Previous gene expression and co-immunoprecipitation studies led to a model in which heme binding to Rev-erbα recruits nuclear receptor corepressor 1 (NCoR1) into an active repressor complex. However, in contradiction, biochemical and crystallographic studies have shown that heme decreases the affinity of the ligand-binding domain of Rev-erb NRs for NCoR1 peptides. One explanation for this discrepancy is that the ligand-binding domain and NCoR1 peptides used for in vitro studies cannot replicate the key features of the full-length proteins used in cellular studies. However, the combined in vitro and cellular results described here demonstrate that heme does not directly promote interactions between full-length Rev-erbβ (FLRev-erbβ) and an NCoR1 construct encompassing all three NR interaction domains. NCoR1 tightly binds both apo- and heme-replete FLRev-erbβ·DNA complexes; furthermore, heme, at high concentrations, destabilizes the FLRev-erbβ·NCoR1 complex. The interaction between FLRev-erbβ and NCoR1 as well as Rev-erbβ repression at the Bmal1 promoter appear to be modulated by another cellular factor(s), at least one of which is related to the ubiquitin-proteasome pathway. Our studies suggest that heme is involved in regulating the degradation of Rev-erbβ in a manner consistent with its role in circadian rhythm maintenance. Finally, the very slow rate constant (10(-6) s(-1)) of heme dissociation from Rev-erbβ rules out a prior proposal that Rev-erbβ acts as an intracellular heme sensor.

  15. Tsetse salivary gland proteins 1 and 2 are high affinity nucleic acid binding proteins with residual nuclease activity.

    Directory of Open Access Journals (Sweden)

    Guy Caljon

    Full Text Available Analysis of the tsetse fly salivary gland EST database revealed the presence of a highly enriched cluster of putative endonuclease genes, including tsal1 and tsal2. Tsal proteins are the major components of tsetse fly (G. morsitans morsitans saliva where they are present as monomers as well as high molecular weight complexes with other saliva proteins. We demonstrate that the recombinant tsetse salivary gland proteins 1&2 (Tsal1&2 display DNA/RNA non-specific, high affinity nucleic acid binding with K(D values in the low nanomolar range and a non-exclusive preference for duplex. These Tsal proteins exert only a residual nuclease activity with a preference for dsDNA in a broad pH range. Knockdown of Tsal expression by in vivo RNA interference in the tsetse fly revealed a partially impaired blood digestion phenotype as evidenced by higher gut nucleic acid, hematin and protein contents.

  16. Structure-based identification of new high-affinity nucleosome binding sequences.

    Science.gov (United States)

    Battistini, Federica; Hunter, Christopher A; Moore, Irene K; Widom, Jonathan

    2012-06-29

    The substrate for the proteins that express genetic information in the cell is not naked DNA but an assembly of nucleosomes, where the DNA is wrapped around histone proteins. The organization of these nucleosomes on genomic DNA is influenced by the DNA sequence. Here, we present a structure-based computational approach that translates sequence information into the energy required to bend DNA into a nucleosome-bound conformation. The calculations establish the relationship between DNA sequence and histone octamer binding affinity. In silico selection using this model identified several new DNA sequences, which were experimentally found to have histone octamer affinities comparable to the highest-affinity sequences known. The results provide insights into the molecular mechanism through which DNA sequence information encodes its organization. A quantitative appreciation of the thermodynamics of nucleosome positioning and rearrangement will be one of the key factors in understanding the regulation of transcription and in the design of new promoter architectures for the purposes of tuning gene expression dynamics.

  17. Oral zinc supplementation restore high molecular weight seminal zinc binding protein to normal value in Iraqi infertile men

    Directory of Open Access Journals (Sweden)

    Hadwan Mahmoud

    2012-11-01

    Full Text Available Abstract Background Zinc in human seminal plasma is divided into three types of ligands which are high (HMW, intermediate (IMW, and low molecular weight ligands (LMW. The present study was aimed to study the effect of Zn supplementation on the quantitative and qualitative characteristics of semen along with Zinc Binding Protein levels in the seminal plasma in asthenozoospermic patients. Methods Semen samples were obtained from 37 fertile and 37 asthenozoospermic infertile men with matched age. The subfertile group was treated with zinc sulfate, every participant took two capsules per day for three months (each one 220mg. Semen samples were obtained (before and after zinc sulfate supplementation. After liquefaction seminal fluid at room temperature, routine semen analyses were performed. For determination of the amount of zinc binding proteins, the gel filtration of seminal plasma on Sephadex G-75 was performed. All the fractions were investigated for protein and for zinc concentration by atomic absorption spectrophotometry. Evaluation of chromatograms was made directly from the zinc concentration in each fraction. Results A significant high molecular weight zinc binding ligands percentage (HMW-Zn % was observed in seminal plasma of fertile males compared with subfertile males. However, seminal low molecular weight ligands (LMW-Zn have opposite behavior. The mean value of semen volume, progressive sperm motility percentage and total normal sperm count were increased after zinc sulfate supplementation. Conclusions Zinc supplementation restores HMW-Zn% in seminal plasma of asthenozoospermic subjects to normal value. Zinc supplementation elevates LMW-Zn% in seminal plasma of asthenozoospermic subjects to more than normal value. Trial registration ClinicalTrials.gov identifier NCT01612403

  18. High avidity binding of engineered papaya mosaic virus virus-like particles to resting spores of Plasmodiophora brassicae.

    Science.gov (United States)

    Morin, Hélène; Tremblay, Marie-Hélène; Plante, Edith; Paré, Christine; Majeau, Nathalie; Hogue, Richard; Leclerc, Denis

    2007-02-01

    Papaya mosaic virus (PapMV) like particles (VLPs) were used as a platform for fusion of affinity peptides binding to resting spores of Plasmodiophora brassicae-a major pathogen of crucifers. Three peptides with specific affinity to the target were isolated and cloned at the C-terminus of the PapMV coat protein (CP), generating three different high avidity VLPs. The peptides were exposed at the surface of the VLPs and their avidity to resting spores of P. brassicae was measured by flow cytometry. NLP-A, with the peptide DPAPRPR, showed the highest avidity. The binding avidity of NLP-A to P. brassicae spores was comparable to that of a polyclonal antibody. NLP-A was also shown to be more specific than the antibody. Fusion of the affinity peptide to a monomeric form (mCP) of the CP [Lecours, K., Tremblay, M.-H., Laliberté Gagné, M.-E., Gagné, S.M., Leclerc, D., 2006. Purification and biochemical characterization of a monomeric form of papaya mosaic potexvirus coat protein. Protein Express. Purific. 47, 273-280] generated a fusion protein that was unable to assemble into VLPs, and mCP-A fusions failed to bind resting spores. The avidity of VLP-A was increased by adding a glycine spacer between the C-terminus of the PapMV CP and the peptide, and improved even further by using a duplicated A peptide in the fusion protein. The use of high avidity VLPs has advantages over polyclonal antibodies because of target specificity. VLPs offers the specificity of monoclonal antibodies but can be more easily generated using the powerful selection of phage display.

  19. Novel cyclic gamma-hydroxybutyrate (GHB) analogs with high affinity and stereoselectivity of binding to GHB sites in rat brain.

    Science.gov (United States)

    Wellendorph, Petrine; Høg, Signe; Greenwood, Jeremy R; de Lichtenberg, Anne; Nielsen, Birgitte; Frølund, Bente; Brehm, Lotte; Clausen, Rasmus P; Bräuner-Osborne, Hans

    2005-10-01

    Gamma-hydroxybutyrate (GHB) is a psychotropic compound endogenous to the brain. Despite its potentially great physiological significance, its exact molecular mechanism of action is unknown. GHB is a weak agonist at GABA(B) receptors, but there is also evidence of specific GHB receptor sites, the molecular cloning of which remains a challenge. Ligands with high affinity and specificity for the reported GHB binding site are needed for pharmacological dissection of the GHB and GABA(B) effects and for mapping the structural requirements of the GHB receptor-ligand interactions. For this purpose, we have synthesized and assayed three conformationally restricted GHB analogs for binding against the GHB-specific ligand [3H]NCS-382 [(E,RS)-(6,7,8,9-tetrahydro-5-hydroxy-5H-benzocyclohept-6-ylidene-)acetic acid] in rat brain homogenate. The cyclohexene and cyclopentene analogs, 3-hydroxycyclohex-1-enecarboxylic acid [(RS)-HOCHCA] and 3-hydroxycyclopent-1-enecarboxylic acid [(RS)-HOCPCA], were found to be high-affinity GHB ligands, with IC50 values in the nanomolar range, and had 9 and 27 times, respectively, higher affinity than GHB. The stereo-selectively synthesized R,R-isomer of the trans-cyclopropyl GHB analog, HOCPrCA, proved to have 10-fold higher affinity than its enantiomer. Likewise, the R-enantiomers of HOCHCA and HOCPCA selectively inhibited [3H]NCS-382 binding. The best inhibitor of these, (R)-HOCPCA, has an affinity 39 times higher than GHB and is thus among the best GHB ligands reported to date. Neither of the cycloalkenes showed any affinity (IC50 > 1 mM) for GABA(A) or GABA(B) receptors. These compounds show excellent potential as lead structures and novel tools for studying specific GHB receptor-mediated pharmacology.

  20. Plasma Levels of Fatty Acid-Binding Protein 4, Retinol-Binding Protein 4, High-Molecular-Weight Adiponectin, and Cardiovascular Mortality Among Men With Type 2 Diabetes: A 22-Year Prospective Study.

    Science.gov (United States)

    Liu, Gang; Ding, Ming; Chiuve, Stephanie E; Rimm, Eric B; Franks, Paul W; Meigs, James B; Hu, Frank B; Sun, Qi

    2016-11-01

    To examine select adipokines, including fatty acid-binding protein 4, retinol-binding protein 4, and high-molecular-weight (HMW) adiponectin in relation to cardiovascular disease (CVD) mortality among patients with type 2 diabetes mellitus. Plasma levels of fatty acid-binding protein 4, retinol-binding protein 4, and HMW adiponectin were measured in 950 men with type 2 diabetes mellitus in the Health Professionals Follow-up Study. After an average of 22 years of follow-up (1993-2015), 580 deaths occurred, of whom 220 died of CVD. After multivariate adjustment for covariates, higher levels of fatty acid-binding protein 4 were significantly associated with a higher CVD mortality: comparing extreme tertiles, the hazard ratio and 95% confidence interval of CVD mortality was 1.78 (1.22-2.59; P trend=0.001). A positive association was also observed for HMW adiponectin: the hazard ratio (95% confidence interval) was 2.07 (1.42-3.06; P trend=0.0002), comparing extreme tertiles, whereas higher retinol-binding protein 4 levels were nonsignificantly associated with a decreased CVD mortality with an hazard ratio (95% confidence interval) of 0.73 (0.50-1.07; P trend=0.09). A Mendelian randomization analysis suggested that the causal relationships of HMW adiponectin and retinol-binding protein 4 would be directionally opposite to those observed based on the biomarkers, although none of the Mendelian randomization associations achieved statistical significance. These data suggest that higher levels of fatty acid-binding protein 4 and HMW adiponectin are associated with elevated CVD mortality among men with type 2 diabetes mellitus. Biological mechanisms underlying these observations deserve elucidation, but the associations of HMW adiponectin may partially reflect altered adipose tissue functionality among patients with type 2 diabetes mellitus. © 2016 American Heart Association, Inc.

  1. Targeted deletion of a high-affinity GATA-binding site in the GATA-1 promoter leads to selective loss of the eosinophil lineage in vivo

    National Research Council Canada - National Science Library

    Yu, Channing; Cantor, Alan B; Yang, Haidi; Browne, Carol; Wells, Richard A; Fujiwara, Yuko; Orkin, Stuart H

    2002-01-01

    .... Here we demonstrate that deletion of a high-affinity GATA-binding site in the GATA-1 promoter, an element presumed to mediate positive autoregulation of GATA-1 expression, leads to selective loss...

  2. Involvement of nitrogen functional groups in high-affinity copper binding in tomato and wheat root apoplasts: spectroscopic and thermodynamic evidence.

    Science.gov (United States)

    Guigues, Stéphanie; Bravin, Matthieu N; Garnier, Cédric; Masion, Armand; Chevassus-Rosset, Claire; Cazevieille, Patrick; Doelsch, Emmanuel

    2016-03-01

    Carboxylic groups located in plant cell walls (CW) are generally considered to be the main copper binding sites in plant roots, despite the presence of other functional groups. The aim of this study was to investigate sites responsible for copper binding in root apoplasts, i.e. CW and outer surface of the plasma membrane (PM) continuum. Binding sites in root apoplasts were investigated by comparing isolated CW of a monocotyledon (Triticum aestivum L.) and dicotyledon (Solanum lycopersicum L.) crop with their respective whole roots. Copper speciation was examined by X-ray absorption (XAS) and (13)C-nuclear magnetic resonance spectroscopies while the affinity of ligands involved in copper binding was investigated by modeling copper sorption isotherms. Homogeneous speciation and binding of copper was found in wheat and tomato root apoplasts. Only Cu-N and Cu-O bonds were detected in wheat and tomato root apoplasts. Nitrogen/oxygen ligands were identified in slightly higher proportions (40-70%) than single oxygen ligands. Furthermore, low- and high-affinity binding sites contributed in an almost equivalent proportion to copper binding in root apoplasts. The high-affinity N functional groups embedded in root apoplasts participated in copper binding in the same magnitude than the low-affinity carboxylic groups.

  3. High Throughput Sequencing Identifies Misregulated Genes in the Drosophila Polypyrimidine Tract-Binding Protein (hephaestus) Mutant Defective in Spermatogenesis.

