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Sample records for affinity dissociation constant

  1. Determination of equilibrium dissociation constants for recombinant antibodies by high-throughput affinity electrophoresis.

    PubMed

    Pan, Yuchen; Sackmann, Eric K; Wypisniak, Karolina; Hornsby, Michael; Datwani, Sammy S; Herr, Amy E

    2016-12-23

    High-quality immunoreagents enhance the performance and reproducibility of immunoassays and, in turn, the quality of both biological and clinical measurements. High quality recombinant immunoreagents are generated using antibody-phage display. One metric of antibody quality - the binding affinity - is quantified through the dissociation constant (KD) of each recombinant antibody and the target antigen. To characterize the KD of recombinant antibodies and target antigen, we introduce affinity electrophoretic mobility shift assays (EMSAs) in a high-throughput format suitable for small volume samples. A microfluidic card comprised of free-standing polyacrylamide gel (fsPAG) separation lanes supports 384 concurrent EMSAs in 30 s using a single power source. Sample is dispensed onto the microfluidic EMSA card by acoustic droplet ejection (ADE), which reduces EMSA variability compared to sample dispensing using manual or pin tools. The KD for each of a six-member fragment antigen-binding fragment library is reported using ~25-fold less sample mass and ~5-fold less time than conventional heterogeneous assays. Given the form factor and performance of this micro- and mesofluidic workflow, we have developed a sample-sparing, high-throughput, solution-phase alternative for biomolecular affinity characterization.

  2. Determination of equilibrium dissociation constants for recombinant antibodies by high-throughput affinity electrophoresis

    PubMed Central

    Pan, Yuchen; Sackmann, Eric K.; Wypisniak, Karolina; Hornsby, Michael; Datwani, Sammy S.; Herr, Amy E.

    2016-01-01

    High-quality immunoreagents enhance the performance and reproducibility of immunoassays and, in turn, the quality of both biological and clinical measurements. High quality recombinant immunoreagents are generated using antibody-phage display. One metric of antibody quality – the binding affinity – is quantified through the dissociation constant (KD) of each recombinant antibody and the target antigen. To characterize the KD of recombinant antibodies and target antigen, we introduce affinity electrophoretic mobility shift assays (EMSAs) in a high-throughput format suitable for small volume samples. A microfluidic card comprised of free-standing polyacrylamide gel (fsPAG) separation lanes supports 384 concurrent EMSAs in 30 s using a single power source. Sample is dispensed onto the microfluidic EMSA card by acoustic droplet ejection (ADE), which reduces EMSA variability compared to sample dispensing using manual or pin tools. The KD for each of a six-member fragment antigen-binding fragment library is reported using ~25-fold less sample mass and ~5-fold less time than conventional heterogeneous assays. Given the form factor and performance of this micro- and mesofluidic workflow, we have developed a sample-sparing, high-throughput, solution-phase alternative for biomolecular affinity characterization. PMID:28008969

  3. Improved accuracy of low affinity protein-ligand equilibrium dissociation constants directly determined by electrospray ionization mass spectrometry.

    PubMed

    Jaquillard, Lucie; Saab, Fabienne; Schoentgen, Françoise; Cadene, Martine

    2012-05-01

    There is continued interest in the determination by ESI-MS of equilibrium dissociation constants (K(D)) that accurately reflect the affinity of a protein-ligand complex in solution. Issues in the measurement of K(D) are compounded in the case of low affinity complexes. Here we present a K(D) measurement method and corresponding mathematical model dealing with both gas-phase dissociation (GPD) and aggregation. To this end, a rational mathematical correction of GPD (f(sat)) is combined with the development of an experimental protocol to deal with gas-phase aggregation. A guide to apply the method to noncovalent protein-ligand systems according to their kinetic behavior is provided. The approach is validated by comparing the K(D) values determined by this method with in-solution K(D) literature values. The influence of the type of molecular interactions and instrumental setup on f(sat) is examined as a first step towards a fine dissection of factors affecting GPD. The method can be reliably applied to a wide array of low affinity systems without the need for a reference ligand or protein.

  4. Pinealectomy increases ouabain high-affinity binding sites and dissociation constant in rat cerebral cortex.

    PubMed

    Acuña Castroviejo, D; del Aguila, C M; Fernández, B; Gomar, M D; Castillo, J L

    1991-06-24

    The effect of the pineal gland on the ouabain high-affinity binding sites (Kd = 3.1 +/- 0.4 nM, Bmax = 246.4 +/- 18.4 fmol/mg protein) in rat cerebral cortex was studied. Pinealectomy increased Bmax (940.7 +/- 42.8 fmol/mg protein) and Kd (7.6 +/- 1.5 nM) while melatonin injection (100 micrograms/kg b.wt.) counteracted these effects, restoring kinetic parameters (Kd = 1.9 +/- 0.05 nM; Bmax = 262.2 +/- 29.6 fmol/mg prot) to control values. Melatonin activity on ouabain binding in vitro did not depend upon a direct effect on the binding sites themselves. However, in competition experiments, melatonin increased binding affinity of ouabain as shown by the decreased IC50 values.

  5. Automatic gesture analysis using constant affine velocity.

    PubMed

    Cifuentes, Jenny; Boulanger, Pierre; Pham, Minh Tu; Moreau, Richard; Prieto, Flavio

    2014-01-01

    Hand human gesture recognition has been an important research topic widely studied around the world, as this field offers the ability to identify, recognize, and analyze human gestures in order to control devices or to interact with computer interfaces. In particular, in medical training, this approach is an important tool that can be used to obtain an objective evaluation of a procedure performance. In this paper, some obstetrical gestures, acquired by a forceps, were studied with the hypothesis that, as the scribbling and drawing movements, they obey the one-sixth power law, an empirical relationship which connects path curvature, torsion, and euclidean velocity. Our results show that obstetrical gestures have a constant affine velocity, which is different for each type of gesture and based on this idea this quantity is proposed as an appropriate classification feature in the hand human gesture recognition field.

  6. Determination of rate constants and equilibrium constants for solution-phase drug-protein interactions by ultrafast affinity extraction.

    PubMed

    Zheng, Xiwei; Li, Zhao; Podariu, Maria I; Hage, David S

    2014-07-01

    A method was created on the basis of ultrafast affinity extraction to determine both the dissociation rate constants and equilibrium constants for drug-protein interactions in solution. Human serum albumin (HSA), an important binding agent for many drugs in blood, was used as both a model soluble protein and as an immobilized binding agent in affinity microcolumns for the analysis of free drug fractions. Several drugs were examined that are known to bind to HSA. Various conditions to optimize in the use of ultrafast affinity extraction for equilibrium and kinetic studies were considered, and several approaches for these measurements were examined. The dissociation rate constants obtained for soluble HSA with each drug gave good agreement with previous rate constants reported for the same drugs or other solutes with comparable affinities for HSA. The equilibrium constants that were determined also showed good agreement with the literature. The results demonstrated that ultrafast affinity extraction could be used as a rapid approach to provide information on both the kinetics and thermodynamics of a drug-protein interaction in solution. This approach could be extended to other systems and should be valuable for high-throughput drug screening or biointeraction studies.

  7. Determination of Rate Constants and Equilibrium Constants for Solution-Phase Drug–Protein Interactions by Ultrafast Affinity Extraction

    PubMed Central

    2015-01-01

    A method was created on the basis of ultrafast affinity extraction to determine both the dissociation rate constants and equilibrium constants for drug–protein interactions in solution. Human serum albumin (HSA), an important binding agent for many drugs in blood, was used as both a model soluble protein and as an immobilized binding agent in affinity microcolumns for the analysis of free drug fractions. Several drugs were examined that are known to bind to HSA. Various conditions to optimize in the use of ultrafast affinity extraction for equilibrium and kinetic studies were considered, and several approaches for these measurements were examined. The dissociation rate constants obtained for soluble HSA with each drug gave good agreement with previous rate constants reported for the same drugs or other solutes with comparable affinities for HSA. The equilibrium constants that were determined also showed good agreement with the literature. The results demonstrated that ultrafast affinity extraction could be used as a rapid approach to provide information on both the kinetics and thermodynamics of a drug–protein interaction in solution. This approach could be extended to other systems and should be valuable for high-throughput drug screening or biointeraction studies. PMID:24911267

  8. Measurement by SPR of very low dissociation rates: oxidation-mediated loss of biotin-streptavidin affinity.

    PubMed

    Rebhan, Mario A E; Brunschweiger, Andreas; Hall, Jonathan

    2013-11-04

    Long-term relationship: biotin labels on RNAs, and possibly other biomacromolecules, are easily oxidized causing a dramatic loss of affinity for streptavidin and adversely affecting the measurement of high-affinity interactions. A new SPR method has been developed for measuring the very low rate-dissociation constants of biotin- and biotin oxide-conjugated RNAs with streptavidin.

  9. Deriving the therapeutic concentrations for clozapine and haloperidol: the apparent dissociation constant of a neuroleptic at the dopamine D2 or D4 receptor varies with the affinity of the competing radioligand.

    PubMed

    Seeman, P; Van Tol, H H

    1995-10-15

    The apparent dissociation constant, Ki, for a neuroleptic at the dopamine D2 or D4 receptor was consistently higher when competed against [3H]nemonapride than against [3H]spiperone which was in turn higher than that against [3H]raclopride. This finding obtained for all four types of dopamine receptors studied, including the native dopamine D2 receptor in the anterior pituitary tissue, the human D2long receptor, the human D2short receptor and the human D4.4 receptor. Some neuroleptics revealed a difference of over 10-fold between the Ki using [3H]nemonapride and the Ki using [3H]raclopride. The KD values of the three 3H-ligands and the neuroleptic Ki values were lower when using a much lower concentration of tissue, indicating that depletion of ligand presumably accounted for the phenomenon. The Ki values of each neuroleptic were related to the the tissue/buffer partition coefficients of the three 3H-ligands. Extrapolating the neuroleptic Ki value down to a tissue/buffer partition coefficient of unity or zero led to a Ki value for competition versus a water-soluble ligand such as dopamine. Clozapine extrapolated to a Ki value of 1.3 nM. Direct measurement gave a Ki value of 1.6 nM for [3H]clozapine at the dopamine D4 receptor. When competing versus endogenous dopamine, this clozapine value of 1.6 nM would rise to 20 nM for the blockade of 75% of dopamine D4 receptors, matching the observed therapeutic concentration of 18 nM. These data also explain why clozapine occupies 48% of the D2 receptors in patients when measured with [11C]raclopride, but between 0% and 22% when measured with [18F]methylspiperone or [18F]fluoroethylspiperone.

  10. Direct measurement of equilibrium constants for high-affinity hemoglobins.

    PubMed

    Kundu, Suman; Premer, Scott A; Hoy, Julie A; Trent, James T; Hargrove, Mark S

    2003-06-01

    The biological functions of heme proteins are linked to their rate and affinity constants for ligand binding. Kinetic experiments are commonly used to measure equilibrium constants for traditional hemoglobins comprised of pentacoordinate ligand binding sites and simple bimolecular reaction schemes. However, kinetic methods do not always yield reliable equilibrium constants with more complex hemoglobins for which reaction mechanisms are not clearly understood. Furthermore, even where reaction mechanisms are clearly understood, it is very difficult to directly measure equilibrium constants for oxygen and carbon monoxide binding to high-affinity (K(D) < 1 micro M) hemoglobins. This work presents a method for direct measurement of equilibrium constants for high-affinity hemoglobins that utilizes a competition for ligands between the "target" protein and an array of "scavenger" hemoglobins with known affinities. This method is described for oxygen and carbon monoxide binding to two hexacoordinate hemoglobins: rice nonsymbiotic hemoglobin and Synechocystis hemoglobin. Our results demonstrate that although these proteins have different mechanisms for ligand binding, their affinities for oxygen and carbon monoxide are similar. Their large affinity constants for oxygen, 285 and approximately 100 micro M(-1) respectively, indicate that they are not capable of facilitating oxygen transport.

  11. Semiclassical Calculation of Reaction Rate Constants for Homolytical Dissociations

    NASA Technical Reports Server (NTRS)

    Cardelino, Beatriz H.

    2002-01-01

    There is growing interest in extending organometallic chemical vapor deposition (OMCVD) to III-V materials that exhibit large thermal decomposition at their optimum growth temperature, such as indium nitride. The group III nitrides are candidate materials for light-emitting diodes and semiconductor lasers operating into the blue and ultraviolet regions. To overcome decomposition of the deposited compound, the reaction must be conducted at high pressures, which causes problems of uniformity. Microgravity may provide the venue for maintaining conditions of laminar flow under high pressure. Since the selection of optimized parameters becomes crucial when performing experiments in microgravity, efforts are presently geared to the development of computational OMCVD models that will couple the reactor fluid dynamics with its chemical kinetics. In the present study, we developed a method to calculate reaction rate constants for the homolytic dissociation of III-V compounds for modeling OMCVD. The method is validated by comparing calculations with experimental reaction rate constants.

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

    PubMed

    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 (Ka<10(5)M(-1)), and fast dissociation, such as those composed of cyclodextrins and drugs. In this study, a modified peak profiling method based on high 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.

  13. Affinity constants for small molecules from SPR competition experiments.

    PubMed

    de Mol, Nico J

    2010-01-01

    Direct assay of small molecules by SPR in general is troublesome and at least tedious procedures have to be applied. Competition experiments offer an attractive alternative. A small ligand known to bind to the analyte is immobilized on an SPR sensor surface, and the binding of the larger analyte in the presence of compounds under investigation in a concentration range is assayed. The resulting inhibition curves of the equilibrium SPR signal as function of the compound concentration can be analyzed to yield thermodynamic binding constants for the interaction in solution between analyte and the compounds under investigation. An additional advantage of this method is that series of compounds can be analyzed using the same sensor surface, so there is no immobilization needed for each compound. An adaptation of the method to analyze interactions with bivalent analytes (e.g., antibodies) is also included. Some observed different affinities in solution compared to that on the SPR surface are discussed.

  14. Bayesian Estimation of the Active Concentration and Affinity Constants Using Surface Plasmon Resonance Technology.

    PubMed

    Feng, Feng; Kepler, Thomas B

    2015-01-01

    Surface plasmon resonance (SPR) has previously been employed to measure the active concentration of analyte in addition to the kinetic rate constants in molecular binding reactions. Those approaches, however, have a few restrictions. In this work, a Bayesian approach is developed to determine both active concentration and affinity constants using SPR technology. With the appropriate prior probabilities on the parameters and a derived likelihood function, a Markov Chain Monte Carlo (MCMC) algorithm is applied to compute the posterior probability densities of both the active concentration and kinetic rate constants based on the collected SPR data. Compared with previous approaches, ours exploits information from the duration of the process in its entirety, including both association and dissociation phases, under partial mass transport conditions; do not depend on calibration data; multiple injections of analyte at varying flow rates are not necessary. Finally the method is validated by analyzing both simulated and experimental datasets. A software package implementing our approach is developed with a user-friendly interface and made freely available.

  15. High Throughput pharmacokinetic modeling using computationally predicted parameter values: dissociation constants (TDS)

    EPA Science Inventory

    Estimates of the ionization association and dissociation constant (pKa) are vital to modeling the pharmacokinetic behavior of chemicals in vivo. Methodologies for the prediction of compound sequestration in specific tissues using partition coefficients require a parameter that ch...

  16. The vibrational dependence of dissociative recombination: Rate constants for N{sub 2}{sup +}

    SciTech Connect

    Guberman, Steven L.

    2014-11-28

    Dissociative recombination rate constants are reported with electron temperature dependent uncertainties for the lowest 5 vibrational levels of the N{sub 2}{sup +} ground state. The rate constants are determined from ab initio calculations of potential curves, electronic widths, quantum defects, and cross sections. At 100 K electron temperature, the rate constants overlap with the exception of the third vibrational level. At and above 300 K, the rate constants for excited vibrational levels are significantly smaller than that for the ground level. It is shown that any experimentally determined total rate constant at 300 K electron temperature that is smaller than 2.0 × 10{sup −7} cm{sup 3}/s is likely to be for ions that have a substantially excited vibrational population. Using the vibrational level specific rate constants, the total rate constant is in very good agreement with that for an excited vibrational distribution found in a storage ring experiment. It is also shown that a prior analysis of a laser induced fluorescence experiment is quantitatively flawed due to the need to account for reactions with unknown rate constants. Two prior calculations of the dissociative recombination rate constant are shown to be inconsistent with the cross sections upon which they are based. The rate constants calculated here contribute to the resolution of a 30 year old disagreement between modeled and observed N{sub 2}{sup +} ionospheric densities.

  17. Dissociation constants of parabens and limiting conductances of their ions in water.

    PubMed

    Kroflič, Ana; Apelblat, Alexander; Bešter-Rogač, Marija

    2012-02-02

    Precise measurements of electrical conductivities of methylparaben, ethylparaben, propylparaben, and butylparaben sodium salts in dilute aqueous solutions were performed from 278.15 to 313.15 K in 5 K intervals. Experimental conductivity data were analyzed applying the Quint-Viallard conductivity equations by taking into account the salt hydrolysis in aqueous solutions. These evaluations yield the limiting conductances of paraben anions and the dissociation constants of the investigated parabens in water. From temperature dependence of dissociation constants, the thermodynamic functions associated with the dissociation process were estimated. It was discovered that the contributions of enthalpy and entropy to the Gibbs free energy are quite similar. The Walden products of paraben anions in water are independent of temperature, indicating that the hydrodynamic radii are not significantly affected by temperature.

  18. DISSOCIATION OF ARSENITE-PEPTIDE COMPLEXES: TRIPHASIC NATURE, RATE CONSTANTS, HALF LIVES AND BIOLOGICAL IMPORTANCE

    EPA Science Inventory

    We determined the number and the dissociation rate constants of different complexes formed from arsenite and two peptides containing either one (RV AVGNDYASGYHYGV for peptide 20) or three cysteines (LE AWQGK VEGTEHLYSMK K for peptide 10) via radioactive 73As labeled arsenite and ...

  19. Analysis of agonist dissociation constants as assessed by functional antagonism in guinea pig left atria

    SciTech Connect

    Molenaar, P.; Malta, E.