    Science.gov (United States)

    Sridharan, Vinod; Heimiller, Joseph; Robida, Mark D; Singh, Ravinder

    2016-01-01

    The Drosophila polypyrimidine tract-binding protein (dmPTB or hephaestus) plays an important role during spermatogenesis. The heph2 mutation in this gene results in a specific defect in spermatogenesis, causing aberrant spermatid individualization and male sterility. However, the array of molecular defects in the mutant remains uncharacterized. Using an unbiased high throughput sequencing approach, we have identified transcripts that are misregulated in this mutant. Aberrant transcripts show altered expression levels, exon skipping, and alternative 5' ends. We independently verified these findings by reverse-transcription and polymerase chain reaction (RT-PCR) analysis. Our analysis shows misregulation of transcripts that have been connected to spermatogenesis, including components of the actomyosin cytoskeletal apparatus. We show, for example, that the Myosin light chain 1 (Mlc1) transcript is aberrantly spliced. Furthermore, bioinformatics analysis reveals that Mlc1 contains a high affinity binding site(s) for dmPTB and that the site is conserved in many Drosophila species. We discuss that Mlc1 and other components of the actomyosin cytoskeletal apparatus offer important molecular links between the loss of dmPTB function and the observed developmental defect in spermatogenesis. This study provides the first comprehensive list of genes misregulated in vivo in the heph2 mutant in Drosophila and offers insight into the role of dmPTB during spermatogenesis.

  4. An engineered high affinity Fbs1 carbohydrate binding protein for selective capture of N-glycans and N-glycopeptides

    Science.gov (United States)

    Chen, Minyong; Shi, Xiaofeng; Duke, Rebecca M.; Ruse, Cristian I.; Dai, Nan; Taron, Christopher H.; Samuelson, James C.

    2017-01-01

    A method for selective and comprehensive enrichment of N-linked glycopeptides was developed to facilitate detection of micro-heterogeneity of N-glycosylation. The method takes advantage of the inherent properties of Fbs1, which functions within the ubiquitin-mediated degradation system to recognize the common core pentasaccharide motif (Man3GlcNAc2) of N-linked glycoproteins. We show that Fbs1 is able to bind diverse types of N-linked glycomolecules; however, wild-type Fbs1 preferentially binds high-mannose-containing glycans. We identified Fbs1 variants through mutagenesis and plasmid display selection, which possess higher affinity and improved recovery of complex N-glycomolecules. In particular, we demonstrate that the Fbs1 GYR variant may be employed for substantially unbiased enrichment of N-linked glycopeptides from human serum. Most importantly, this highly efficient N-glycopeptide enrichment method enables the simultaneous determination of N-glycan composition and N-glycosites with a deeper coverage (compared to lectin enrichment) and improves large-scale N-glycoproteomics studies due to greatly reduced sample complexity. PMID:28534482

  5. Transcriptome-Based Identification of Highly Similar Odorant-Binding Proteins among Neotropical Stink Bugs and Their Egg Parasitoid.

    Directory of Open Access Journals (Sweden)

    Luciana R Farias

    Full Text Available Olfaction plays a fundamental role in insect survival through resource location and intra and interspecific communications. We used RNA-Seq to analyze transcriptomes for odorant-binding proteins (OBPs from major stink bug pest species in Brazil, Euschistus heros, Chinavia ubica, and Dichelops melacanthus, and from their egg parasitoid, Telenomus podisi. We identified 23 OBPs in E. heros, 25 OBPs in C. ubica, 9 OBPs in D. melacanthus, and 7 OBPs in T. podisi. The deduced amino acid sequences of the full-length OBPs had low intraspecific similarity, but very high similarity between two pairs of OBPs from E. heros and C. ubica (76.4 and 84.0% and between two pairs of OBPs from the parasitoid and its preferred host E. heros (82.4 and 88.5%, confirmed by a high similarity of their predicted tertiary structures. The similar pairs of OBPs from E. heros and C. ubica may suggest that they have derived from a common ancestor, and retain the same biological function to bind a ligand perceived or produced in both species. The T. podisi OBPs similar to E. heros were not orthologous to any known hymenopteran OBPs, and may have evolved independently and converged to the host OBPs, providing a possible basis for the host location of T. podisi using E. heros semiochemical cues.

  6. An engineered high affinity Fbs1 carbohydrate binding protein for selective capture of N-glycans and N-glycopeptides.

    Science.gov (United States)

    Chen, Minyong; Shi, Xiaofeng; Duke, Rebecca M; Ruse, Cristian I; Dai, Nan; Taron, Christopher H; Samuelson, James C

    2017-05-23

    A method for selective and comprehensive enrichment of N-linked glycopeptides was developed to facilitate detection of micro-heterogeneity of N-glycosylation. The method takes advantage of the inherent properties of Fbs1, which functions within the ubiquitin-mediated degradation system to recognize the common core pentasaccharide motif (Man3GlcNAc2) of N-linked glycoproteins. We show that Fbs1 is able to bind diverse types of N-linked glycomolecules; however, wild-type Fbs1 preferentially binds high-mannose-containing glycans. We identified Fbs1 variants through mutagenesis and plasmid display selection, which possess higher affinity and improved recovery of complex N-glycomolecules. In particular, we demonstrate that the Fbs1 GYR variant may be employed for substantially unbiased enrichment of N-linked glycopeptides from human serum. Most importantly, this highly efficient N-glycopeptide enrichment method enables the simultaneous determination of N-glycan composition and N-glycosites with a deeper coverage (compared to lectin enrichment) and improves large-scale N-glycoproteomics studies due to greatly reduced sample complexity.

  7. Constant Proportion Debt Obligations (CPDOs)

    DEFF Research Database (Denmark)

    Cont, Rama; Jessen, Cathrine

    2012-01-01

    be made arbitrarily small—and thus the credit rating arbitrarily high—by increasing leverage, but the ratings obtained strongly depend on assumptions on the credit environment (high spread or low spread). More importantly, CPDO loss distributions are found to exhibit a wide range of tail risk measures......Constant Proportion Debt Obligations (CPDOs) are structured credit derivatives that generate high coupon payments by dynamically leveraging a position in an underlying portfolio of investment-grade index default swaps. CPDO coupons and principal notes received high initial credit ratings from...... the major rating agencies, based on complex models for the joint transition of ratings and spreads for all names in the underlying portfolio. We propose a parsimonious model for analysing the performance of CPDO strategies using a top-down approach that captures the essential risk factors of the CPDO. Our...

  8. High-accuracy measurements of OH(•) reaction rate constants and IR and UV absorption spectra: ethanol and partially fluorinated ethyl alcohols.

    Science.gov (United States)

    Orkin, Vladimir L; Khamaganov, Victor G; Martynova, Larissa E; Kurylo, Michael J

    2011-08-11

    Rate constants for the gas phase reactions of OH(•) radicals with ethanol and three fluorinated ethyl alcohols, CH(3)CH(2)OH (k(0)), CH(2)FCH(2)OH (k(1)), CHF(2)CH(2)OH (k(2)), and CF(3)CH(2)OH (k(3)) were measured using a flash photolysis resonance-fluorescence technique over the temperature range 220 to 370 K. The Arrhenius plots were found to exhibit noticeable curvature for all four reactions. The temperature dependences of the rate constants can be represented by the following expressions over the indicated temperature intervals: k(0)(220-370 K) = 5.98 × 10(-13)(T/298)(1.99) exp(+515/T) cm(3) molecule(-1) s(-1), k(0)(220-298 K) = (3.35 ± 0.06) × 10(-12) cm(3) molecule(-1) s(-1) [for atmospheric modeling purposes, k(0)(T) is essentially temperature-independent below room temperature, k(0)(220-298 K) = (3.35 ± 0.06) × 10(-12) cm(3) molecule(-1) s(-1)], k(1)(230-370 K) = 3.47 × 10(-14)(T/298)(4.49) exp(+977/T) cm(3) molecule(-1) s(-1), k(2)(220-370 K) = 3.87 × 10(-14)(T/298)(4.25) exp(+578/T) cm(3) molecule(-1) s(-1), and k(3)(220-370 K) = 2.48 × 10(-14)(T/298)(4.03) exp(+418/T) cm(3) molecule(-1) s(-1). The atmospheric lifetimes due to reactions with tropospheric OH(•) were estimated to be 4, 16, 62, and 171 days, respectively, under the assumption of a well-mixed atmosphere. UV absorption cross sections of all four ethanols were measured between 160 and 215 nm. The IR absorption cross sections of the three fluorinated ethanols were measured between 400 and 1900 cm(-1), and their global warming potentials were estimated.

  9. High throughput techniques for discovering new glycine receptor modulators and their binding sites

    Directory of Open Access Journals (Sweden)

    Daniel F Gilbert

    2009-10-01

    Full Text Available The inhibitory glycine receptor (GlyR is a member of the Cys-loop receptor family that mediates inhibitory neurotransmission in the central nervous system. These receptors are emerging as potential drug targets for inflammatory pain, immunomodulation, spasticity and epilepsy. Antagonists that specifically inhibit particular GlyR isoforms are also required as pharmacological probes for elucidating the roles of particular GlyR isoforms in health and disease. Although a substantial number of both positive and negative GlyR modulators have been identified, very few of these are specific for the GlyR over other receptor types. Thus, the potential of known compounds as either therapeutic leads or pharmacological probes is limited. It is therefore surprising that there have been few published studies describing attempts to discover novel GlyR isoform-specific compounds. The first aim of this review is to consider various methods for efficiently screening compounds against these receptors. We conclude that an anion sensitive yellow fluorescent protein is optimal for primary screening and that automated electrophysiology of cells stably expressing GlyRs is useful for confirming hits and quantitating the actions of identified compounds. The second aim of this review is to demonstrate how these techniques are used in our laboratory for the purpose of both discovering novel GlyR-active compounds and characterizing their binding sites. We also describe a reliable, cost effective method for transfecting HEK293 cells in single wells of a 384 well plate using nanogram quantities of cDNA.

  10. High protein binding and cidal activity against penicillin-resistant S. pneumoniae: a cefditoren in vitro pharmacodynamic simulation.

    Directory of Open Access Journals (Sweden)

    David Sevillano

    Full Text Available BACKGROUND: Although protein binding is a reversible phenomenon, it is assumed that antibacterial activity is exclusively exerted by the free (unbound fraction of antibiotics. METHODOLOGY/PRINCIPAL FINDINGS: Activity of cefditoren, a highly protein bound 3(rd generation cephalosporin, over 24h after an oral 400 mg cefditoren-pivoxil bid regimen was studied against six S. pneumoniae strains (penicillin/cefditoren MICs; microg/ml: S1 (0.12/0.25, S2 (0.25/0.25, S3 and S4 (0.5/0.5, S5 (1/0.5 and S6 (4/0.5. A computerized pharmacodynamic simulation with media consisting in 75% human serum and 25% broth (mean albumin concentrations = 4.85+/-0.12 g/dL was performed. Protein binding was measured. The cumulative percentage of a 24h-period that drug concentrations exceeded the MIC for total (T > MIC and unbound concentrations (fT > MIC, expressed as percentage of the dosing interval, were determined. Protein binding was 87.1%. Bactericidal activity (> or = 99.9% initial inocula reduction was obtained against strains S1 and S2 at 24h (T > MIC = 77.6%, fT > MIC = 23.7%. With T > MIC of 61.6% (fT > MIC = 1.7%, reductions against S3 and S4 ranged from 90% to 97% at 12h and 24h; against S5, reduction was 45.1% at 12h and up to 85.0% at 24h; and against S6, reduction was 91.8% at 12h, but due to regrowth of 52.9% at 24h. Cefditoren physiological concentrations exerted antibacterial activity against strains exhibiting MICs of 0.25 and 0.5 microg/ml under protein binding conditions similar to those in humans. CONCLUSIONS/SIGNIFICANCE: The results of this study suggest that, from the pharmacodynamic perspective, the presence of physiological albumin concentrations may not preclude antipneumococcal activity of highly bound cephalosporins as cefditoren.

  11. The Application of Frequency Converter in Constant Pressure Water Supply System of High-rize Buildings%变频器在高层建筑恒压供水系统中的应用

    Institute of Scientific and Technical Information of China (English)

    杨洪升

    2015-01-01

    Using frequency control method to enable water pump motor keep the pressure constant in water supply system of high-rise building’s construction, can reach the target that high-rise building's water supply capacity changes with the water flow. By the use of Siemens frequency converter to control the water pump motor, the article introduces the composi⁃tion of variable frequency constant pressure water supply system, the PID regulatory function of Siemens inverter, the set⁃ting method of PID parameters and briefly introduces the control scheme of frequency converter in constant pressure water supply system of high-rise building’s construction . Results show that the constant pressure water supply system con⁃trolled by Siemens inverter in high-rise building’s construction can ensure the of constant pressure water supply, at the same time has achieved remarkable energy saving effect. At present, constant pressure water supply system controlled by the Siemens inverter has been widely used.%采用对水泵电机变频调速的方法,使高层建筑施工供水系统压力恒定,可达到高层建筑供水能力随用水流量而变化的目标。本文通过使用西门子变频器控制水泵电机,介绍了变频恒压供水系统的构成、西门子变频器的PID调节功能、PID参数的设置方法并简要介绍了变频器在高层建筑施工恒压供水系统中的控制方案。结果表明西门子变频器控制的恒压供水系统在实现高层建筑施工恒压供水的同时,达到了显著的节能效果。目前西门子变频器控制的恒压供水系统已经得到了广泛的应用。

  12. High Precision and High Yield Fabrication of Dense Nanoparticle Arrays onto DNA Origami at Statistically Independent Binding Sites †

    OpenAIRE

    Takabayashi, Sadao; Klein, William P.; Onodera, Craig; Rapp, Blake; Flores-Estrada, Juan; Lindau, Elias; Snowball, Lejmarc; Sam, Joseph Tyler; Padilla, Jennifer E.; Lee, Jeunghoon; Knowlton, William B.; Graugnard, Elton; Yurke, Bernard; Kuang, Wan; Hughes, William L.