    1986-04-01

    In electrically driven guinea pig left atria, positive inotropic responses to (-)-isoprenaline and the selective beta 1-adrenoceptor agonist RO363 were obtained in the absence and in the presence of the functional antagonists adenosine, carbachol, gallopamil, nifedipine, and Ro 03-7894. Each of the functional antagonists reduced the maximum response to both agonists and produced nonparallel rightward shifts in the cumulative concentration effect curves. For both agonists, dissociation constants (KA) were calculated using the equation described by Furchgott (1966) for irreversible antagonism. For RO363, which is a partial agonist with high agonist activity, the equations outlined for functional interaction by Mackay (1981) were also employed to calculate KA values. The KA values obtained by each method were compared with the dissociation constants (KD) for the two agonists determined from their ability to displace the radioligand (-)-(/sup 125/I)iodocyanopindolol from beta 1-adrenoceptors in guinea pig left atrial membrane preparations. The estimates of KA varied substantially from KD values. The KD values were taken as more accurate estimates of the true values for the dissociation constants because a high degree of correlation exists between pKD and pD2 values for a number of other beta-adrenoceptor agonists that behave as partial agonists and between pKD and pKB values for a number of beta-adrenoceptor antagonists. Thus, it appears that there are serious limitations in the current theory for using functional antagonism as a means of obtaining agonist dissociation constants.

  20. Relative Proton Affinities from Kinetic Energy Release Distributions for Dissociation of Proton-Bound Dimers

    SciTech Connect

    Hache, John J.; Laskin, Julia ); Futrell, Jean H. )

    2002-12-19

    Kinetic energy release distributions (KERDs) upon dissociation of proton-bound dimers are utilized along with Finite Heat Bath theory analysis to obtain relative proton affinities of monomeric species composing the dimer. The proposed approach allows accurate measurement of relative proton affinities based on KERD measurements for the compound with unknown thermochemical properties versus a single reference base. It also allows distinguishing the cases when dissociation of proton-bound dimers is associated with reverse activation barrier, for which both our approach and the kinetic method become inapplicable. Results are reported for the n-butanol-n-propanol dimer, for which there is no significant difference in entropy effects for two reactions and for the pyrrolidine-1,2-ethylenediamine dimer, which is characterized by a significant difference in entropy effects for the two competing reactions. Relative protonation affinities of -1.0?0.3 kcal/mol for the n-butanol-n-propanol pair and 0.27?0.10 kcal/mol for the pyrrolidine-1,2-ethylenediamine pair are in good agreement with literature values. Relative reaction entropies were extracted from the branching ratio and KERD measurements. Good correspondence was found between the relative reaction entropies for the n-butanol-n-propanol dimer (D(DS?)=-0.3?1.5 cal/mol K) and the relative protonation entropy for the two monomers (D(DSp)=0). However, the relative reaction entropy for the pyrrolidine-1,2-ethylenediamine dimer is higher than the difference in protonation entropies (D(DS?)=8.2?0.5 cal/mol K vs. D(DSp)=5 cal/mol K).

  1. Determination of Henry's constant, the dissociation constant, and the buffer capacity of the bicarbonate system in ruminal fluid.

    PubMed

    Hille, Katharina T; Hetz, Stefan K; Rosendahl, Julia; Braun, Hannah-Sophie; Pieper, Robert; Stumpff, Friederike

    2016-01-01

    Despite the clinical importance of ruminal acidosis, ruminal buffering continues to be poorly understood. In particular, the constants for the dissociation of H2CO3 and the solubility of CO2 (Henry's constant) have never been stringently determined for ruminal fluid. The pH was measured in parallel directly in the rumen and the reticulum in vivo, and in samples obtained via aspiration from 10 fistulated cows on hay- or concentrate-based diets. The equilibrium constants of the bicarbonate system were measured at 38°C both using the Astrup technique and a newly developed method with titration at 2 levels of partial pressure of CO2 (pCO2; 4.75 and 94.98 kPa), yielding mean values of 0.234 ± 0.005 mmol ∙ L(-1) ∙ kPa(-1) and 6.11 ± 0.02 for Henry's constant and the dissociation constant, respectively (n/n = 31/10). Both reticular pH and the pH of samples measured after removal were more alkalic than those measured in vivo in the rumen (by ΔpH = 0.87 ± 0.04 and 0.26 ± 0.04). The amount of acid or base required to shift the pH of ruminal samples to 6.4 or 5.8 (base excess) differed between the 2 feeding groups. Experimental results are compared with the mathematical predictions of an open 2-buffer Henderson-Hasselbalch equilibrium model. Because pCO2 has pronounced effects on ruminal pH and can decrease rapidly in samples removed from the rumen, introduction of a generally accepted protocol for determining the acid-base status of ruminal fluid with standard levels of pCO2 and measurement of base excess in addition to pH should be considered.

  2. A Unified Kinetics and Equilibrium Experiment: Rate Law, Activation Energy, and Equilibrium Constant for the Dissociation of Ferroin

    ERIC Educational Resources Information Center

    Sattar, Simeen

    2011-01-01

    Tris(1,10-phenanthroline)iron(II) is the basis of a suite of four experiments spanning 5 weeks. Students determine the rate law, activation energy, and equilibrium constant for the dissociation of the complex ion in acid solution and base dissociation constant for phenanthroline. The focus on one chemical system simplifies a daunting set of…

  3. Differences in agonist dissociation constant estimates for 5-HT at 5-HT2-receptors: a problem of acute desensitization?

    PubMed Central

    Leff, P.; Martin, G. R.

    1988-01-01

    1. The agonist dissociation constant for 5-hydroxytryptamine (5-HT) was estimated in the guinea-pig isolated trachea by the method of receptor inactivation. The value obtained (pKA = 6.45) was significantly lower than estimates previously obtained in the rabbit aorta and rat jugular vein, although all three tissues are supposed to contain the same 5-HT2 class of receptor. 2. The antagonist dissociation constant for alpha,alpha-dimethyltryptamine was also estimated in the guinea-pig trachea. The pKB value (5.43) was not significantly different from previous estimates in the rabbit aorta and rat jugular vein, consistent with receptor homogeneity between the three tissues. 3. The effect-time profiles corresponding to individual 5-HT applications were more transient in the guinea-pig trachea than in the rabbit aorta. This difference could be accounted for using a simple model of acute receptor desensitization (Leff, 1986), assuming that the conversion of active agonist-receptor complexes into inactive ones was faster in the guinea-pig trachea than in the rabbit aorta. 4. Computer simulation of the desensitization model showed that the discrepancy of pKA estimates for 5-HT between the rabbit aorta and guinea-pig trachea could also be explained using the same rate constant difference that accounted for the difference in effect-time profiles. This analysis indicated that the estimate made in the trachea was erroneously low, whereas that made in the aorta was concluded to be correct. 5. The apparent association between transience of response and pKA estimates is discussed with particular attention to the reliability of agonist affinity estimates in receptor classification. PMID:3228675

  4. Electron-ion dissociative recombination rate constants relevant to the Titan atmosphere and the Interstellar Medium

    SciTech Connect

    Osborne, David; Lawson, Patrick; Adams, Nigel

    2014-01-21

    Following the arrival of Cassini at Titan in 2004, the Titan atmosphere has been shown to contain large complex polycyclic-aromatic hydrocarbons. Since Cassini has provided a great deal of data, there exists a need for kinetic rate data to help with modeling this atmosphere. One type of kinetic data needed is electron-ion dissociative recombination (e-IDR) rate constants. These data are not readily available for larger compounds, such as naphthalene, or oxygen containing compounds, such as 1,4 dioxane or furan. Here, the rate constants for naphthalene, 1,4 dioxane, and furan have been measured and their temperature dependencies are determined when possible, using the University of Georgia's Variable Temperature Flowing Afterglow. The rate constants are compared with those previously published for other compounds; these show trends which illustrate the effects which multi-rings and oxygen heteroatoms substitutions have upon e-IDR rate constants.

  5. Electron-ion dissociative recombination rate constants relevant to the Titan atmosphere and the Interstellar Medium

    NASA Astrophysics Data System (ADS)

    Osborne, David; Lawson, Patrick; Adams, Nigel

    2014-01-01

    Following the arrival of Cassini at Titan in 2004, the Titan atmosphere has been shown to contain large complex polycyclic-aromatic hydrocarbons. Since Cassini has provided a great deal of data, there exists a need for kinetic rate data to help with modeling this atmosphere. One type of kinetic data needed is electron-ion dissociative recombination (e-IDR) rate constants. These data are not readily available for larger compounds, such as naphthalene, or oxygen containing compounds, such as 1,4 dioxane or furan. Here, the rate constants for naphthalene, 1,4 dioxane, and furan have been measured and their temperature dependencies are determined when possible, using the University of Georgia's Variable Temperature Flowing Afterglow. The rate constants are compared with those previously published for other compounds; these show trends which illustrate the effects which multi-rings and oxygen heteroatoms substitutions have upon e-IDR rate constants.

  6. Determination of acid dissociation constants (pKa) of cephalosporin antibiotics: Computational and experimental approaches.