    2014-01-01

    High precision, high yield, and high density self-assembly of nanoparticles into arrays is essential for nanophotonics. Spatial deviations as small as a few nanometers can alter the properties of near-field coupled optical nanostructures. Several studies have reported assemblies of few nanoparticle structures with controlled spacing using DNA nanostructures with variable yield. Here, we report multi-tether design strategies and attachment yields for homo- and hetero-nanoparticle arrays templa...

  13. String theory, cosmology and varying constants

    Science.gov (United States)

    Damour, Thibault

    In string theory the coupling `constants' appearing in the low-energy effective Lagrangian are determined by the vacuum expectation values of some (a priori) massless scalar fields (dilaton, moduli). This naturally leads one to expect a correlated variation of all the coupling constants, and an associated violation of the equivalence principle. We review some string-inspired theoretical models which incorporate such a spacetime variation of coupling constants while remaining naturally compatible both with phenomenological constraints coming from geochemical data (Oklo; Rhenium decay) and with present equivalence principle tests. Barring a very unnatural fine-tuning of parameters, a variation of the fine-structure constant as large as that recently `observed' by Webb et al. in quasar absorption spectra appears to be incompatible with these phenomenological constraints. Independently of any model, it is emphasized that the best experimental probe of varying constants are high-precision tests of the universality of free fall, such as MICROSCOPE and STEP.

  14. String theory, cosmology and varying constants

    CERN Document Server

    Damour, Thibault Marie Alban Guillaume

    2002-01-01

    In string theory the coupling ``constants'' appearing in the low-energy effective Lagrangian are determined by the vacuum expectation values of some (a priori) massless scalar fields (dilaton, moduli). This naturally leads one to expect a correlated variation of all the coupling constants, and an associated violation of the equivalence principle. We review some string-inspired theoretical models which incorporate such a spacetime variation of coupling constants while remaining naturally compatible both with phenomenological constraints coming from geochemical data (Oklo; Rhenium decay) and with present equivalence principle tests. Barring a very unnatural fine-tuning of parameters, a variation of the fine-structure constant as large as that recently ``observed'' by Webb et al. in quasar absorption spectra appears to be incompatible with these phenomenological constraints. Independently of any model, it is emphasized that the best experimental probe of varying constants are high-precision tests of the universa...

  15. High-order above-threshold ionization the uniform approximation and the effect of the binding potential

    CERN Document Server

    De Faria, C F M; Becker, W

    2002-01-01

    A versatile semiclassical approximation for intense laser-atom processes is presented. This uniform approximation is no more complicated than the frequently-used multi-dimensional saddle-point approximation and far superior, since it applies for all energies, both close to as well as away from classical cutoffs. In the latter case, it reduces to the standard saddle-point approximation. The uniform approximation agrees accurately with numerical evaluations for potentials, for which these are feasible, and constitutes a practicable method of calculation in general. The method is applied to the calculation of high-order above-threshold ionization spectra with various binding potentials: Coulomb, Yukawa, and shell potentials which may model C$_{60}$ molecules or clusters. The shell potentials generate high-order ATI spectra that are more structured and may feature an apparently higher cutoff.

  16. The presence of high-affinity, low-capacity estradiol-17β binding in rainbow trout scale indicates a possible endocrine route for the regulation of scale resorption

    Science.gov (United States)

    Persson, Petra; Shrimpton, J.M.; McCormick, S.D.; Bjornsson, Bjorn Thrandur

    2000-01-01

    High-affinity, low-capacity estradiol-17β (E2) binding is present in rainbow trout scale. The Kd and Bmax of the scale E2 binding are similar to those of the liver E2 receptor (Kd is 1.6 ± 0.1 and 1.4 ± 0.1 nM, and Bmax is 9.1 ± 1.2 and 23.1 ± 2.2 fmol x mg protein-1, for scale and liver, respectively), but different from those of the high-affinity, low-capacity E2 binding in plasma (Kd is 4.0 ± 0.4 nM and Bmax is 625.4 ± 63.1 fmol x mg protein-1). The E2 binding in scale was displaced by testosterone, but not by diethylstilbestrol. Hence, the ligand binding specificity is different from that of the previously characterized liver E2 receptor, where E2 is displaced by diethylstilbestrol, but not by testosterone. The putative scale E2 receptor thus appears to bind both E2 and testosterone, and it is proposed that the increased scale resorption observed during sexual maturation in both sexes of several salmonid species may be mediated by this receptor. No high-affinity, low-capacity E2 binding could be detected in rainbow trout gill or skin.

  17. Mannosylerythritol lipid, a yeast extracellular glycolipid, shows high binding affinity towards human immunoglobulin G

    OpenAIRE

    Ikegami Toru; Yanagishita Hiroshi; Nakane Takashi; Im Jae Hong; Kitamoto Dai

    2001-01-01

    Abstract Background There have been many attempts to develop new materials with stability and high affinity towards immunoglobulins. Some of glycolipids such as gangliosides exhibit a high affinity toward immunoglobulins. However, it is considerably difficult to develop these glycolipids into the practical separation ligand due to their limited amounts. We thus focused our attention on the feasible use of "mannosylerythritol lipid A", a yeast glycolipid biosurfactant, as an alternative ligand...

  18. Comparison of high affinity binding of {sup 3}H-proadifen and {sup 3}H-(-)-cocaine t rat liver membranes

    Energy Technology Data Exchange (ETDEWEB)

    Ross, S.B. [Astra Arcus AB, Dept. of Neuropharmacology, Soedertaelje (Sweden)

    1995-06-01

    The characteristics of the binding of {sup 3}H-proadifen to rat liver membranes were studied and compared to those of {sup 3}H-cocaine. It was found that {sup 3}H-proadifen was bound reversibly with high affinity (K{sub D}=1.8{+-}0.5 nM) and large capacity (B{sub max}=2010{+-}340 pmol/g wet tissue) to liver membranes. The corresponding values for the {sup 3}H-cocaine binding were 3.5 nM and 1000 pmol/g wet tissue. The binding of {sup 3}H-proadifen was mainly localised to the microsomal fraction. The number of binding sites was not increased by treatment of rats with phenobarbitone. With 1 {mu}M CdCl{sub 2} in the incubation buffer it was possible to differentiate between two {sup 3}H-cocaine binding sites with K{sub d} values of 1.6 and 7.7 nM and B{sub max} values of 280 and 940 pmol/g wet liver tissue. S-(-)-Alaproclate inhibited the binding of {sup 3}H-proadifen and {sup 3}H-cocaine inhibited the binding of {sup 3}H-proadifen (IC{sub 50}=10 nM) and proadifen that of {sup 3}H-cocaine (IC{sub 50}=1 nM). There was a high correlation coefficient (r{sub r}=0.972; P<0.01; n=12) in the Spearman rank test between the inhibitory potencies of compounds examined in both systems. Beside some potent alaproclate analogues a couple of compounds had moderately high affinity (IC{sub 50}=100-500 nM): chloroquine, phenoxybenzamine, amitriptyline, ajmaline, remoxipride, imipramine and (-)-alaprenolol. CdCl{sub 2}, ZnCl{sub 2} and CuCl{sub 2} inhibited the binding of both ligands with low Hill coefficients, indicating heterogeneous binding sites. The inhibition curve of Cd{sup 2+} on the cocaine binding was biphasic with a high affinity part around 50 nM and a low affinity part at 15{mu}M. The similarity of the characteristics of the binding of these ligands with that of {sup 3}H-alaproclate is discussed. It is suggested that all three compounds bind to the same sites, although additional binding sites seem to exist for proadifen. (au) (9 refs.).

  19. Effects of Midgut-Protein-Preparative and Ligand Binding Procedures on the Toxin Binding Characteristics of BT-R1, a Common High-Affinity Receptor in Manduca sexta for Cry1A Bacillus thuringiensis Toxins

    Science.gov (United States)

    Keeton, Timothy P.; Francis, Brian R.; Maaty, Walid S. A.; Bulla, Lee A.

    1998-01-01

    The identity of the physiologically important Cry1A receptor protein(s) in the lepidopteran Manduca sexta has been a matter of dispute due to the multiple proteins which bind the Cry1Ac toxin. Cry1Aa, Cry1Ab, and Cry1Ac exhibit essentially identical toxicities toward M. sexta larvae and show a high degree of sequence and presumed structural identities. These similarities make it likely that there is a common mechanism of toxicity in these lepidopteran-specific toxins in terms of both mode of action and the receptor proteins through which these toxins exert their lepidopteran-specific toxicity. Investigators in our laboratory previously demonstrated that the cloned 210-kDa glycoprotein BT-R1 binds all three Cry1A toxins (T. P. Keeton and L. A. Bulla, Jr., Appl. Environ. Microbiol. 63:3419–3425, 1997). This protein remains a common binding protein even after being subjected to various midgut membrane preparation and processing protocols. The method used to isolate proteins from the M. sexta larval midgut in no significant way affects the results of ligand binding and vacuum blotting experiments, and we have been unable to detect specific, high-affinity binding of any Cry1A toxin to Cry1Ac binding proteins other than BT-R1. Alterations in blot substrate and blocking, hybridization, and washing buffers support these conclusions. Collectively, these results indicate that in M. sexta the cadherin-like BT-R1 protein is a common high-affinity receptor protein for the Cry1A family of toxins. PMID:9603829

  20. Joint Binding of OTX2 and MYC in Promotor Regions Is Associated with High Gene Expression in Medulloblastoma

    NARCIS (Netherlands)

    Bunt, J.; Hasselt, N.E.; Zwijnenburg, D.A.; Koster, J.; Versteeg, R.; Kool, M.

    2011-01-01

    Both OTX2 and MYC are important oncogenes in medulloblastoma, the most common malignant brain tumor in childhood. Much is known about MYC binding to promoter regions, but OTX2 binding is hardly investigated. We used ChIP-on-chip data to analyze the binding patterns of both transcription factors in D

  1. The Interacting and Non-constant Cosmological Constant

    CERN Document Server

    Verma, Murli Manohar

    2009-01-01

    We propose a time-varying cosmological constant with a fixed equation of state, which evolves mainly through its interaction with the background during most of the long history of the universe. However, such interaction does not exist in the very early and the late-time universe and produces the acceleration during these eras when it becomes very nearly a constant. It is found that after the initial inflationary phase, the cosmological constant, that we call as lambda parameter, rolls down from a large constant value to another but very small constant value and further dominates the present epoch showing up in form of the dark energy driving the acceleration.

  2. High Precision Momentum Calibration of the Magnetic Spectrometers at MAMI for Hypernuclear Binding Energy Determination

    CERN Document Server

    Margaryan, A; Achenbach, P; Ajvazyan, R; Elbakyan, H; Montgomery, R; Nakamura, S N; Pochodzalla, J; Schulz, F; Toyama, Y; Zhamkochyan, S

    2016-01-01

    We propose a new method for absolute momentum calibration of magnetic spectrometers used in nuclear physics, using the time-of-flight (TOF), differences of pairs of particles with different masses. In cases where the flight path is not known, a calibration can be determined by using the TOF differences of two pair combinations of three particles. A Cherenkov detector, read out by a radio frequency photomultiplier tube, is considered as the high-resolution and highly stable TOF detector. By means of Monte Carlo simulations it is demonstrated that the magnetic spectrometers at the MAMI electron-scattering facility can be calibrated absolutely with an accuracy $\\delta p/p\\leq 10^{-4}$, which will be crucial for high precision determination of hypernuclear masses.

  3. Quantum Theory without Planck's Constant

    CERN Document Server

    Ralston, John P

    2012-01-01

    Planck's constant was introduced as a fundamental scale in the early history of quantum mechanics. We find a modern approach where Planck's constant is absent: it is unobservable except as a constant of human convention. Despite long reference to experiment, review shows that Planck's constant cannot be obtained from the data of Ryberg, Davisson and Germer, Compton, or that used by Planck himself. In the new approach Planck's constant is tied to macroscopic conventions of Newtonian origin, which are dispensable. The precision of other fundamental constants is substantially improved by eliminating Planck's constant. The electron mass is determined about 67 times more precisely, and the unit of electric charge determined 139 times more precisely. Improvement in the experimental value of the fine structure constant allows new types of experiment to be compared towards finding "new physics." The long-standing goal of eliminating reliance on the artifact known as the International Prototype Kilogram can be accompl...

  4. Binding of complement inhibitor C4b-binding protein to a highly virulent Streptococcus pyogenes M1 strain is mediated by protein H and enhances adhesion to and invasion of endothelial cells.

    Science.gov (United States)

    Ermert, David; Weckel, Antonin; Agarwal, Vaibhav; Frick, Inga-Maria; Björck, Lars; Blom, Anna M

    2013-11-08

    Streptococcus pyogenes AP1, a strain of the highly virulent M1 serotype, uses exclusively protein H to bind the complement inhibitor C4b-binding protein (C4BP). We found a strong correlation between the ability of AP1 and its isogenic mutants lacking protein H to inhibit opsonization with complement C3b and binding of C4BP. C4BP bound to immobilized protein H or AP1 bacteria retained its cofactor activity for degradation of (125)I-C4b. Furthermore, C4b deposited from serum onto AP1 bacterial surfaces was processed into C4c/C4d fragments, which did not occur on strains unable to bind C4BP. Recombinant C4BP mutants, which (i) lack certain CCP domains or (ii) have mutations in single aa as well as (iii) mutants with additional aa between different CCP domains were used to determine that the binding is mainly mediated by a patch of positively charged amino acid residues at the interface of domains CCP1 and CCP2. Using recombinant protein H fragments, we narrowed down the binding site to the N-terminal domain A. With a peptide microarray, we identified one single 18-amino acid-long peptide comprising residues 92-109, which specifically bound C4BP. Biacore was used to determine KD = 6 × 10(-7) M between protein H and a single subunit of C4BP. C4BP binding also correlated with elevated levels of adhesion and invasion to endothelial cells. Taken together, we identified the molecular basis of C4BP-protein H interaction and found that it is not only important for decreased opsonization but also for invasion of endothelial cells by S. pyogenes.

  5. Protein imprinted ionic liquid polymer on the surface of multiwall carbon nanotubes with high binding capacity for lysozyme.

    Science.gov (United States)

    Yuan, Shifang; Deng, Qiliang; Fang, Guozhen; Wu, Jianhua; Li, Wangwang; Wang, Shuo

    2014-06-01

    In this research, ionic liquid as functional monomer to prepare molecularly imprinted polymers for protein recognition was for the first time demonstrated, in which, 1-vinyl-3-butylimidazolium chloride was selected as functional monomer, acrylamide as co-functional monomer and lysozyme (Lyz) as template protein to synthesize imprinted polymers on the surface of multiwall carbon nanotubes in aqueous medium. The results indicated that ionic liquid was helpful to improve binding capacity of imprinted polymers, which had a maximum binding capacity of 763.36 mg/g in the optimum adsorption conditions. The prepared imprinted polymers had a fast adsorption rate and a much higher adsorption capacity than the corresponding non-imprinted polymers, with the difference in adsorption capacity up to 258.31 mg/g. The obtained polymer was evaluated by Lyz, bovine serum albumin (BSA), bovine hemoglobin (BHb), equine myoglobin (MB) and cytochrome c (Cyt c). The selectivity factor (β) for Lyz/BSA, Lyz/Mb, Lyz/BHb, and Lyz/Cyt c were 7.17, 2.12, 1.76 and 1.57, respectively, indicating the imprinted polymers had a good selectivity. Easy preparation of the imprinted polymers as well as high ability and selectivity to adsorb template proteins makes this polymer attractive and broadly applicable in biomacromolecular separation, biotechnology and sensors.

  6. A highly Conserved Aspartic Acid Residue of the Chitosanase from Bacillus Sp. TS Is Involved in the Substrate Binding.

    Science.gov (United States)

    Zhou, Zhanping; Zhao, Shuangzhi; Liu, Yang; Chang, Zhengying; Ma, Yanhe; Li, Jian; Song, Jiangning

    2016-11-01

    The chitosanase from Bacillus sp. TS (CsnTS) is an enzyme belonging to the glycoside hydrolase family 8. The sequence of CsnTS shares 98 % identity with the chitosanase from Bacillus sp. K17. Crystallography analysis and site-direct mutagenesis of the chitosanase from Bacillus sp. K17 identified the important residues involved in the catalytic interaction and substrate binding. However, despite progress in understanding the catalytic mechanism of the chitosanase from the family GH8, the functional roles of some residues that are highly conserved throughout this family have not been fully elucidated. This study focused on one of these residues, i.e., the aspartic acid residue at position 318. We found that apart from asparagine, mutation of Asp318 resulted in significant loss of enzyme activity. In-depth investigations showed that mutation of this residue not only impaired enzymatic activity but also affected substrate binding. Taken together, our results showed that Asp318 plays an important role in CsnTS activity.

  7. Engineering a Highly Hydrophilic PVDF Membrane via Binding TiO₂Nanoparticles and a PVA Layer onto a Membrane Surface.

    Science.gov (United States)

    Qin, Aiwen; Li, Xiang; Zhao, Xinzhen; Liu, Dapeng; He, Chunju

    2015-04-29

    A highly hydrophilic PVDF membrane was fabricated through chemically binding TiO2 nanoparticles and a poly(vinyl alcohol) (PVA) layer onto a membrane surface simultaneously. The chemical composition of the modified membrane surface was determined by X-ray photoelectron spectroscopy, and the binding performance of TiO2 nanoparticles and the PVA layer was investigated by a rinsing test. The results indicated that the TiO2 nanoparticles were uniformly and strongly tailored onto the membrane surface, while the PVA layer was firmly attached onto the surface of TiO2 nanoparticles and the membrane by adsorption-cross-linking. The possible mechanisms during the modification process and filtration performance, i.e., water permeability and bovine serum albumin (BSA) rejection, were investigated as well. Furthermore, antifouling property was discussed through multicycles of BSA solution filtration tests, where the flux recovery ratio was significantly increased from 20.0% for pristine PVDF membrane to 80.5% for PVDF/TiO2/PVA-modified membrane. This remarkable promotion is mainly ascribed to the improvement of surface hydrophilicity, where the water contact angle of the membrane surface was decreased from 84° for pristine membrane to 24° for PVDF/TiO2/PVA membrane. This study presents a novel and varied strategy for immobilization of nanoparticles and PVA layer on substrate surface, which could be easily adapted for a variety of materials for surface modification.

  8. Rational Design of Highly Potent and Slow-Binding Cytochrome bc1 Inhibitor as Fungicide by Computational Substitution Optimization

    Science.gov (United States)

    Hao, Ge-Fei; Yang, Sheng-Gang; Huang, Wei; Wang, Le; Shen, Yan-Qing; Tu, Wen-Long; Li, Hui; Huang, Li-Shar; Wu, Jia-Wei; Berry, Edward A.; Yang, Guang-Fu

    2015-01-01

    Hit to lead (H2L) optimization is a key step for drug and agrochemical discovery. A critical challenge for H2L optimization is the low efficiency due to the lack of predictive method with high accuracy. We described a new computational method called Computational Substitution Optimization (CSO) that has allowed us to rapidly identify compounds with cytochrome bc1 complex inhibitory activity in the nanomolar and subnanomolar range. The comprehensively optimized candidate has proved to be a slow binding inhibitor of bc1 complex, ~73-fold more potent (Ki = 4.1 nM) than the best commercial fungicide azoxystrobin (AZ; Ki = 297.6 nM) and shows excellent in vivo fungicidal activity against downy mildew and powdery mildew disease. The excellent correlation between experimental and calculated binding free-energy shifts together with further crystallographic analysis confirmed the prediction accuracy of CSO method. To the best of our knowledge, CSO is a new computational approach to substitution-scanning mutagenesis of ligand and could be used as a general strategy of H2L optimisation in drug and agrochemical design.

  9. High throughput functional assays of the variant antigen PfEMP1 reveal a single domain in the 3D7 Plasmodium falciparum genome that binds ICAM1 with high affinity and is targeted by naturally acquired neutralizing antibodies.

    Directory of Open Access Journals (Sweden)

    Andrew V Oleinikov

    2009-04-01

    Full Text Available Plasmodium falciparum-infected erythrocytes bind endothelial receptors to sequester in vascular beds, and binding to ICAM1 has been implicated in cerebral malaria. Binding to ICAM1 may be mediated by the variant surface antigen family PfEMP1: for example, 6 of 21 DBLbetaC2 domains from the IT4 strain PfEMP1 repertoire were shown to bind ICAM1, and the PfEMP1 containing these 6 domains are all classified as Group B or C type. In this study, we surveyed binding of ICAM1 to 16 DBLbetaC2 domains of the 3D7 strain PfEMP1 repertoire, using a high throughput Bioplex assay format. Only one DBL2betaC2 domain from the Group A PfEMP1 PF11_0521 showed strong specific binding. Among these 16 domains, DBL2betaC2(PF11_0521 best preserved the residues previously identified as conserved in ICAM1-binding versus non-binding domains. Our analyses further highlighted the potential role of conserved residues within predominantly non-conserved flexible loops in adhesion, and, therefore, as targets for intervention. Our studies also suggest that the structural/functional DBLbetaC2 domain involved in ICAM1 binding includes about 80 amino acid residues upstream of the previously suggested DBLbetaC2 domain. DBL2betaC2(PF11_0521 binding to ICAM1 was inhibited by immune sera from east Africa but not by control US sera. Neutralizing antibodies were uncommon in children but common in immune adults from east Africa. Inhibition of binding was much more efficient than reversal of binding, indicating a strong interaction between DBL2betaC2(PF11_0521 and ICAM1. Our high throughput approach will significantly accelerate studies of PfEMP1 binding domains and protective antibody responses.

  10. High familial risk for mood disorder is associated with low dorsolateral prefrontal cortex serotonin transporter binding

    DEFF Research Database (Denmark)

    Frokjaer, Vibe G; Vinberg, Maj; Erritzoe, David;

    2009-01-01

    was measured with [(11)C]DASB PET. The volumes of interest included the orbitofrontal cortex, the dorsolateral prefrontal cortex, the ventrolateral prefrontal cortex, anterior cingulate, caudate, putamen, thalamus, and midbrain. We found that individuals at high familial risk for mood disorders had a 35...

  11. GHB receptor targets in the CNS: Focus on high-affinity binding sites

    DEFF Research Database (Denmark)

    Bay, Tina; Eghorn, Laura Friis; Klein, Anders Bue;

    2014-01-01

    γ-Hydroxybutyric acid (GHB) is an endogenous compound in the mammalian brain with both low- and high-affinity receptor targets. GHB is used clinically in the treatment of symptoms of narcolepsy and alcoholism, but also illicitly abused as the recreational drug Fantasy. Major pharmacological effects...

  12. High-throughput sequencing enhanced phage display identifies peptides that bind mycobacteria

    CSIR Research Space (South Africa)

    Ngubane, NAC

    2013-11-01

    Full Text Available tuberculosis. Chest 95: 1193–1197. 7. Davis JL, Huang L, Worodria W, Masur H, Cattamanchi A, et al. (2011) Nucleic acid amplification tests for diagnosis of smear-negative TB in a high HIV-prevalence setting: a prospective cohort study. PLoS One 6: e16321. 8...

  13. The roles of the dielectric constant and the relative level of conduction band of high-k composite with Si in improving the memory performance of charge-trapping memory devices

    Directory of Open Access Journals (Sweden)

    Jianxin Lu

    2014-11-01

    Full Text Available The memory structures Pt/Al2O3/(TiO2x(Al2O31−x/Al2O3/p-Si(nominal composition x = 0.05, 0.50 and 0.70 were fabricated by using rf-magnetron sputtering and atomic layer deposition techniques, in which the dielectric constant and the bottom of the conduction band of the high-k composite (TiO2x(Al2O31−x were adjusted by controlling the partial composition of Al2O3. With the largest dielectric constant and the lowest deviation from the bottom of the conduction band of Si, (TiO20.7(Al2O30.3 memory devices show the largest memory window of 7.54 V, the fast programming/erasing speed and excellent endurance and retention characteristics, which were ascribed to the special structural design, proper combination of dielectric constant and band alignment in the high-k composite (TiO20.7(Al2O30.3.

  14. High affinity and temperature sensitivity of blood oxygen binding in Pangasianodon hypophthalmus due to lack of chloride-hemoglobin allosteric interaction.

    Science.gov (United States)

    Damsgaard, Christian; Phuong, Le My; Huong, Do Thi Thanh; Jensen, Frank B; Wang, Tobias; Bayley, Mark

    2015-06-01

    Air-breathing fishes represent interesting organisms in terms of understanding the physiological changes associated with the terrestrialization of vertebrates, and, further, are of great socio-economic importance for aquaculture in Southeast Asia. To understand how environmental factors, such as high temperature, affect O2 transport in air-breathing fishes, this study assessed the effects of temperature on O2 binding of blood and Hb in the economically important air-breathing fish Pangasianodon hypophthalmus. To determine blood O2 binding properties, blood was drawn from resting cannulated fishes and O2 binding curves made at 25°C and 35°C. To determine the allosteric regulation and thermodynamics of Hb O2 binding, Hb was purified, and O2 equilibria were recorded at five temperatures in the absence and presence of ATP and Cl(-). Whole blood had a high O2 affinity (O2 tension at half saturation P50 = 4.6 mmHg at extracellular pH 7.6 and 25°C), a high temperature sensitivity of O2 binding (apparent heat of oxygenation ΔH(app) = -28.3 kcal/mol), and lacked a Root effect. Further, the data on Hb revealed weak ATP binding and a complete lack of Cl(-) binding to Hb, which, in part, explains the high O2 affinity and high temperature sensitivity of blood O2 binding. This study demonstrates how a potent mechanism for increasing O2 affinity is linked to increased temperature sensitivity of O2 transport and provides a basic framework for a better understanding of how hypoxia-adapted species will react to increasing temperatures.

  15. Positive allosteric modulation of the GHB high-affinity binding site by the GABAA receptor modulator monastrol and the flavonoid catechin.

    Science.gov (United States)

    Eghorn, Laura F; Hoestgaard-Jensen, Kirsten; Kongstad, Kenneth T; Bay, Tina; Higgins, David; Frølund, Bente; Wellendorph, Petrine

    2014-10-05

    γ-Hydroxybutyric acid (GHB) is a metabolite of γ-aminobutyric acid (GABA) and a proposed neurotransmitter in the mammalian brain. We recently identified α4βδ GABAA receptors as possible high-affinity GHB targets. GABAA receptors are highly sensitive to allosteric modulation. Thus to investigate whether GHB high-affinity binding sites are also sensitive to allosteric modulation, we screened both known GABAA receptor ligands and a library of natural compounds in the rat cortical membrane GHB specific high-affinity [3H]NCS-382 binding assay. Two hits were identified: Monastrol, a positive allosteric modulator of GABA function at δ-containing GABAA receptors, and the naturally occurring flavonoid catechin. These compounds increased [3H]NCS-382 binding to 185-272% in high micromolar concentrations. Monastrol and (+)-catechin significantly reduced [3H]NCS-382 dissociation rates and induced conformational changes in the binding site, demonstrating a positive allosteric modulation of radioligand binding. Surprisingly, binding of [3H]GHB and the GHB high-affinity site-specific radioligands [125I]BnOPh-GHB and [3H]HOCPCA was either decreased or only weakly increased, indicating that the observed modulation was critically probe-dependent. Both monastrol and (+)-catechin were agonists at recombinant α4β3δ receptors expressed in Xenopus laevis oocytes. When monastrol and GHB were co-applied no changes were seen compared to the individual responses. In summary, we have identified the compounds monastrol and catechin as the first allosteric modulators of GHB high-affinity binding sites. Despite their relatively weak affinity, these compounds may aid in further characterization of the GHB high-affinity sites that are likely to represent certain GABAA receptors.