    PubMed

    Ribeiro, Alyson R; Schmidt, Torsten C

    2017-02-01

    Cefapirin (CEPA) and ceftiofur (CEF) are two examples of widely used veterinarian cephalosporins presenting multiple ionization centers. However, the acid dissociation constants (pKa) of CEF are missing and experimental data about CEPA are rare. The same is true for many cephalosporins, where available data are either incomplete or even wrong. Environmentally relevant biotic and abiotic processes depend primordially on the antibiotic pH-dependent speciation. Consequently, this physicochemical parameter should be reliable, including the correct ionization center identification. In this direction, two experimental techniques, potentiometry and spectrophotometry, along with two well-known pKa predictors, Marvin and ACD/Percepta, were used to study the macro dissociation constants of CEPA and CEF. Additionally, the experimental dissociation constants of 14 cephalosporins available in the literature were revised, compiled and compared with data obtained in silico. Only one value was determined experimentally for CEF (2.68 ± 0.05), which was associated to the carboxylic acid group deprotonation. For CEPA two values were obtained experimentally: 2.74 ± 0.01 for the carboxylic acid deprotonation and 5.13 ± 0.01 for the pyridinium ring deprotonation. In general, experimentally obtained values agree with the in silico predicted data (ACD/Percepta RMSE: 0.552 and Marvin RMSE: 0.706, n = 88). However, for cephalosporins having imine and aminothiazole groups structurally close, Marvin presented problems in pKa predictions. For the biological and environmental fate and effect discussion, it is important to recognize that CEPA and CEF, as well as many other cephalosporins, are present as anionic species in the biologic and environmentally relevant pH values of 6-7.5.

  7. Dissociation constants of KSO4- from 10°-50°C

    USGS Publications Warehouse

    Truesdell, A.H.; Hostetler, P.B.

    1968-01-01

    A cell without liquid junction was used to obtain dissociation constants for the reaction: KSO4− = K+ +SO42−. At 10°, 25°, 38° and 50°C, values for Kdiss KSO4− are, respectively, 0.195, 0.142, 0.117, and 0.095. At 25°C, , and  values for the KSO4− ion are −245.96 and −274.02 kcal mole−1, and S° is +42.3 cal mole−1 deg−1.

  8. Enthalpies and constants of dissociation for D,L-Alanyl-D,L-Serine at 298 K

    NASA Astrophysics Data System (ADS)

    Gridchin, S. N.; Pyreu, D. F.

    2015-01-01

    Protolytic equilibria in aqueous solutions of D,L-alanyl-D,L-serine are studied by means of potentiometry and calorimetry. The dissociation constants and thermal effects of this reaction of the dipeptide are determined at 298.15 K and ionic strengths of 0.1, 0.3, 0.5, and 1.0 (KNO3). The standard thermodynamic characteristics (p K°, Δr G°, Δr H°, and Δr S°) of the studied equilibria are calculated. The final results are compared with the corresponding data on the related compounds.

  9. Acid-base titration curves for acids with very small ratios of successive dissociation constants.

    PubMed

    Campbell, B H; Meites, L

    1974-02-01

    The shapes of the potentiometric acid-base titration curves obtained in the neutralizations of polyfunctional acids or bases for which each successive dissociation constant is smaller than the following one are examined. In the region 0 < < 1 (where is the fraction of the equivalent volume of reagent that has been added) the slope of the titration curve decreases as the number j of acidic or basic sites increases. The difference between the pH-values at = 0.75 and = 0.25 has (1 j)log 9 as the lower limit of its maximum value.

  10. Acid Dissociation Constants of Melamine Derivatives from Density Functional Theory Calculations

    NASA Astrophysics Data System (ADS)

    Jang, Yun Hee; Hwang, Sungu; Chang, Seo Bong; Ku, Jamin; Chung, Doo Soo

    2009-10-01

    Melamine and its hydrolysis products (ammeline, ammelide, and cyanuric acid) recently attracted great attention as major food contaminants. Developing analytical tools to quantify them requires exact knowledge of their acid dissociation constants (pKa values). Herein, we calculated the pKa values of these melamine derivatives in water, using a density functional theory quantum mechanical method [B3LYP/6-311++G(d,p)] in combination with the Poisson-Boltzmann continuum solvation model. The excellent agreement of the calculated values with the experimental ones shows that our method can be used to predict such properties of other food contaminants.

  11. Determination of aqueous acid-dissociation constants of aspartic acid using PCM/DFT method

    NASA Astrophysics Data System (ADS)

    Sang-Aroon, Wichien; Ruangpornvisuti, Vithaya

    Determination of acid-dissociation constants, pKa, of aspartic acid in aqueous solution, using density functional theory calculations combined with the conductor-like polarizable continuum model (CPCM) and with integral-equation-formalism polarizable continuum model (IEFPCM) based on the UAKS and UAHF radii, was carried out. The computed pKa values derived from the CPCM and IEFPCM with UAKS cavity model of bare structures of the B3LYP/6-31+G(d,p)-optimized tetrahydrated structures of aspartic acid species are mostly close to the experimental pKa values.0

  12. Spectrophotometric determination of the dissociation constants of methyl yellow in mixed protic solvents.

    PubMed

    Fan, J; Shen, X; Wang, J

    1999-07-12

    The concentration dissociation constants (pK(a)) of methyl yellow, MY (H(+)In) in mixed aqueous solvents of methanol, ethanol, iso-propanol, tert-butanol have been accurately determined from spectrophotometric measurements at 25 degrees C and a constant ionic strength of 0.1 mol l(-1). It has been shown that in these solvents, the pK(a) values decrease with increasing composition of the organic co-solvent. A linear relationship between pK(a) and the mole fraction (x(2)) of the co-solvent was observed in a limited range of the compositions for each of the solvent systems. The results have been discussed in the light of transfer thermodynamic properties of the species existing in the dissociation equilibrium, solvent basicity and solute-solvent interactions. Furthermore, it was also observed that with the change of the solvents, the absorption spectra of MY shifted apparently and the color transition changed accordingly. The solvent effect on the spectra has been attributed to the isomerization equilibria of MY. A simple application of MY was also shown to the sodium acetate-hydrochloric acid titrations in the mixed solvents.

  13. Using electrophoretic mobility shift assays to measure equilibrium dissociation constants: GAL4-p53 binding DNA as a model system.

    PubMed

    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(D) ) for a protein/DNA interaction using electrophoretic mobility shift assays (EMSAs). An EMSA monitors the migration of DNA through a native gel; the DNA migrates more slowly when bound to a protein. To determine a K(D) the amount of unbound and protein-bound DNA in the gel is measured as the protein concentration increases. By performing this experiment, students will be introduced to making affinity measurements and gain experience in performing quantitative EMSAs. The experiment describes measuring the K(D) for the interaction between the chimeric protein GAL4-p53 and its DNA recognition site; however, the techniques are adaptable to other DNA binding proteins. In addition, the basic experiment described can be easily expanded to include additional inquiry-driven experimentation. © 2012 by The International Union of Biochemistry and Molecular Biology.

  14. Complex unconstrained three-dimensional hand movement and constant equi-affine speed.

    PubMed

    Maoz, Uri; Berthoz, Alain; Flash, Tamar

    2009-02-01

    One long-established simplifying principle behind the large repertoire and high versatility of human hand movements is the two-thirds power law-an empirical law stating a relationship between local geometry and kinematics of human hand trajectories during planar curved movements. It was further generalized not only to various types of human movements, but also to motion perception and prediction, although it was unsuccessful in explaining unconstrained three-dimensional (3D) movements. Recently, movement obeying the power law was proved to be equivalent to moving with constant planar equi-affine speed. Generalizing such motion to 3D space-i.e., to movement at constant spatial equi-affine speed-predicts the emergence of a new power law, whose utility for describing spatial scribbling movements we have previously demonstrated. In this empirical investigation of the new power law, subjects repetitively traced six different 3D geometrical shapes with their hand. We show that the 3D power law explains the data consistently better than both the two-thirds power law and an additional power law that was previously suggested for spatial hand movements. We also found small yet systematic modifications of the power-law's exponents across the various shapes, which further scrutiny suggested to be correlated with global geometric factors of the traced shape. Nevertheless, averaging over all subjects and shapes, the power-law exponents are generally in accordance with constant spatial equi-affine speed. Taken together, our findings provide evidence for the potential role of non-Euclidean geometry in motion planning and control. Moreover, these results seem to imply a relationship between geometry and kinematics that is more complex than the simple local one stipulated by the two-thirds power law and similar models.

  15. Determination of the dissociation constants for Ca2+ and calmodulin from the plasma membrane Ca2+ pump by a lipid probe that senses membrane domain changes.

    PubMed

    Mangialavori, Irene; Ferreira-Gomes, Mariela; Pignataro, María F; Strehler, Emanuel E; Rossi, Juan Pablo F C

    2010-01-01

    The purpose of this work was to obtain information about conformational changes of the plasma membrane Ca(2+)-pump (PMCA) in the membrane region upon interaction with Ca(2+), calmodulin (CaM) and acidic phospholipids. To this end, we have quantified labeling of PMCA with the photoactivatable phosphatidylcholine analog [(125)I]TID-PC/16, measuring the shift of conformation E(2) to the auto-inhibited conformation E(1)I and to the activated E(1)A state, titrating the effect of Ca(2+) under different conditions. Using a similar approach, we also determined the CaM-PMCA dissociation constant. The results indicate that the PMCA possesses a high affinity site for Ca(2+) regardless of the presence or absence of activators. Modulation of pump activity is exerted through the C-terminal domain, which induces an apparent auto-inhibited conformation for Ca(2+) transport but does not modify the affinity for Ca(2+) at the transmembrane domain. The C-terminal domain is affected by CaM and CaM-like treatments driving the auto-inhibited conformation E(1)I to the activated E(1)A conformation and thus modulating the transport of Ca(2+). This is reflected in the different apparent constants for Ca(2+) in the absence of CaM (calculated by Ca(2+)-ATPase activity) that sharply contrast with the lack of variation of the affinity for the Ca(2+) site at equilibrium. This is the first time that equilibrium constants for the dissociation of Ca(2+) and CaM ligands from PMCA complexes are measured through the change of transmembrane conformations of the pump. The data further suggest that the transmembrane domain of the PMCA undergoes major rearrangements resulting in altered lipid accessibility upon Ca(2+) binding and activation.