  16. Core-shell structured polystyrene/BaTiO3 hybrid nanodielectrics prepared by in situ RAFT polymerization: a route to high dielectric constant and low loss materials with weak frequency dependence.

    Science.gov (United States)

    Yang, Ke; Huang, Xingyi; Xie, Liyuan; Wu, Chao; Jiang, Pingkai; Tanaka, Toshikatsu

    2012-11-23

    A novel route to prepare core-shell structured nanocomposites with excellent dielectric performance is reported. This approach involves the grafting of polystyrene (PS) from the surface of BaTiO(3) by an in situ RAFT polymerization. The core-shell structured PS/BaTiO(3) nanocomposites not only show significantly increased dielectric constant and very low dielectric loss, but also have a weak frequency dependence of dielectric properties over a wide range of frequencies. In addition, the dielectric constant of the nanocomposites can also be easily tuned by varying the thickness of the PS shell. Our method is very promising for preparing high-performance nanocomposites used in energy-storage devices.

  17. POCKET RADIUS OF A HADRON AND STRONG COUPLING CONSTANT AT HIGH ENERGY%强子口袋半径及高能情况下的强耦合常数

    Institute of Scientific and Technical Information of China (English)

    于长丰; 阎坤; 杨新铁

    2004-01-01

    according to the relationship between the strong interaction force range and strong coupling constant among quarks in a hadron, a brief computational method on the pocket radius of a hadron is given in this paper. The strong coupling constant at high energy is also calculated and quite consistent with the experimental value made at at Tevatron -p p Collider.%根据夸克间强相互作用力程随强跑动耦合常数的变化关系,给出了强子口袋半径的一种简明的计算方法.同时计算了高能情况下的强耦合常数,与最近Tevatron对撞机上的实验结果相比较,吻合较好.

  18. Crystal quality and inhibitor binding by aspartic proteinases; preparation of high quality crystals of mouse renin

    Science.gov (United States)

    Badasso, M.; Sibanda, B. L.; Cooper, J. B.; Dealwis, C. G.; Wood, S. P.

    1992-08-01

    Renin from mouse submandibular glands has been highly purified and co-crystallized with a synthetic nonapeptide fragment of rat angiotensionogen in which the scissile Leu-Leu bond has been modified as a hydroxyethylene mimic of the transition state. The strong diffraction from these crystals compared to the native form is discussed in relation to the behaviour of other members of the aspartic proteinase family in crystallisation.

  19. 多 MOSFET 并联均流的高稳定度恒流源研究%Research on the high stable constant current source based on multi-MOSFETs in paralleling connection

    Institute of Scientific and Technical Information of China (English)

    张存凯; 李正坤; 陈乐; 张钟华

    2016-01-01

    In this paper , an approach for a high stable constant current source is presented , which is based on multi-ple power MOSFETs paralleling connection method , the maxim output of current source is up to 30 A, and it is con-tinuously adjustable .The performance of the designed constant current source has been tested , and the measurement results indicate that the stability of constant source is better than 5 ppm when the current is 30 A.The working princi-ple of the designed constant current circuit is analyzed in detail , and the selection of key components is explained clearly.We use the high precision reference voltage source and current series negative feedback based on high -power precision sample resistor to increase the stability of system .In order to realize the high stable current , multiple MOS-FETs in paralleling connection are used as regulator , and systematic analysis on factors affecting MOSFETs in paralle-ling work are presented as well .The constant current source that we designed has a certain reference value for the de-sign of DC current source .%提出了一种基于多MOSFET并联均流的高稳定度大电流恒流源研究方案,输出电流最高可达30 A且连续可调,输出30 A时,30分钟内电流稳定度优于5 ppm。文中详细分析了恒流源基本稳流电路的工作原理,并对电路中关键元器件的选型进行了分析说明。通过高精度基准电压芯片和基于大功率精密电阻采样的电流串联负反馈方法提高系统的稳流精度,通过多只功率MOSFET并联均流工作作为调整管,实现了输出大电流的高稳定度,同时对影响功率MOSFET并联工作的因素进行了系统分析。提出的恒流源方案,对高稳定度直流电源的设计具有一定的参考价值。

  20. Irreversible blockade of the high and low affinity ( sup 3 H) naloxone binding sites by C-6 derivatives of morphinane-6-ones

    Energy Technology Data Exchange (ETDEWEB)

    Krizsan, D. (EGIS Pharmaceutical Works, Budapest (Hungary)); Varga, E.; Benyhe, S.; Szucs, M.; Borsodi, A. (Biological Research Center of the Hungarian Academy of Sciences, Szeged (Hungary)); Hosztafi, S. (Alkaloida Chemical Works, Tiszavasvari (Hungary))

    1991-01-01

    C-6 derivatives-hydrazones, phenylhydrazones, dinitrophenylhydrazones, oximes and semicarbazones - of morphinane-6-ones were synthesized and their binding characteristics were studied on rat brain membranes. The dihydromorphinone and oxymorphone derivatives compete for the ({sup 3}H)naloxone binding sites with high affinity, while the dihydrocodeinone and oxycodone derivatives are less potent. The affinity of the new compounds is decreased for the delta sites as compared to the parent ligands. The ligands bearing bulky substituents also bind with low affinity to the kappa sites. The modification decreased the Na{sup +}-index of compounds indicating their mixed agonist-antagonist character. The dihydromorphinone derivatives are all capable to block irreversibly the high affinity binding site of ({sup 3}H)naloxone, whereas the dihydrocodeinone derivatives block irreversibly the low affinity site. A possible mechanism for the inhibition is suggested.

  1. A maize gene encoding an NADPH binding enzyme highly homologous to isoflavone reductases is activated in response to sulfur starvation.

    Science.gov (United States)

    Petrucco, S; Bolchi, A; Foroni, C; Percudani, R; Rossi, G L; Ottonello, S

    1996-01-01

    we isolated a novel gene that is selectively induced both in roots and shoots in response to sulfur starvation. This gene encodes a cytosolic, monomeric protein of 33 kD that selectively binds NADPH. The predicted polypeptide is highly homologous ( > 70%) to leguminous isoflavone reductases (IFRs), but the maize protein (IRL for isoflavone reductase-like) belongs to a novel family of proteins present in a variety of plants. Anti-IRL antibodies specifically recognize IFR polypeptides, yet the maize protein is unable to use various isoflavonoids as substrates. IRL expression is correlated closely to glutathione availability: it is persistently induced in seedlings whose glutathione content is about fourfold lower than controls, and it is down-regulated rapidly when control levels of glutathione are restored. This glutathione-dependent regulation indicates that maize IRL may play a crucial role in the establishment of a thiol-independent response to oxidative stress under glutathione shortage conditions.

  2. Differential refractometric determination of binding of sodium dodecyl sulfate to protein using high-performance gel chromatography.

    Science.gov (United States)

    Rao, P F; Takagi, T

    1988-10-01

    When sodium dodecyl sulfate (SDS) is added to a high-performance gel chromatographic column equilibrated with a buffer solution containing SDS at a level above the critical micelle concentration, the surplus SDS migrates as micelles giving a sharp peak. The presence of an unfolded protein in the sample solution gives a polypeptide peak in advance of the SDS micelle peak. As the result of SDS binding to the polypeptide, the SDS micelle peak is attenuated in comparison to that in the absence of protein. Thus the amount of SDS bound to the polypeptide can be determined accurately and simply from the decrease in the area of the SDS micelle peak. This approach is particularly useful for precise determination of bound SDS, which is pertinent to understanding the state of the protein polypeptide-SDS complex under the conditions of SDS-polyacrylamide gel electrophoresis.

  3. Novel cyclic gamma-hydroxybutyrate (GHB) analogs with high affinity and stereoselectivity of binding to GHB sites in rat brain

    DEFF Research Database (Denmark)

    Wellendorph, Petrine; Høg, Signe; Greenwood, Jeremy R

    2005-01-01

    acid [(RS)-HOCHCA] and 3-hydroxycyclopent-1-enecarboxylic acid [(RS)-HOCPCA], were found to be high-affinity GHB ligands, with IC50 values in the nanomolar range, and had 9 and 27 times, respectively, higher affinity than GHB. The stereo-selectively synthesized R,R-isomer of the trans-cyclopropyl GHB...... analog, HOCPrCA, proved to have 10-fold higher affinity than its enantiomer. Likewise, the R-enantiomers of HOCHCA and HOCPCA selectively inhibited [3H]NCS-382 binding. The best inhibitor of these, (R)-HOCPCA, has an affinity 39 times higher than GHB and is thus among the best GHB ligands reported......Gamma-hydroxybutyrate (GHB) is a psychotropic compound endogenous to the brain. Despite its potentially great physiological significance, its exact molecular mechanism of action is unknown. GHB is a weak agonist at GABA(B) receptors, but there is also evidence of specific GHB receptor sites...

  4. High-stringency screening of target-binding partners using a microfluidic device

    Energy Technology Data Exchange (ETDEWEB)

    Soh, Hyongsok; Lou, Xinhui; Lagally, Eric

    2015-12-01

    The invention provides a method of screening a library of candidate agents by contacting the library with a target in a reaction mixture under a condition of high stringency, wherein the target includes a tag that responds to a controllable force applied to the tag, and passing the members of the library through a microfluidic device in a manner that exposes the library members to the controllable force, thereby displacing members of the library that are bound to the target relative to their unbound counterparts. Kits and systems for use with the methods of the invention are also provided.

  5. High glucose decreases the expression of ATP-binding cassette transporter G1 in human vascular smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    Jiahong Xue; Zuyi Yuan; Yue Wu; Yan Zhao; Zhaofei Wan

    2008-01-01

    Objective:ATP-binding cassette transporters(ABC) A1 and G1 play an important role in mediating cholesterol efflux and preventing macrophage foam cell formation. In this study, we examined the regulation of ABC transporters by high glucose in human vascular smooth muscle cells(VSMCs), the other precursor of foam cells. Methods:Incubation of human VSMCs with D-ghicose(5 to 30 mM) for 1 to 7 days in the presence or absence of antioxidant and nuclear factor(NF)-kB inhibitors, the expressions of ABCA1 and ABCG1 were analyzed by real time PCR and Western blotting. Results:High glucose decreased ABCG1 mRNA and protein expression in cultured VSMCs, whereas the expression of ABCA1 was not significantly decreased. Down-regulation of ABCG1 mRNA expression by high glucose was abolished by antioxidant N-acetyl-L-cysteine(NAC) and NF-kB inhibitors, BAY 11-7085 and tosyl-phenylalanine chloromethyl-ketone(TPCK). Conclusion:High glucose suppresses the expression of ABCG1 in VSMCs, which is the possible mechanism of VSMC derived foam cell transformation.

  6. Anti-human IgE monoclonal antibodies recognizing epitopes related to the binding sites of high and low affinity IgE receptors.

    Science.gov (United States)

    Takemoto, H; Nishimura, S; Kosada, Y; Hata, S; Takagi, S; Hosoi, S; Ezumi, K; Ide, M; Harada, S

    1994-01-01

    Anti-human IgE monoclonal antibodies (mAbs) were produced and eight clones recognizing epitopes on native IgE were selected. Epitopes were mapped by a competitive inhibition enzyme-linked immunosorbent assay, Western blotting and a multi-pin peptide technology. Four sites (one each in the C epsilon 1, C epsilon 2, C epsilon 2/C epsilon 3 junction and C epsilon 3) were recognized by the mAbs. The relationship between the four epitopes and the binding sites of high and low affinity IgE receptors (Fc epsilon RI and Fc epsilon RII, respectively) was studied using a monovalent Fab fragment of each mAb as a binding inhibitor. The IgE-Fc epsilon RII binding was clearly inhibited by the mAb recognizing the C epsilon 2/C epsilon 3 junction, suggesting that Fc epsilon RII binds to a rather limited area around the C epsilon 2/C epsilon 3 junction. The IgE-Fc epsilon RI binding, on the other hand, was scarcely inhibited by any single mAb. However, the binding was inhibited when the epitope in C epsilon 2 was blocked simultaneously with that at the C epsilon 2/C epsilon 3 junction or with that in C epsilon 3, indicating that these three distinct epitopes are related to the Fc epsilon RI binding sites. When these three epitopes were shown in the stereograph of human IgE, the Fc epsilon RI binding area was spread largely on the groove side between C epsilon 2 and C epsilon 3 domains. These results suggest that Fc epsilon RI acquires the high affinity through multiple bindings.