  16. Determination of acid dissociation constants of warfarin and hydroxywarfarins by capillary electrophoresis.

    PubMed

    Nowak, Paweł; Olechowska, Paulina; Mitoraj, Mariusz; Woźniakiewicz, Michał; Kościelniak, Paweł

    2015-08-10

    In this work the acid dissociation constants--pKa of warfarin and its all important oxidative metabolites have been determined by capillary electrophoresis-based methods. It has resulted in a complete description of two acid-base dissociation equilibria, yet not investigated experimentally for phase I metabolites of warfarin. The capillary electrophoresis (CE) method based on the relation between effective electrophoretic mobilities and pH has proven to be a suitable tool for pKa determination, while the spectrophotometric (CE-DAD) and the internal standard methods (IS-CE), have appeared to be promising alternative approaches. The CE-DAD approach based on the change in absorbance spectra between the acidic and basic forms is a combination between capillary electrophoresis and spectrophotometric titration, and yields very consistent values of pKa1 with CE. The IS-CE, in turn, enables an estimation of pKa1 and pKa2 from only two analytical runs, however, less accurate than CE and CE-DAD. The Debye-Hückel model has been confirmed experimentally as a good predictor of pKa values at various ionic strengths. Therefore, it has been used in determination of thermodynamic pKa1 and pKa2, referring to the zero ionic strength. The results are important from the analytical, pharmacological, and theoretical points of view.

  17. Simultaneous determination of equivalence volumes and acid dissociation constants from potentiometric titration data.

    PubMed

    Papanastasiou, G; Ziogas, I

    1995-06-01

    New iterative methods for analysis of potentiometric titration data of (a) mixtures of weak monoprotic acids with their conjugate bases, (b) solutions of polyprotic (di- and triprotic) acids, and (c) mixtures of two diprotic acids are presented. These methods, using data exclusively resulting from the acidic region of the titration curve permits the accurate determination of the analytical concentration of one or more acids even if the titration is stopped well before the end point of the titration. For the titration of a solution containing a conjugate acid/base pair, the proposed procedure enables the extraction of the initial composition of the mixture, as well as the dissociation constant of the concerned acid. Thus, it is possible by this type of analysis to distinguish whether a weak acid has been contaminated by a strong base and define the extent of the contamination. On the other hand, for the titration of polyprotic acids, the proposed approach enables the extraction of the accurate values of the equivalence volume and the dissociation constants K(i) even when the ionization stages overlap. Finally, for the titration of a mixture of two diprotic acids the proposed procedure enables the determination of the composition of the mixture even if the sum of the concentrations of the acids is not known. This method can be used in the analysis of solutions containing two diastereoisomeric forms of a weak diprotic acid. The test of the proposed procedures by means of ideal and Monte Carlo simulated data revealed that these methods are fairly applicable even when the titration data are considerably obscured by 'noise' or contain an important systematic error. The proposed procedures were also successfully applied to experimental titration data.

  18. Is the dissociative recombination of H3(+) really slow. A new spectroscopic measurement of the rate constant

    SciTech Connect

    Amano, T.

    1988-06-01

    The dissociative rate constant for H3(+) occupies a key role in model calculations of the interstellar chemical evolution. The decay of an infrared absorption signal of H3(+) was measured as a function of time, and the decay curve was analyzed and found to fit very well to the form expected for a recombination decay. The signal decay is attributed to the dissociative recombination with electrons, and the rate constant was determined to be (1./8 + or - 0.2) x 10 to the -7th cu cm/s, which disagrees with the recent value obtained with the flowing afterglow/Langmuir probe technique. 22 references.

  19. A new sensitive method of dissociation constants determination based on the isohydric solutions principle.

    PubMed

    Michałowski, Tadeusz; Pilarski, Bogusław; Asuero, Agustin G; Dobkowska, Agnieszka

    2010-10-15

    The paper provides a new formulation and analytical proposals based on the isohydric solutions concept. It is particularly stated that a mixture formed, according to titrimetric mode, from a weak acid (HX, C(0)mol/L) and a strong acid (HB, Cmol/L) solutions, assumes constant pH, independently on the volumes of the solutions mixed, provided that the relation C(0)=C+C(2)·10(pK(1)) is valid, where pK(1)=-log K(1), K(1) the dissociation constant for HX. The generalized formulation, referred to the isohydric solutions thus obtained, was extended also to more complex acid-base systems. Particularly in the (HX, HB) system, the titration occurs at constant ionic strength (I) value, not resulting from presence of a basal electrolyte. This very advantageous conjunction of the properties provides, among others, a new, very sensitive method for verification of pK(1) value. The new method is particularly useful for weak acids HX characterized by low pK(1) values. The method was tested experimentally on four acid-base systems (HX, HB), in aqueous and mixed-solvent media and compared with the literature data. Some useful (linear and hyperbolic) correlations were stated and applied for validation of pK(1) values. Finally, some practical applications of analytical interest of the isohydricity (pH constancy) principle as one formulated in this paper were enumerated, proving the usefulness of such a property which has its remote roots in the Arrhenius concept.

  20. Experimental-like affinity constants and enantioselectivity estimates from flexible docking.

    PubMed

    Gumede, N J; Singh, P; Sabela, M I; Bisetty, K; Escuder-Gilabert, L; Medina-Hernández, M J; Sagrado, S

    2012-10-22

    Experimental-like affinity constants and enantioselectivity estimates, not predicted so far computationally, were obtained using a novel flexible modeling/docking combined strategy. The S- and R-warfarin-human serum albumin (HSA, site I) complexes were used as an interaction model. The process for a verified estimation includes the following: (i) ionized open chain forming at physiological pH (a recent focus); (ii) conformational search (molecular mechanics and Monte Carlo methods); (iii) rigid protein-flexible ligand docking (GlideXP) generating low energy paired S- and R-poses; (iv) graphical comparison against the X-ray crystal structure (unsatisfactory verification step); (v) quantum polarized ligand docking (insufficient verification step); (vi) induced fit docking (one pose satisfying the verification criterion; selection step); (vii) converting docking scores to affinity and enantioselectivity estimates (log K(S) = 5.43, log K(R) = 5.34, ES = K(S)/K(R) = 1.23) and numerical comparison against equivalent literature data from bioanalytical techniques (validation step); (viii) intermolecular forces explaining ES (hydrogen bonding and π-π interactions).

  1. The universal statistical distributions of the affinity, equilibrium constants, kinetics and specificity in biomolecular recognition.

    PubMed

    Zheng, Xiliang; Wang, Jin

    2015-04-01

    We uncovered the universal statistical laws for the biomolecular recognition/binding process. We quantified the statistical energy landscapes for binding, from which we can characterize the distributions of the binding free energy (affinity), the equilibrium constants, the kinetics and the specificity by exploring the different ligands binding with a particular receptor. The results of the analytical studies are confirmed by the microscopic flexible docking simulations. The distribution of binding affinity is Gaussian around the mean and becomes exponential near the tail. The equilibrium constants of the binding follow a log-normal distribution around the mean and a power law distribution in the tail. The intrinsic specificity for biomolecular recognition measures the degree of discrimination of native versus non-native binding and the optimization of which becomes the maximization of the ratio of the free energy gap between the native state and the average of non-native states versus the roughness measured by the variance of the free energy landscape around its mean. The intrinsic specificity obeys a Gaussian distribution near the mean and an exponential distribution near the tail. Furthermore, the kinetics of binding follows a log-normal distribution near the mean and a power law distribution at the tail. Our study provides new insights into the statistical nature of thermodynamics, kinetics and function from different ligands binding with a specific receptor or equivalently specific ligand binding with different receptors. The elucidation of distributions of the kinetics and free energy has guiding roles in studying biomolecular recognition and function through small-molecule evolution and chemical genetics.

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

    ERIC Educational Resources Information Center

    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;…

  3. Enrichment and Analysis of Nonenzymatically Glycated Peptides: Boronate Affinity Chromatography Coupled with Electron-Transfer Dissociation Mass Spectrometry

    SciTech Connect

    Zhang, Qibin; Tang, Ning; Brock, Jonathan W.; Mottaz, Heather M.; Ames, Jennifer M.; Baynes, John; Smith, Richard D.; Metz, Thomas O.

    2007-06-01

    Non-enzymatic glycation of peptides and proteins by D-glucose has important implications in the pathogenesis of diabetes mellitus, particularly in the development of diabetic complications. However, no effective high-throughput methods exist for identifying proteins containing this low abundance post-translational modification in bottom-up proteomic studies. In this report, phenylboronate affinity chromatography was used in a two-step enrichment scheme to selectively isolate first glycated proteins and then glycated, tryptic peptides from human serum glycated in vitro. Enriched peptides were subsequently analyzed by alternating electron transfer dissociation (ETD) and collision induced dissociation (CID) tandem mass spectrometry. It was observed that ETD fragmentation mode resulted in a significantly higher number of glycated peptide identifications (87.6% of all identified peptides) versus CID mode (17.0% of all identified peptides), when utilizing dual glycation enrichment on both the protein and peptide level. This study illustrates that phenylboronate affinity chromatography coupled with LC-MS/MS with ETD as the fragmentation mode is an efficient approach for analyses of glycated proteins and can have broad applications in studies of diabetes mellitus.

  4. Determination of dissociation constant of the NFκB p50/p65 heterodimer using fluorescence cross-correlation spectroscopy in the living cell

    SciTech Connect

    Tiwari, Manisha; Mikuni, Shintaro; Muto, Hideki; Kinjo, Masataka

    2013-07-05

    Highlights: •We used two-laser-beam FCCS to determine the dissociation constant (K{sub d}) of IPT domain of p50/p65 heterodimer in living cell. •Interaction of p50 and p65 was analyzed in the cytoplasm and nucleus of single living cell. •Binding affinity of p50/p65 heterodimer is higher in cytoplasm than that of nucleus. -- Abstract: Two-laser-beam fluorescence cross-correlation spectroscopy (FCCS) is promising technique that provides quantitative information about the interactions of biomolecules. The p50/p65 heterodimer is the most abundant and well understood of the NFκB dimers in most cells. However, the quantitative value of affinity, namely the K{sub d}, for the heterodimer in living cells is not known yet. To quantify the heterodimerization of the IPT domain of p50/p65 in the living cell, we used two-laser-beam FCCS. The K{sub d} values of mCherry{sub 2}- and EGFP-fused p50 and p65 were determined to be 0.46 μM in the cytoplasm and 1.06 μM in the nucleus of the living cell. These results suggest the different binding affinities of the p50/p65 heterodimer in the cytoplasm and nucleus of the living cell and different complex formation in each region.

  5. The Kinetics of N-Ethylmaleimide Inhibition of a Vacuolar H+-ATPase and Determination of Nucleotide Dissociation Constants.

    PubMed Central

    Hunt, I. E.; Sanders, D.

    1996-01-01

    All eukaryotic vacuolar (V-type) ATPases share the property of being inhibited by low concentrations (1-2 [mu]M) if N-ethylmaleimide (NEM). This distinguishes them from P-type ATPases, which are inhibited by higher concentrations of NEM (0.1-1 mM), and F-type ATPases, which are virtually resistant to inhibition by NEM. Using tonoplast vesicles from Beta vulgaris we have determined the kinetics of NEM inactivation of the V-type ATPase to be pseudo-first order. The concentration dependence of the reaction indicates interaction with a single class of inhibitory site with a rate constant of 4.1 x 104 M-1 min-1. Nucleotides protect against inactivation with an efficacy that agrees with their capacity to act as enzyme substrates. The dissociation constant for MgATP has been determined from protection experiments to be 0.44 mM, which is close to the observed Km for hydrolysis (0.39 mM). Likewise, the dissociation constant for protection by MgADP (127 [mu]M) is close to its inhibition constant as a competitive inhibitor (110 [mu]M). Taken together, these findings suggest that NEM inactivation is associated with nucleotide protectable exposure of a single cysteine residue on the catalytic subunit and confirm the utility of this residue for the determination of ligand dissociation constants through protection of maleimide inhibition. PMID:12226173

  6. High kinetic stability of Zn(II) coordinated by the tris(histidine) unit of carbonic anhydrase towards solvolytic dissociation studied by affinity capillary electrophoresis.

    PubMed

    Sato, Yosuke; Hoshino, Hitoshi; Iki, Nobuhiko

    2016-08-01

    Solvolytic dissociation rate constants (kd) of bovine carbonic anhydrase II (CA) and its metallovariants (M-CAs, M=Co(II), Ni(II), Cu(II), Zn(II), and Cd(II)) were estimated by a ligand substitution reaction, which was monitored by affinity capillary electrophoresis to selectively detect the undissociated CAs in the reaction mixture. Using EDTA as the competing ligand for Zn-CA, the dissociation followed the unimolecular nucleophilic substitution (SN1) mechanism with kd=1.0×10(-7)s(-1) (pH7.4, 25°C). The corresponding solvolysis half-life (t1/2) was 80days, showing the exceptionally high kinetic stability of t Zn-CA, in contrast to the highly labile [Zn(II)(H2O)6](2+), where the water exchange rate (kex) is high. This behavior is attributed to the tetrahedral coordination geometry supported by the tris(histidine) unit (His3) of CA. In the case of Co-CA, it showed a somewhat larger kd value (5.7×10(-7)s(-1), pH7.4, 25°C) even though it shares the same tetrahedral coordination environment with Zn-CA, suggesting that the d(7) electronic configuration of Co(II) in the transition state of the dissociation is stabilized by the ligand field. Among M-CAs, only Ni-CA showed a bimolecular nucleophilic substitution (SN2) reaction path in its reaction with EDTA, implying that the large coordination number (6) of Ni(II) in Ni-CA allows EDTA to form an EDTA-Ni-CA intermediate. Overall, kd values roughly correlated with kex values among M-CAs, with the kd value of Zn-CA deviating strongly from the trend and highlighting the exceptionally high kinetic stabilization of Zn-CA by the His3 unit.

  7. SARS CoV main proteinase: The monomer-dimer equilibrium dissociation constant.

    PubMed

    Graziano, Vito; McGrath, William J; Yang, Lin; Mangel, Walter F

    2006-12-12

    The SARS coronavirus main proteinase (SARS CoV main proteinase) is required for the replication of the severe acute respiratory syndrome coronavirus (SARS CoV), the virus that causes SARS. One function of the enzyme is to process viral polyproteins. The active form of the SARS CoV main proteinase is a homodimer. In the literature, estimates of the monomer-dimer equilibrium dissociation constant, KD, have varied more than 65,0000-fold, from <1 nM to more than 200 microM. Because of these discrepancies and because compounds that interfere with activation of the enzyme by dimerization may be potential antiviral agents, we investigated the monomer-dimer equilibrium by three different techniques: small-angle X-ray scattering, chemical cross-linking, and enzyme kinetics. Analysis of small-angle X-ray scattering data from a series of measurements at different SARS CoV main proteinase concentrations yielded KD values of 5.8 +/- 0.8 microM (obtained from the entire scattering curve), 6.5 +/- 2.2 microM (obtained from the radii of gyration), and 6.8 +/- 1.5 microM (obtained from the forward scattering). The KD from chemical cross-linking was 12.7 +/- 1.1 microM, and from enzyme kinetics, it was 5.2 +/- 0.4 microM. While each of these three techniques can present different, potential limitations, they all yielded similar KD values.

  8. SARS CoV Main Proteinase: The Monomer-Dimer Equilibrium Dissociation Constant

    SciTech Connect

    Graziano,V.; McGrath, W.; Yang, L.; Mangel, W.

    2006-01-01

    The SARS coronavirus main proteinase (SARS CoV main proteinase) is required for the replication of the severe acute respiratory syndrome coronavirus (SARS CoV), the virus that causes SARS. One function of the enzyme is to process viral polyproteins. The active form of the SARS CoV main proteinase is a homodimer. In the literature, estimates of the monomer-dimer equilibrium dissociation constant, K{sub D}, have varied more than 650000-fold, from <1 nM to more than 200 {mu}M. Because of these discrepancies and because compounds that interfere with activation of the enzyme by dimerization may be potential antiviral agents, we investigated the monomer-dimer equilibrium by three different techniques: small-angle X-ray scattering, chemical cross-linking, and enzyme kinetics. Analysis of small-angle X-ray scattering data from a series of measurements at different SARS CoV main proteinase concentrations yielded K{sub D} values of 5.8 {+-} 0.8 {mu}M (obtained from the entire scattering curve), 6.5 {+-} 2.2 {mu}M (obtained from the radii of gyration), and 6.8 {+-} 1.5 {mu}M (obtained from the forward scattering). The K{sub D} from chemical cross-linking was 12.7 {+-} 1.1 {mu}M, and from enzyme kinetics, it was 5.2 {+-} 0.4 {mu}M. While each of these three techniques can present different, potential limitations, they all yielded similar K{sub D} values.

  9. Determination of the Acid-Base Dissociation Constant of Acid-Degradable Hexamethylenetetramine by Capillary Zone Electrophoresis.

    PubMed

    Takayanagi, Toshio; Shimakami, Natsumi; Kurashina, Masashi; Mizuguchi, Hitoshi; Yabutani, Tomoki

    2016-01-01

    The acid-base equilibrium of hexamethylenetetramine (hexamine) was analyzed with its effective electrophoretic mobility by capillary zone electrophoresis. Although hexamine is degradable in a weakly acidic aqueous solution, and the degraded products of ammonia and formaldehyde can be formed, the effective electrophoretic mobility of hexamine was measured in the pH range between 2.8 and 6.9. An acid-base dissociation equilibrium of the protonated hexamine was analyzed based on the mobility change, and an acid dissociation constant of pKa = 4.93 ± 0.01 (mean ± standard error, ionic strength: 0.020 mol dm(-3)) was determined. The monoprotic acid-base equilibrium of hexamine was confirmed through comparisons of its electrophoretic mobility with the N-ethylquinolinium ion and with the monocationic N-ethyl derivative of hexamine, as well as a slope analysis of the dissociation equilibrium.