  7. Binding characteristics of the muscarinic receptor subtype in rabbit pancreas

    Energy Technology Data Exchange (ETDEWEB)

    van Zwam, A.J.; Willems, P.H.; Rodrigues de Miranda, J.F.; de Pont, J.J.; van Ginneken, C.A. (Catholic Univ. of Nijmegen (Netherlands))

    1990-01-01

    The muscarinic receptor in the rabbit pancreas was characterized with the use of the labeled ligand ({sup 3}H)-(-)-quinuclidinyl-benzylate (({sup 3}H)-(-)-QNB). Specific binding of ({sup 3}H)-(-)-QNB to pancreatic acini was found to be reversible and of high affinity, with an equilibrium dissociation constant (KD) of 68 pmol/l and a receptor density (RT) of 170 fmol/mg protein. Agonist binding behaviour was investigated by displacement of ({sup 3}H)-(-)-QNB binding by eight agonists like arecoline, arecadine-propargylester (APE) and carbachol, yielding only low affinity binding sites. The inhibition of ({sup 3}H)-(-)-QNB binding by the selective antagonists pirenzepine, hexahydrosiladifenidol (HHSiD) and (11-(2-(diethyl-amino)-methyl-1-piperidinyl)acetyl)-5,11-dihydro-6H-pyr ido (2,3-b) (1,4) benzodiazepin-6-one (AF-DX 116) confirmed the M3 nature of the rabbit pancreatic receptor.

  8. Construction and experimental testing of the constant-bandwidth constant-temperature anemometer.

    Science.gov (United States)

    Ligeza, P

    2008-09-01

    A classical constant-temperature hot-wire anemometer enables the measurement of fast-changing flow velocity fluctuations, although its transmission bandwidth is a function of measured velocity. This may be a source of significant dynamic errors. Incorporation of an adaptive controller into the constant-temperature system results in hot-wire anemometer operating with a constant transmission bandwidth. The construction together with the results of experimental testing of a constant-bandwidth hot-wire anemometer prototype are presented in this article. During the testing, an approximately constant transmission bandwidth of the anemometer was achieved. The constant-bandwidth hot-wire anemometer can be used in measurements of high-frequency variable flows characterized by a wide range of velocity changes.

  9. High resolution infrared synchrotron study of CH2D81Br: ground state constants and analysis of the ν5, ν6 and ν9 fundamentals

    DEFF Research Database (Denmark)

    Baldacci, A.; Stoppa, P.; Visinoni, R.

    2012-01-01

    The high resolution infrared absorption spectrum of CH2D81Br has been recorded by Fourier transform spectroscopy in the range 550–1075 cm1, with an unapodized resolution of 0.0025 cm-1, employing a synchrotron radiation source. This spectral region is characterized by the ν6 (593.872 cm-1), ν5 (768...

  10. High dielectric constant PrY(x)O(y) sensing films electrolyte-insulator-semiconductor pH-sensor for the detection of urea.

    Science.gov (United States)

    Wu, Min-Hsien; Lee, Cheng-Da; Pan, Tung-Ming

    2009-09-28

    In this paper, we describe the structural and sensing properties of high-k PrY(x)O(y) sensing films deposited on Si substrates through reactive co-sputtering. Secondary ion mass spectrometry and atomic force microscopy were employed to analyze the compositional and morphological features of these films after annealing at various temperatures. The electrolyte-insulator-semiconductor (EIS) device incorporating a PrY(x)O(y) sensing membrane that had been annealed at 800 degrees C exhibited good sensing characteristics, including a high sensitivity (59.07 mV pH(-1) in solutions from pH 2 to 12), a low hysteresis voltage (2.4 mV in the pH loop 7-->4-->7-->10-->7), and a small drift rate (0.62 mV h(-1) in the buffer solution at pH 7). The PrY(x)O(y) EIS device also showed a high selective response towards H(+). This improvement can be attributed to the small number of crystal defects and the large surface roughness. In addition, the enzymatic EIS-based urea biosensor incorporating a high-k PrY(x)O(y) sensing film annealed at 800 degrees C allowed the potentiometric analysis of urea, at concentrations ranging from 1 to 16 mM, with a sensitivity of 9.59 mV mM(-1).

  11. Specificity of Bacillus thuringiensis endotoxins is correlated with the presence of high-affinity binding sites in the brush border membrane of target insect midguts

    Energy Technology Data Exchange (ETDEWEB)

    Hofmann, C.; Vanderbruggen, H.; Hoefte, H.; Van Rie, J.; Jansens, S.; Van Mellaert, H. (J. Plateaustraat, Gent (Belgium))

    1988-11-01

    Binding studies were performed with two {sup 125}I-labeled Bacillus thuringiensis {delta}-endotoxins on brush border membrane vesicles prepared from the larval midgut of the tobacco hornworm Manduca sexta or the cabbage butterfly Pieris brassicae. One {delta}-endotoxin, Bt2-protoxin, is a 130-kDa recombinant crystalline protein from B. thuringiensis subsp. berliner. It kills larvae of both insect species. The active Bt2-toxin is a 60-kDa proteolytic fragment of the Bt2-protoxin. It binds saturably and with high affinity to brush border membrane vesicles from the midgut of both species. The other {delta}-endotoxin, Bt4412-protoxin, is a 136-kDa crystalline protein from B. thuringiensis subsp. thuringiensis, which is highly toxic for P. brassicae, but not for M. sexta larvae. Bt4412-toxin, obtained after proteolytic activation of Bt4412-protoxin, shows high-affinity saturable binding to P. brassicae vesicles but not to M. sexta vesicles. The correlation between toxicity and specific binding is further strengthened by competition studies. Other B. thuringiensis {delta}-endotoxins active against M. sexta compete for binding of {sup 125}I-labeled Bt2-toxin to M. sexta vesicles, whereas toxins active against dipteran or coleopteran larvae do not compete. Bt2-toxin and Bt4412-toxin bind to different sites on P. brassicae vesicles.

  12. Application of Neural network PID Controller in Constant Temperature and Constant Liquid-level System

    Institute of Scientific and Technical Information of China (English)

    Chen,Guochu; Zhang,Lin; Hao,Ninmei; Liu,Xianguang; Wang,Junhong

    2003-01-01

    Guided by the principle of neural network, an intelligent PID controller based on neural network is devised and applied to control of constant temperature and constant liquidlevel system. The experiment results show that this controller has high accuracy and strong robustness and good characters.

  13. Molecular basis for the high-affinity binding and stabilization of firefly luciferase by PTC124

    Energy Technology Data Exchange (ETDEWEB)

    Auld, Douglas S.; Lovell, Scott; Thorne, Natasha; Lea, Wendy A.; Maloney, David J.; Shen, Min; Rai, Ganesha; Battaile, Kevin P.; Thomas, Craig J.; Simeonov, Anton; Hanzlik, Robert P.; Inglese, James (NIH); (Kansas); (HWMRI)

    2010-04-07

    Firefly luciferase (FLuc), an ATP-dependent bioluminescent reporter enzyme, is broadly used in chemical biology and drug discovery assays. PTC124 Ataluren; (3-[5-(2-fluorophenyl)-1,2,4-oxadiazol-3-yl]benzoic acid) discovered in an FLuc-based assay targeting nonsense codon suppression, is an unusually potent FLuc-inhibitor. Paradoxically, PTC124 and related analogs increase cellular FLuc activity levels by posttranslational stabilization. In this study, we show that FLuc inhibition and stabilization is the result of an inhibitory product formed during the FLuc-catalyzed reaction between its natural substrate, ATP, and PTC124. A 2.0 {angstrom} cocrystal structure revealed the inhibitor to be the acyl-AMP mixed-anhydride adduct PTC124-AMP, which was subsequently synthesized and shown to be a high-affinity multisubstrate adduct inhibitor (MAI; KD = 120 pM) of FLuc. Biochemical assays, liquid chromatography/mass spectrometry, and near-attack conformer modeling demonstrate that formation of this novel MAI is absolutely dependent upon the precise positioning and reactivity of a key meta-carboxylate of PTC124 within the FLuc active site. We also demonstrate that the inhibitory activity of PTC124-AMP is relieved by free coenzyme A, a component present at high concentrations in luciferase detection reagents used for cell-based assays. This explains why PTC124 can appear to increase, instead of inhibit, FLuc activity in cell-based reporter gene assays. To our knowledge, this is an unusual example in which the 'off-target' effect of a small molecule is mediated by an MAI mechanism.

  14. Molecular basis for the high-affinity binding and stabilization of firefly luciferase by PTC124

    Science.gov (United States)

    Auld, Douglas S.; Lovell, Scott; Thorne, Natasha; Lea, Wendy A.; Maloney, David J.; Shen, Min; Rai, Ganesha; Battaile, Kevin P.; Thomas, Craig J.; Simeonov, Anton; Hanzlik, Robert P.; Inglese, James

    2010-01-01

    Firefly luciferase (FLuc), an ATP-dependent bioluminescent reporter enzyme, is broadly used in chemical biology and drug discovery assays. PTC124 (Ataluren; (3-[5-(2-fluorophenyl)-1,2,4-oxadiazol-3-yl]benzoic acid) discovered in an FLuc-based assay targeting nonsense codon suppression, is an unusually potent FLuc-inhibitor. Paradoxically, PTC124 and related analogs increase cellular FLuc activity levels by posttranslational stabilization. In this study, we show that FLuc inhibition and stabilization is the result of an inhibitory product formed during the FLuc-catalyzed reaction between its natural substrate, ATP, and PTC124. A 2.0 Å cocrystal structure revealed the inhibitor to be the acyl-AMP mixed-anhydride adduct PTC124-AMP, which was subsequently synthesized and shown to be a high-affinity multisubstrate adduct inhibitor (MAI; KD = 120 pM) of FLuc. Biochemical assays, liquid chromatography/mass spectrometry, and near-attack conformer modeling demonstrate that formation of this novel MAI is absolutely dependent upon the precise positioning and reactivity of a key meta-carboxylate of PTC124 within the FLuc active site. We also demonstrate that the inhibitory activity of PTC124-AMP is relieved by free coenzyme A, a component present at high concentrations in luciferase detection reagents used for cell-based assays. This explains why PTC124 can appear to increase, instead of inhibit, FLuc activity in cell-based reporter gene assays. To our knowledge, this is an unusual example in which the “off-target” effect of a small molecule is mediated by an MAI mechanism. PMID:20194791

  15. High- and low-affinity binding sites for Cd on the bacterial cell walls of Bacillus subtilis and Shewanella oneidensis

    Science.gov (United States)

    Mishra, Bhoopesh; Boyanov, Maxim; Bunker, Bruce A.; Kelly, Shelly D.; Kemner, Kenneth M.; Fein, Jeremy B.

    2010-08-01

    Bulk Cd adsorption isotherm experiments, thermodynamic equilibrium modeling, and Cd K edge EXAFS were used to constrain the mechanisms of proton and Cd adsorption to bacterial cells of the commonly occurring Gram-positive and Gram-negative bacteria, Bacillus subtilis and Shewanella oneidensis, respectively. Potentiometric titrations were used to characterize the functional group reactivity of the S. oneidensis cells, and we model the titration data using the same type of non-electrostatic surface complexation approach as was applied to titrations of B. subtilis suspensions by Fein et al. (2005). Similar to the results for B. subtilis, the S. oneidensis cells exhibit buffering behavior from approximately pH 3-9 that requires the presence of four distinct sites, with p Ka values of 3.3 ± 0.2, 4.8 ± 0.2, 6.7 ± 0.4, and 9.4 ± 0.5, and site concentrations of 8.9(±2.6) × 10 -5, 1.3(±0.2) × 10 -4, 5.9(±3.3) × 10 -5, and 1.1(±0.6) × 10 -4 moles/g bacteria (wet mass), respectively. The bulk Cd isotherm adsorption data for both species, conducted at pH 5.9 as a function of Cd concentration at a fixed biomass concentration, were best modeled by reactions with a Cd:site stoichiometry of 1:1. EXAFS data were collected for both bacterial species as a function of Cd concentration at pH 5.9 and 10 g/L bacteria. The EXAFS results show that the same types of binding sites are responsible for Cd sorption to both bacterial species at all Cd loadings tested (1-200 ppm). Carboxyl sites are responsible for the binding at intermediate Cd loadings. Phosphoryl ligands are more important than carboxyl ligands for Cd binding at high Cd loadings. For the lowest Cd loadings studied here, a sulfhydryl site was found to dominate the bound Cd budgets for both species, in addition to the carboxyl and phosphoryl sites that dominate the higher loadings. The EXAFS results suggest that both Gram-positive and Gram-negative bacterial cell walls have a low concentration of very high

  16. High dielectric constant PrY{sub x}O{sub y} sensing films electrolyte-insulator-semiconductor pH-sensor for the detection of urea

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Min-Hsien [Graduate Institute of Biochemical and Biomedical Engineering, Chang Gung University, Taoyuan 333, Taiwan (China); Lee, Cheng-Da [Department of Electronics Engineering, Chang Gung University, Taoyuan 333, Taiwan (China); Pan, Tung-Ming, E-mail: tmpan@mail.cgu.edu.tw [Department of Electronics Engineering, Chang Gung University, Taoyuan 333, Taiwan (China)

    2009-09-28

    In this paper, we describe the structural and sensing properties of high-k PrY{sub x}O{sub y} sensing films deposited on Si substrates through reactive co-sputtering. Secondary ion mass spectrometry and atomic force microscopy were employed to analyze the compositional and morphological features of these films after annealing at various temperatures. The electrolyte-insulator-semiconductor (EIS) device incorporating a PrY{sub x}O{sub y} sensing membrane that had been annealed at 800 {sup o}C exhibited good sensing characteristics, including a high sensitivity (59.07 mV pH{sup -1} in solutions from pH 2 to 12), a low hysteresis voltage (2.4 mV in the pH loop 7 {yields} 4 {yields} 7 {yields} 10 {yields} 7), and a small drift rate (0.62 mV h{sup -1} in the buffer solution at pH 7). The PrY{sub x}O{sub y} EIS device also showed a high selective response towards H{sup +}. This improvement can be attributed to the small number of crystal defects and the large surface roughness. In addition, the enzymatic EIS-based urea biosensor incorporating a high-k PrY{sub x}O{sub y} sensing film annealed at 800 {sup o}C allowed the potentiometric analysis of urea, at concentrations ranging from 1 to 16 mM, with a sensitivity of 9.59 mV mM{sup -1}.

  17. Antipsychotic drug binding in the substantia nigra: an examination of high metoclopramide binding in the brains of normal, Alzheimer's disease, Huntington's disease, and Multiple Sclerosis patients, and its relation to tardive dyskinesia.

    Science.gov (United States)

    Chen, Sheng; Seeman, Philip; Liu, Fang

    2011-02-01

    This project was done in order to determine why the annual incidence of metoclopramide-associated tardive dyskinesia is much higher than that for the commonly used antipsychotics. To test the hypothesis that metoclopramide tardive dyskinesia may be associated with high concentrations of metoclopramide in the substantia nigra under clinical conditions, the nonspecific binding of tritiated antipsychotics to the dissected melaninized regions of postmortem human substantia nigra was measured. The nonspecific binding at 1 nM [³H]ligand was 7.3, 4.2, 2.6, 0.91 and 0.66 fmoles/mg for [³H]haloperidol, [³H]clozapine, [³H]raclopride, [³H]metoclopramide, and [³H]olanzapine, respectively. After adjusting these values for the known free concentrations of these drugs in plasma or spinal fluid, the amounts that would be bound under clinical conditions would be 231, 113, 15, 11, and 3.4 fmoles/mg for metoclopramide, clozapine, raclopride, haloperidol, and olanzapine, respectively. Using rat striatum as baseline to define antipsychotic binding to nonnigral tissue, the excess amount of binding to the Alzheimer nigral tissue under clinical conditions would be 209, 19, 0, 3.4 and 0.8 fmole/mg for metoclopramide, clozapine, raclopride, haloperidol, and olanzapine, respectively, with a similar pattern for nigral tissues from Huntington and Multiple Sclerosis patients. The high accumulation of metoclopramide is sufficiently high to cause nigral nerve cell membrane damage by metoclopramide's detergent-like action, possibly explaining metoclopramide's toxic ability to elicit early tardive dyskinesia. In addition, the nonspecific binding of metoclopramide was much higher in Alzheimer-diseased substantia nigra, consistent with the fact that older individuals are relatively more vulnerable to metoclopramide tardive dyskinesia.

  18. Determination of the sedimentation constants for total suspended solids and the algal component in a full-scale primary facultative pond operating at high wind velocities under tropical conditions.

    Science.gov (United States)

    Saraiva, L B; Ribeiro Meneses, C G; de Souza Melo, H N; Calado Araújo, A L; Pearson, H

    2005-01-01

    This study evaluated the amount, distribution and sedimentation constant of solids in a full-scale primary facultative pond operating mostly under high wind conditions and the contribution made by the algal biomass. Solids deposition rates were measured using sedimentation traps placed in the inlet and outlet zones of the pond. Most sludge accumulation occurred, not surprisingly, in the inlet zone A1 with a sludge volume of 9072.m3 accumulating over an operating time of approximately 3 years. However, sludge deposition within this zone was uneven and affected by wind action. Mean proportionality constant (K) values for solids sedimentation were 3.02 and 5.70 for depths of 50 cm and 100 cm respectively for A1. In contrast in zone A3, (the outlet zone), reduced K values of 1.38 and 3.22 were obtained for depths of 50 cm and 100 cm respectively. The algal sedimentation constant varied from 0.8 d(-1) in zone A1 to 0.02 d(-1) in A3. These data suggest that in this large facultative pond the wind, blowing predominantly from the direction of the outlets towards the pond inlets, had a greater influence on solids deposition than the bulk hydraulic flow and also kept the pond completely mixed for most of the time.

  19. The highs that bind: school context, social status and marijuana use.

    Science.gov (United States)

    Vogel, Matt; Rees, Chris E; McCuddy, Timothy; Carson, Dena C

    2015-05-01

    Substance use has been closely linked with the structural characteristics of adolescent social networks. Those who drink, smoke, and use drugs typically enjoy an elevated status among their peers. Rates of substance use vary substantially across schools, and indicators of school structure and climate account for at least part of this variation. Emerging research suggests peer-group processes are contingent on school context, but questions remain regarding the school-level mechanisms which condition the influence of network characteristics on substance use. The present study uses multilevel logistic regression models to examine the moderating influence of school connectedness, school drug culture, and global network density on the association between peer network status and marijuana use. The analyses draw on self, peer, and parental data from a sample of 7,548 high-school aged youth nested within 106 schools participating in the National Longitudinal Study of Adolescent to Adult Health (mean age = 15.2; % white = 59 %; male = 45 %). The results indicate that school connectedness significantly reduces the effect of social status on marijuana use. This provides evidence that school-level mechanisms can reduce the instrumentality of marijuana consumption in the status attainment process in adolescence.

  20. Zirconium oxide aerogel for effective enrichment of phosphopeptides with high binding capacity.

    Science.gov (United States)

    Zhang, Liyuan; Xu, Jin; Sun, Liangliang; Ma, Junfeng; Yang, Kaiguang; Liang, Zhen; Zhang, Lihua; Zhang, Yukui

    2011-04-01

    In this study, zirconium oxide (ZrO(2)) aerogel was synthesized via a green sol-gel approach, with zirconium oxychloride, instead of the commonly used alkoxide with high toxicity, as the precursor. With such material, phosphopeptides from the digests of 4 pmol of β-casein with the coexistence of 100 times (mol ratio) BSA could be selectively captured, and identified by MALDI-TOF MS. Due to the large surface area (416.0 m(2) g(-1)) and the mesoporous structure (the average pore size of 10.2 nm) of ZrO(2) aerogel, a 20-fold higher loading capacity for phosphopeptide, YKVPQLEIVPN[pS]AEER (MW 1952.12), was obtained compared to that of commercial ZrO(2) microspheres (341.5 vs. 17.87 mg g(-1)). The metal oxide aerogel was further applied in the enrichment of phosphopeptides from 100 ng nonfat milk, and 17 phosphopeptides were positively identified, with a 1.5-fold improvement in phosphopeptide detection compared with previously reported results. These results demonstrate that ZrO(2) aerogel can be a powerful enrichment material for phosphoproteome study.

  1. Radiosynthesis and Evaluation of [(11)C]3-Hydroxycyclopent-1-enecarboxylic Acid as Potential PET Ligand for the High-Affinity γ-Hydroxybutyric Acid Binding Sites.

    Science.gov (United States)

    Jensen, Claus H; Hansen, Hanne D; Bay, Tina; Vogensen, Stine B; Lehel, Szabolcs; Thiesen, Louise; Bundgaard, Christoffer; Clausen, Rasmus P; Knudsen, Gitte M; Herth, Matthias M; Wellendorph, Petrine; Frølund, Bente

    2017-01-18

    γ-Hydroxybutyric acid (GHB) is an endogenous neuroactive substance and proposed neurotransmitter with affinity for both low- and high-affinity binding sites. A radioligand with high and specific affinity toward the high-affinity GHB binding site would be a unique tool toward a more complete understanding of this population of binding sites. With its high specific affinity and monocarboxylate transporter (MCT1) mediated transport across the blood-brain barrier in pharmacological doses, 3-hydroxycyclopent-1-enecarboxylic acid (HOCPCA) seems like a suitable PET radiotracer candidate. Here, we report the (11)C-labeling and subsequent evaluation of [(11)C]HOCPCA in a domestic pig, as a PET-radioligand for visualization of the high-affinity GHB binding sites in the live pig brain. To investigate the regional binding of HOCPCA in pig brain prior to in vivo PET studies, in vitro quantitative autoradiography on sections of pig brain was performed using [(3)H]HOCPCA. In vivo evaluation of [(11)C]HOCPCA showed no brain uptake, possibly due to a limited uptake of HOCPCA by the MCT1 transporter at tracer doses of [(11)C]HOCPCA.

  2. High statistics determination of the strong coupling constant in Taylor scheme and its OPE Wilson coefficient from lattice QCD with a dynamical charm

    CERN Document Server

    Blossier, B; Brinet, M; De Soto, F; Morenas, V; Pène, O; Petrov, K; Rodríguez-Quintero, J

    2014-01-01

    This paper reports on the determination of $\\alpha_S$ from lattice simulations with 2+1+1 twisted-mass dynamical flavours {\\it via} the computation of the ghost-gluon coupling renormalized in the MOM Taylor scheme. A high-statistics sample of gauge configurations, used to evaluate the coupling from ghost and gluon propagators, allows for the appropriate update of previous results, now performing an improved analysis of data with reduced statistical errors and the systematical uncertainties under a better control.

  3. Computational modeling of the class I low-mass protostar Elias 29 applying optical constants of ices processed by high energy cosmic ray analogs

    CERN Document Server

    Rocha, W R M

    2015-01-01

    We present the study of the effects of high energy cosmic rays (CRs) over the astrophysical ices, observed toward the embedded class I protostar Elias 29, by using computational modeling and laboratory data. Its spectrum was observed with {\\it Infrared Space Observatory - ISO}, covering 2.3 - 190 $\\mu$m. The modeling employed the three-dimensional Monte Carlo radiative transfer code RADMC-3D (Dullemond et al. 2012) and laboratory data of bombarded ice grains by CRs analogs, and unprocessed ices (not bombarded). We are assuming that Elias 29 has a self-irradiated disk with inclination $i =$ 60$^{\\circ}$, surrounded by an envelope with bipolar cavity. The results show that absorption features toward Elias 29, are better reproduced by assuming a combination between unprocessed astrophysical ices at low temperature (H$_2$O, CO, CO$_2$) and bombarded ices (H$_2$O:CO$_2$) by high energy CRs. Evidences of the ice processing around Elias 29 can be observed by the good fitting around 5.5-8.0 $\\mu$m, by polar and apola...

  4. Reactions of a fluorescent ATP analog, 2'-(5-dimethyl-aminonaphthalene-1-sulfonyl) amino-2'-deoxyATP, with E. coli F1-ATPase and its subunits: the roles of the high affinity binding site in the alpha subunit and the low affinity binding site in the beta subunit.

    Science.gov (United States)

    Matsuoka, I; Takeda, K; Futai, M; Tonomura, Y

    1982-11-01

    We performed kinetic studies on the reactions of a fluorescent ATP analog, 2'-(5-dimethyl-aminonaphthalene-1-sulfonyl) amino-2'-deoxyATP (DNS-ATP), with E. coli F1-ATPase (EF1) and its subunits, to clarify the role of each subunit in the ATPase reaction. The following results were obtained. 1. One mol of EF1, which contains nonexchangeable 2 mol ATP and 0.5 mol ADP, binds 3 mol of DNS-ATP. The apparent dissociation constant, in the presence of Mg2+, was 0.23 microM. Upon binding, the fluorescence intensity of DNS-ATP at 520 nm increased exponentially with t1/2 of 35 s, and reached 3.5 times the original fluorescence level. Following the fluorescence increase, DNS-ATP was hydrolyzed, and the fluorescence intensity maintained its enhanced level. 2. The addition of an excess of ATP over the EF1-DNS-nucleotide complex, in the presence of Mg2+, decreased the fluorescence intensity rapidly, indicating the acceleration of DNS-nucleotide release from EF1. ADP and GTP also decreased the fluorescence intensity. 3. DCCD markedly inhibited the accelerating effect of ATP on DNS-nucleotide release from EF1 and the EF1-DNS-ATPase or -ATPase activity in a steady state. On the other hand, DCCD only slightly inhibited the fluorescence increase of DNS-ATP, due to its binding to EF1, and the rate of single cleavage of 1 mol of DNS-ATP per mol of alpha subunit of EF1. 4. In the presence of Mg2+, 0.65-0.82 mol of DNS-ATP binds to 1 mol of the isolated alpha subunit of EF1 with an apparent dissociation constant of 0.06-0.07 microM. Upon binding, the fluorescence intensity of DNS-ATP at 520 nm increased 1.55 fold very rapidly (t1/2 less than 1 s). No hydrolysis of DNS-ATP was observed upon the addition of the isolated alpha subunit. The fluorescence intensity of DNS-ATP was unaffected by the addition of the isolated beta subunit. DNS-ATP was also unhydrolyzed by the isolated beta subunit. 5. EF1-ATPase was reconstituted from alpha, beta, and gamma subunits in the presence of Mg2+ and ATP

  5. Analyzing radioligand binding data.

    Science.gov (United States)

    Motulsky, Harvey; Neubig, Richard

    2002-08-01

    Radioligand binding experiments are easy to perform, and provide useful data in many fields. They can be used to study receptor regulation, discover new drugs by screening for compounds that compete with high affinity for radioligand binding to a particular receptor, investigate receptor localization in different organs or regions using autoradiography, categorize receptor subtypes, and probe mechanisms of receptor signaling, via measurements of agonist binding and its regulation by ions, nucleotides, and other allosteric modulators. This unit reviews the theory of receptor binding and explains how to analyze experimental data. Since binding data are usually best analyzed using nonlinear regression, this unit also explains the principles of curve fitting with nonlinear regression.

  6. RBscore&NBench: a high-level web server for nucleic acid binding residues prediction with a large-scale benchmarking database.

    Science.gov (United States)

    Miao, Zhichao; Westhof, Eric

    2016-07-08

    RBscore&NBench combines a web server, RBscore and a database, NBench. RBscore predicts RNA-/DNA-binding residues in proteins and visualizes the prediction scores and features on protein structures. The scoring scheme of RBscore directly links feature values to nucleic acid binding probabilities and illustrates the nucleic acid binding energy funnel on the protein surface. To avoid dataset, binding site definition and assessment metric biases, we compared RBscore with 18 web servers and 3 stand-alone programs on 41 datasets, which demonstrated the high and stable accuracy of RBscore. A comprehensive comparison led us to develop a benchmark database named NBench. The web server is available on: http://ahsoka.u-strasbg.fr/rbscorenbench/.