  10. Measuring affinity constants of 1450 monoclonal antibodies to peptide targets with a microarray-based label-free assay platform.

    PubMed

    Landry, J P; Ke, Yaohuang; Yu, Guo-Liang; Zhu, X D

    2015-02-01

    Monoclonal antibodies (mAbs) are major reagents for research and clinical diagnosis. For their inherently high specificities to intended antigen targets and thus low toxicity in general, they are pursued as one of the major classes of new drugs. Yet binding properties of most monoclonal antibodies are not well characterized in terms of affinity constants and how they vary with presentations and/or conformational isomers of antigens, buffer compositions, and temperature. We here report a microarray-based label-free assay platform for high-throughput measurements of monoclonal antibody affinity constants to antigens immobilized on solid surfaces. Using this platform we measured affinity constants of over 1410 rabbit monoclonal antibodies and 46 mouse monoclonal antibodies to peptide targets that are immobilized through a terminal cysteine residue to a glass surface. The experimentally measured affinity constants vary from 10 pM to 200 pM with the median value at 66 pM. We compare the results obtained from the microarray-based platform with those from a benchmarking surface-plasmon-resonance-based (SPR) sensor (Biacore 3000).

  11. Determination of acid-base dissociation constants of amino- and guanidinopurine nucleotide analogs and related compounds by capillary zone electrophoresis.

    PubMed

    Solínová, Veronika; Kasicka, Václav; Koval, Dusan; Cesnek, Michal; Holý, Antonín

    2006-03-01

    CZE has been applied for determination of acid-base dissociation constants (pKa) of ionogenic groups of newly synthesized amino- and (amino)guanidinopurine nucleotide analogs, such as acyclic nucleoside phosphonate, acyclic nucleoside phosphonate diesters and other related compounds. These compounds bear characteristic pharmacophores contained in various important biologically active substances, such as cytostatics and antivirals. The pKa values of ionogenic groups of the above compounds were determined by nonlinear regression analysis of the experimentally measured pH dependence of their effective electrophoretic mobilities. The effective mobilities were measured by CZE performed in series of BGEs in a broad pH range (3.50-11.25), at constant ionic strength (25 mM) and temperature (25 degrees C). pKa values were determined for the protonated guanidinyl group in (amino)guanidino 9-alkylpurines and in (amino)guanidinopurine nucleotide analogs, such as acyclic nucleoside phosphonates and acyclic nucleoside phosphonate diesters, for phosphonic acid to the second dissociation degree (-2) in acyclic nucleoside phosphonates of amino and (amino)guanidino 9-alkylpurines, and for protonated nitrogen in position 1 (N1) of purine moiety in acyclic nucleoside phosphonates of amino 9-alkylpurines. Thermodynamic pKa of protonated guanidinyl group was estimated to be in the range of 7.75-10.32, pKa of phosphonic acid to the second dissociation degree achieved values of 6.64-7.46, and pKa of protonated nitrogen in position 1 of purine was in the range of 4.13-4.89, depending on the structure of the analyzed compounds.

  12. Photodetachment of an electron from selenide ion - The electron affinity and spin-orbit coupling constant for SeH.

    NASA Technical Reports Server (NTRS)

    Smyth, K. C.; Brauman, J. I.

    1972-01-01

    The relative cross section for the gas phase photodetachment of an electron from SeH(-) was determined in the wavelength region 428 to 578 nm. An ion cyclotron resonance spectrometer was used to generate, trap, and detect the negative ions, and a 1000-W xenon arc lamp with a grating monochromator was employed as the light source. The cross section exhibited two sharp thresholds, whose positions remained unchanged for the photodetachment of SeD(-). As a result of these thresholds, the electron affinity and the spin-orbit coupling constant were evaluated.

  13. Optimization of Electrospray Ionization by Statistical Design of Experiments and Response Surface Methodology: Protein-Ligand Equilibrium Dissociation Constant Determinations.

    PubMed

    Pedro, Liliana; Van Voorhis, Wesley C; Quinn, Ronald J

    2016-09-01

    Electrospray ionization mass spectrometry (ESI-MS) binding studies between proteins and ligands under native conditions require that instrumental ESI source conditions are optimized if relative solution-phase equilibrium concentrations between the protein-ligand complex and free protein are to be retained. Instrumental ESI source conditions that simultaneously maximize the relative ionization efficiency of the protein-ligand complex over free protein and minimize the protein-ligand complex dissociation during the ESI process and the transfer from atmospheric pressure to vacuum are generally specific for each protein-ligand system and should be established when an accurate equilibrium dissociation constant (KD) is to be determined via titration. In this paper, a straightforward and systematic approach for ESI source optimization is presented. The method uses statistical design of experiments (DOE) in conjunction with response surface methodology (RSM) and is demonstrated for the complexes between Plasmodium vivax guanylate kinase (PvGK) and two ligands: 5'-guanosine monophosphate (GMP) and 5'-guanosine diphosphate (GDP). It was verified that even though the ligands are structurally similar, the most appropriate ESI conditions for KD determination by titration are different for each. Graphical Abstract ᅟ.

  14. Optimization of Electrospray Ionization by Statistical Design of Experiments and Response Surface Methodology: Protein-Ligand Equilibrium Dissociation Constant Determinations

    NASA Astrophysics Data System (ADS)

    Pedro, Liliana; Van Voorhis, Wesley C.; Quinn, Ronald J.

    2016-09-01

    Electrospray ionization mass spectrometry (ESI-MS) binding studies between proteins and ligands under native conditions require that instrumental ESI source conditions are optimized if relative solution-phase equilibrium concentrations between the protein-ligand complex and free protein are to be retained. Instrumental ESI source conditions that simultaneously maximize the relative ionization efficiency of the protein-ligand complex over free protein and minimize the protein-ligand complex dissociation during the ESI process and the transfer from atmospheric pressure to vacuum are generally specific for each protein-ligand system and should be established when an accurate equilibrium dissociation constant (KD) is to be determined via titration. In this paper, a straightforward and systematic approach for ESI source optimization is presented. The method uses statistical design of experiments (DOE) in conjunction with response surface methodology (RSM) and is demonstrated for the complexes between Plasmodium vivax guanylate kinase ( PvGK) and two ligands: 5'-guanosine monophosphate (GMP) and 5'-guanosine diphosphate (GDP). It was verified that even though the ligands are structurally similar, the most appropriate ESI conditions for KD determination by titration are different for each.

  15. Enthalpies and constants of dissociation of several neutral and cationic acids in aqueous and methanol/water solutions at various temperatures.

    PubMed

    Shoghi, Elham; Romero, Lilian; Reta, Mario; Ràfols, Clara; Bosch, Elisabeth

    2009-05-01

    The acidic dissociation enthalpies and constants of anilinium, protonated tris(hydroxymethyl)aminomethane (HTris(+)), benzoic and acetic acids, have been determined at several temperatures in pure water and in methanol/water mixtures by potentiometry and by isothermal titration microcalorimetry (ITC). The pK(a) values determined by both techniques are in accordance when the dissociation process involves large amounts of heat. However, for the neutral acids the ITC technique gave slightly lower pK(a) values than those from potentiometry at the highest temperatures studied due to the small amounts of heat involved in the acidic dissociation. The dissociation enthalpies have been determined directly by calorimetry and the obtained values slightly decrease with the increase of temperature. Therefore, only a rough estimation of the dissociation enthalpies can be obtained from potentiometric pK(a) by means of the Van't Hoff approach.

  16. Resolution of a Challenge for Solvation Modeling: Calculation of Dicarboxylic Acid Dissociation Constants Using Mixed Discrete-Continuum Solvation Models

    SciTech Connect

    Marenich, Aleksandr; Ding, Wendu; Cramer, Christopher J.; Truhlar, Donald G.

    2012-06-07

    First and second dissociation constants (pKa values) of oxalic acid, malonic acid, and adipic acid were computed by using a number of theoretical protocols based on density functional theory and using both continuum solvation models and mixed discrete-continuum solvation models. We show that fully implicit solvation models (in which the entire solvent is represented by a dielectric continuum) fail badly for dicarboxylic acids with mean unsigned errors averaged over six pKa values) of 2.4-9.0 log units, depending on the particular implicit model used. The use of water-solute clusters and accounting for multiple conformations in solution significantly improve the performance of both generalized Born solvation models and models that solve the nonhomogeneous dielectric Poisson equation for bulk electrostatics. The four most successful models have mean unsigned errors of only 0.6-0.8 log units.

  17. Interaction between DNA and Drugs Having Protonable Basic Groups: Characterization through Affinity Constants, Drug Release Kinetics, and Conformational Changes.