  7. Novel radioiodinated {gamma}-hydroxybutyric acid analogues for radiolabeling and Photolinking of high-affinity {gamma}-hydroxybutyric acid binding sites

    DEFF Research Database (Denmark)

    Wellendorph, Petrine; Høg, Signe; Sabbatini, Paola;

    2010-01-01

    ¿-Hydroxybutyric acid (GHB) is a therapeutic drug, a drug of abuse, and an endogenous substance that binds to low- and high-affinity sites in the mammalian brain. To target the specific GHB binding sites, we have developed a (125)I-labeled GHB analog and characterized its binding in rat brain...... homogenate and slices. Our data show that [(125)I]4-hydroxy-4-[4-(2-iodobenzyloxy)phenyl]butanoate ([(125)I]BnOPh-GHB) binds to one site in rat brain cortical membranes with low nanomolar affinity (K(d), 7 nM; B(max), 61 pmol/mg protein). The binding is inhibited by GHB and selected analogs......, but not by ¿-aminobutyric acid. Autoradiography using horizontal slices from rat brain demonstrates the highest density of binding in hippocampus and cortical regions and the lowest density in the cerebellum. Altogether, the findings correlate with the labeling and brain regional distribution of high-affinity GHB sites...

  8. Novel Radioiodinated γ-Hydroxybutyric Acid Analogues for Radiolabeling and Photolinking of High-Affinity γ-Hydroxybutyric Acid Binding Sites

    DEFF Research Database (Denmark)

    Wellendorph, Petrine; Høg, Signe; Sabbatini, Paola;

    2010-01-01

    γ-Hydroxybutyric acid (GHB) is a therapeutic drug, a drug of abuse, and an endogenous substance that binds to low- and high-affinity sites in the mammalian brain. To target the specific GHB binding sites, we have developed a 125I-labeled GHB analog and characterized its binding in rat brain...... homogenate and slices. Our data show that [125I]4-hydroxy-4-[4-(2-iodobenzyloxy)phenyl]butanoate ([125I]BnOPh-GHB) binds to one site in rat brain cortical membranes with low nanomolar affinity (Kd, 7 nM; Bmax, 61 pmol/mg protein). The binding is inhibited by GHB and selected analogs, but not by γ......-aminobutyric acid. Autoradiography using horizontal slices from rat brain demonstrates the highest density of binding in hippocampus and cortical regions and the lowest density in the cerebellum. Altogether, the findings correlate with the labeling and brain regional distribution of high-affinity GHB sites or [3H...

  9. Determination of the gravitational constant G

    Institute of Scientific and Technical Information of China (English)

    HU Zhong-kun; LIU Qi; LUO Jun

    2006-01-01

    A precise knowledge of the Newtonian gravitational constant G has an important role in physics and is of considerable meteorological interest.Although G was the first physical constant to be introduced and measured in the history of science,it is still the least precisely determined of all the fundamental constants of nature.The 2002 CODATA recommended value for G,G=(6.6742±0.0010)×10-11m3·kg-1·s-2,has an uncertainty of 150 parts per million (ppm),much larger than that of all other fundamental constants.Reviewed here is the status of our knowledge of the absolute value of G,methods for determining G,and recent high precision experiments for determining G.

  10. A new generation of starch products as excipient in pharmaceutical tablets .1. Preparation and binding properties of high surface area potato starch products

    NARCIS (Netherlands)

    Wierik, GHPT; ArendsScholte, AW; Eissens, AC; Lerk, CF

    1996-01-01

    A new pharmaceutical excipient with a high binding capacity was prepared from potato starch by enzymatic degradation, followed by suitable dehydration of the precipitated and filtered retrograded starch to produce high specific surface area products. Thermal dehydration methods like drying at room o

  11. Multiple reentrant glass transitions of soft spheres at high densities: monotonicity of the curves of constant relaxation time in jamming phase diagrams depending on temperature over pressure and pressure.

    Science.gov (United States)

    Schmiedeberg, Michael

    2013-05-01

    By using molecular-dynamics simulations, we determine the jamming phase diagrams at high densities for a bidisperse mixture of soft spheres that interact according to repulsive power-law pair potentials. We observe that the relaxation time varies nonmonotonically as a function of density at constant temperature. Therefore, the jamming phase diagrams contain multiple reentrant glass transitions if temperature and density are used as control parameters. However, if we consider a new formulation of the jamming phase diagrams where temperature over pressure and pressure are employed as control parameters, no nonmonotonic behavior is observed.

  12. Quantitative autoradiography of the binding sites for ( sup 125 I) iodoglyburide, a novel high-affinity ligand for ATP-sensitive potassium channels in rat brain

    Energy Technology Data Exchange (ETDEWEB)

    Gehlert, D.R.; Gackenheimer, S.L.; Mais, D.E.; Robertson, D.W. (Eli Lilly and Co., Indianapolis, IN (USA))

    1991-05-01

    We have developed a high specific activity ligand for localization of ATP-sensitive potassium channels in the brain. When brain sections were incubated with ({sup 125}I)iodoglyburide (N-(2-((((cyclohexylamino)carbonyl)amino)sulfonyl)ethyl)-5-{sup 125}I-2- methoxybenzamide), the ligand bound to a single site with a KD of 495 pM and a maximum binding site density of 176 fmol/mg of tissue. Glyburide was the most potent inhibitor of specific ({sup 125}I)iodoglyburide binding to rat forebrain sections whereas iodoglyburide and glipizide were slightly less potent. The binding was also sensitive to ATP which completely inhibited binding at concentrations of 10 mM. Autoradiographic localization of ({sup 125}I)iodoglyburide binding indicated a broad distribution of the ATP-sensitive potassium channel in the brain. The highest levels of binding were seen in the globus pallidus and ventral pallidum followed by the septohippocampal nucleus, anterior pituitary, the CA2 and CA3 region of the hippocampus, ventral pallidum, the molecular layer of the cerebellum and substantia nigra zona reticulata. The hilus and dorsal subiculum of the hippocampus, molecular layer of the dentate gyrus, cerebral cortex, lateral olfactory tract nucleus, olfactory tubercle and the zona incerta contained relatively high levels of binding. A lower level of binding (approximately 3- to 4-fold) was found throughout the remainder of the brain. These results indicate that the ATP-sensitive potassium channel has a broad presence in the rat brain and that a few select brain regions are enriched in this subtype of neuronal potassium channels.

  13. High Resolution Structures of Periplasmic Glucose-binding Protein of Pseudomonas putida CSV86 Reveal Structural Basis of Its Substrate Specificity.

    Science.gov (United States)

    Pandey, Suman; Modak, Arnab; Phale, Prashant S; Bhaumik, Prasenjit

    2016-04-01

    Periplasmic substrate-binding proteins (SBPs) bind to the specific ligand with high affinity and mediate their transport into the cytoplasm via the cognate inner membrane ATP-binding cassette proteins. Because of low sequence identities, understanding the structural basis of substrate recognition by SBPs has remained very challenging. There are several structures available for the ligand-bound sugar SBPs, but very few unliganded structures are reported. No structural data are available for sugar SBPs fromPseudomonassp. to date. This study reports the first high resolution crystal structures of periplasmic glucose-binding protein fromPseudomonas putidaCSV86 (ppGBP) in unliganded form (2.5 Å) and complexed with glucose (1.25 Å) and galactose (1.8 Å). Asymmetric domain closure of ppGBP was observed upon substrate binding. The ppGBP was found to have an affinity of ∼ 0.3 μmfor glucose. The structural analysis showed that the sugars are bound to the protein mainly by hydrogen bonds, and the loss of two strong hydrogen bonds between ppGBP and galactose compared with glucose may be responsible for lowering its affinity toward galactose. The higher stability of ppGBP-glucose complex was also indicated by an 8 °C increase in the melting temperature compared with unliganded form and ppGBP-galactose complex. ppGBP binds to monosaccharide, but the structural features revealed it to have an oligosaccharide-binding protein fold, indicating that during evolution the sugar binding pocket may have undergone structural modulation to accommodate monosaccharide only.

  14. Rational design, synthesis and biological evaluations of amino-noscapine: a high affinity tubulin-binding noscapinoid

    Science.gov (United States)

    Naik, Pradeep K.; Chatterji, Biswa Prasun; Vangapandu, Surya N.; Aneja, Ritu; Chandra, Ramesh; Kanteveri, Srinivas; Joshi, Harish C.

    2011-05-01

    Noscapine and its derivatives are important microtubule-interfering agents shown to have potent anti-tumor activity. The binding free energies (Δ G bind) of noscapinoids computed using linear interaction energy (LIE) method with a surface generalized Born (SGB) continuum solvation model were in agreement with the experimental Δ G bind with average root mean square error of 0.082 kcal/mol. This LIE-SGB model guided us in designing a novel derivative of noscapine, amino-noscapine [(S)-3-((R)-9-amino-4-methoxy-6-methyl-5,6,7,8-tetrahydro [1, 3] dioxolo[4,5- g]isoquinolin-5-yl)-6,7-dimethoxy isobenzo-furan-1(3 H)-one] that has higher tubulin binding activity (predicted Δ G bind = -6.438 kcal/mol and experimental Δ G bind = -6.628 kcal/mol) than noscapine, but does not significantly change the total extent of the tubulin subunit/polymer ratio. The modes of interaction of amino-noscapine with the binding pocket of tubulin involved three hydrogen bonds and are distinct compared to noscapine which involved only one hydrogen bond. Also the patterns of non-bonded interactions are albeit different between both the lignads. The `blind docking' approach (docking of ligand with different binding sites of a protein and their evaluations) as well as the reasonable accuracy of calculating Δ G bind using LIE-SGB model constitutes the first evidence that this class of compounds binds to tubulin at a site overlapping with colchicine-binding site or close to it. Our results revealed that amino-noscapine has better anti-tumor activity than noscapine.

  15. Shark Attack: high affinity binding proteins derived from shark vNAR domains by stepwise in vitro affinity maturation.

    Science.gov (United States)

    Zielonka, Stefan; Weber, Niklas; Becker, Stefan; Doerner, Achim; Christmann, Andreas; Christmann, Christine; Uth, Christina; Fritz, Janine; Schäfer, Elena; Steinmann, Björn; Empting, Martin; Ockelmann, Pia; Lierz, Michael; Kolmar, Harald

    2014-12-10

    A novel method for stepwise in vitro affinity maturation of antigen-specific shark vNAR domains is described that exclusively relies on semi-synthetic repertoires derived from non-immunized sharks. Target-specific molecules were selected from a CDR3-randomized bamboo shark (Chiloscyllium plagiosum) vNAR library using yeast surface display as platform technology. Various antigen-binding vNAR domains were easily isolated by screening against several therapeutically relevant antigens, including the epithelial cell adhesion molecule (EpCAM), the Ephrin type-A receptor 2 (EphA2), and the human serine protease HTRA1. Affinity maturation was demonstrated for EpCAM and HTRA1 by diversifying CDR1 of target-enriched populations which allowed for the rapid selection of nanomolar binders. EpCAM-specific vNAR molecules were produced as soluble proteins and more extensively characterized via thermal shift assays and biolayer interferometry. Essentially, we demonstrate that high-affinity binders can be generated in vitro without largely compromising the desirable high thermostability of the vNAR scaffold.

  16. RIM-binding protein links synaptic homeostasis to the stabilization and replenishment of high release probability vesicles.

    Science.gov (United States)

    Müller, Martin; Genç, Özgür; Davis, Graeme W

    2015-03-04

    Here we define activities of RIM-binding protein (RBP) that are essential for baseline neurotransmission and presynaptic homeostatic plasticity. At baseline, rbp mutants have a ∼10-fold decrease in the apparent Ca(2+) sensitivity of release that we attribute to (1) impaired presynaptic Ca(2+) influx, (2) looser coupling of vesicles to Ca(2+) influx, and (3) limited access to the readily releasable vesicle pool (RRP). During homeostatic plasticity, RBP is necessary for the potentiation of Ca(2+) influx and the expansion of the RRP. Remarkably, rbp mutants also reveal a rate-limiting stage required for the replenishment of high release probability (p) vesicles following vesicle depletion. This rate slows ∼4-fold at baseline and nearly 7-fold during homeostatic signaling in rbp. These effects are independent of altered Ca(2+) influx and RRP size. We propose that RBP stabilizes synaptic efficacy and homeostatic plasticity through coordinated control of presynaptic Ca(2+) influx and the dynamics of a high-p vesicle pool.

  17. Iron-nucleated folding of a metalloprotein in high urea: resolution of metal binding and protein folding events.

    Science.gov (United States)

    Morleo, Anna; Bonomi, Francesco; Iametti, Stefania; Huang, Victor W; Kurtz, Donald M

    2010-08-10

    Addition of iron salts to chaotrope-denatured aporubredoxin (apoRd) leads to nearly quantitative recovery of its single Fe(SCys)(4) site and native protein structure without significant dilution of the chaotrope. This "high-chaotrope" approach was used to examine iron binding and protein folding events using stopped-flow UV-vis absorption and CD spectroscopies. With a 100-fold molar excess of ferrous iron over denatured apoRd maintained in 5 M urea, the folded holoFe(III)Rd structure was recovered in >90% yield with a t(1/2) of Ser iron ligand variants support formation of an unfolded-Fe(SCys)(3) complex between steps 1 and 2, which we propose is the key nucleation event that pulls together distal regions of the protein chain. These results show that folding of chaotrope-denatured apoRd is iron-nucleated and driven by extraordinarily rapid formation of the Fe(SCys)(4) site from an essentially random coil apoprotein. This high-chaotrope, multispectroscopy approach could clarify folding pathways of other [M(SCys)(3)]- or [M(SCys)(4)]-containing proteins.