    PubMed

    Alarcón, Liliana P; Baena, Yolima; Manzo, Rubén H

    2017-01-04

    This paper reports the in vitro characterization of the interaction between the phosphate groups of DNA and the protonated species of drugs with basic groups through the determination of the affinity constants, the reversibility of the interaction, and the effect on the secondary structure of the macromolecule. Affinity constants of the counterionic condensation DNA-drug were in the order of 10⁶. The negative electrokinetic potential of DNA decreased with the increase of the proportion of loading drugs. The drugs were slowly released from the DNA-drug complexes and had release kinetics consistent with the high degree of counterionic condensation. The circular dichroism profile of DNA was not modified by complexation with atenolol, lidocaine, or timolol, but was significantly altered by the more lipophilic drugs benzydamine and propranolol, revealing modifications in the secondary structure of the DNA. The in vitro characterization of such interactions provides a physicochemical basis that would contribute to identify the effects of this kind of drugs in cellular cultures, as well as side effects observed under their clinical use. Moreover, this methodology could also be projected to the fields of intracellular DNA transfection and the use of DNA as a carrier of active drugs.

  18. SNAP-Tag Technology: A Useful Tool To Determine Affinity Constants and Other Functional Parameters of Novel Antibody Fragments.

    PubMed

    Niesen, Judith; Sack, Markus; Seidel, Melanie; Fendel, Rolf; Barth, Stefan; Fischer, Rainer; Stein, Christoph

    2016-08-17

    Antibody derivatives, such as the single chain fragment variable (scFv), can be developed as diagnostic and therapeutic tools in cancer research, especially in the form of fusion proteins. Such derivatives are easier to produce and modify than monoclonal antibodies (mAbs) and achieve better tissue/tumor penetration. The genetic modification of scFvs is also much more straightforward than the challenging chemical modification of mAbs. Therefore, we constructed two scFvs derived from the approved monoclonal antibodies cetuximab (scFv2112) and panitumumab (scFv1711), both of which are specific for the epidermal growth factor receptor (EGFR), a well-characterized solid tumor antigen. Both scFvs were genetically fused to the SNAP-tag, an engineered version of the human DNA repair enzyme O(6)-alkylguanine DNA alkyltransferase that allows the covalent coupling of benzylguanine (BG)-modified substrates such as fluorescent dyes. The SNAP-tag achieves controllable and irreversible protein modification and is an important tool for experimental studies in vitro and in vivo. The affinity constant of a scFv is a key functional parameter, especially in the context of a fusion protein. Therefore, we developed a method to define the affinity constants of scFv-SNAP fusion proteins by surface plasmon resonance (SPR) spectroscopy. We could confirm that both scFvs retained their functionality after fusion to the SNAP-tag in a variety of procedures and assays, including ELISA, flow cytometry, and confocal microscopy. The experimental procedures described herein, and the new protocol for affinity determination by SPR spectroscopy, are suitable for the preclinical evaluation of diverse antibody formats and derivatives.

  19. Calculation of rate constants for dissociative attachment of low-energy electrons to hydrogen halides HCl, HBr, and HI and their deuterated analogs

    SciTech Connect

    Houfek, Karel; Cizek, Martin; Horacek, Jiri

    2002-12-01

    Calculations of rate constants for the process of dissociative attachment of low-energy electrons to hydrogen halides HCl, HBr, and HI and for the reverse process of associative detachment based on the nonlocal resonance model are reported. The calculated data are of importance for the modeling of plasma processes, environmental chemistry, etc. The calculated dissociative attachment rate constants are found to be in good agreement with existing experimental data. It is shown that at low temperatures the rate constants are very sensitive to small changes of the parameters of the nonlocal resonance model used for the calculation of the rate constants and represent a severe test of the theory. The isotopic effect and its dependence on the temperature is also discussed. The calculations of rate constants for the reverse process of associative detachment are also reported and discussed.

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

    PubMed

    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.

  1. Equilibrium and rate constants, and reaction mechanism of the HF dissociation in the HF(H2O)7 cluster by ab initio rare event simulations.

    PubMed

    Elena, Alin Marin; Meloni, Simone; Ciccotti, Giovanni

    2013-12-12

    We perform restrained hybrid Monte Carlo (MC) simulations to compute the equilibrium constant of the dissociation reaction of HF in HF(H2O)7. We find that the HF is a stronger acid in the cluster than in the bulk, and its acidity is higher at lower T. The latter phenomenon has a vibrational entropic origin, resulting from a counterintuitive balance of intra- and intermolecular terms. We find also a temperature dependence of the reactions mechanism. At low T (≤225 K) the dissociation reaction follows a concerted path, with the H atoms belonging to the relevant hydrogen bond chain moving synchronously. At higher T (300 K), the first two hydrogen atoms move together, forming an intermediate metastable state having the structure of an eigen ion (H9O4(+)), and then the third hydrogen migrates completing the reaction. We also compute the dissociation rate constant, kRP. At very low T (≤75 K) kRP depends strongly on the temperature, whereas it gets almost constant at higher T’s. With respect to the bulk, the HF dissociation in the HF(H2O)7 is about 1 order of magnitude faster. This is due to a lower free energy barrier for the dissociation in the cluster.

  2. Determination of drug-polymer binding constants by affinity capillary electrophoresis for aryl propionic acid derivatives and related compounds.

    PubMed

    Jia, Zhongjiang; Choi, Duk Soon; Chokshi, Hitesh

    2013-03-01

    The binding constants (K(b)s) of 17 aryl propionic acid derivatives (APADs) and related compounds with polyvinylpyrrolidone (PVP K30) and vinylpyrrolidone-vinyl acetate copolymer (Kollidon VA64) in aqueous media were determined by affinity capillary electrophoreses (ACE). The K(b)s of APAD to polymers increase with octanol-water partition coefficients of the compounds. Kollidon VA64 is a stronger binder than PVP K30 to APAD compounds. The K(b)s are greater at pH 4 than at pH 9. Both hydrophobic interaction and hydrogen bonding may be involved. However, hydrophobic interaction appears to be dominant. The ACE method is simple and fast, which could be used to study drug-polymer interaction in aqueous media.

  3. Visualization of CD2 interaction with LFA-3 and determination of the two-dimensional dissociation constant for adhesion receptors in a contact area

    PubMed Central

    1996-01-01

    Many adhesion receptors have high three-dimensional dissociation constants (Kd) for counter-receptors compared to the KdS of receptors for soluble extracellular ligands such as cytokines and hormones. Interaction of the T lymphocyte adhesion receptor CD2 with its counter- receptor, LFA-3, has a high solution-phase Kd (16 microM at 37 degrees C), yet the CD2/LFA-3 interaction serves as an effective adhesion mechanism. We have studied the interaction of CD2 with LFA-3 in the contact area between Jurkat T lymphoblasts and planar phospholipid bilayers containing purified, fluorescently labeled LFA-3. Redistribution and lateral mobility of LFA-3 were measured in contact areas as functions of the initial LFA-3 surface density and of time after contact of the cells with the bilayers. LFA-3 accumulated at sites of contact with a half-time of approximately 15 min, consistent with the previously determined kinetics of adhesion strengthening. The two-dimensional Kd for the CD2/LFA-3 interaction was 21 molecules/microns 2, which is lower than the surface densities of CD2 on T cells and LFA-3 on most target or stimulator cells. Thus, formation of CD2/LFA-3 complexes should be highly favored in physiological interactions. Comparison of the two-dimensional (membrane- bound) and three-dimensional (solution-phase) KdS suggest that cell- cell contact favors CD2/LFA-3 interaction to a greater extent than that predicted by the three-dimensional Kd and the intermembrane distance at the site of contact. LFA-3 molecules in the contact site were capable of lateral diffusion in the plane of the phospholipid bilayer and did not appear to be irreversibly trapped in the contact area, consistent with a rapid off-rate. These data provide insights into the function of low affinity interactions in adhesion. PMID:8636222

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

    SciTech Connect

    Cheng, Chih-Ming; Lee, Yuarn-Jang; Wang, Wei-Ting; Hsu, Chien-Ting; Tsai, Jing-Shin; Wu, Chien-Ming; Ou, Keng-Liang; 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. The New Low Value for the Second Dissociation Constant for H2S: Its History, Its Best Value, and Its Impact on the Teaching of Sulfide Equilibria.

    ERIC Educational Resources Information Center

    Myers, Rollie J.

    1986-01-01

    Discusses the historical development of a new value of the second dissociation constant (K2) for hydrogen sulfide (H2S). Describes the differences between the traditional high values for K2. Suggests modification of teaching about sulfide equilibria in light of the new low value for K2. (TW)

  6. Determination of the acid dissociation constant of the biosurfactant monorhamnolipid in aqueous solution by potentiometric and spectroscopic methods.

    PubMed

    Lebrón-Paler, Ariel; Pemberton, Jeanne E; Becker, Bridget A; Otto, William H; Larive, Cynthia K; Maier, Raina M

    2006-11-15

    The acid dissociation constant in water for a monorhamnolipid mixture extracted from Pseudomonas aeruginosa ATCC 9027 has been determined using potentiometry and two spectroscopic approaches at concentrations below and above the critical micelle concentration (cmc). Potentiometric titrations resulted in pKa values ranging from 4.28 +/- 0.16 to 5.50 +/- 0.06 depending on concentration. 1H NMR spectrochemical titrations at concentrations below the cmc revealed a pKa value of 4.39 +/- 0.06. ATR-FT-IR spectrochemical titrations on solutions well above the cmc gave a pKa value of 4.84 +/- 0.05. The value of 4.28 for the free rhamnolipid molecule for concentrations below the cmc differs markedly from that reported previously. However, the pKa of 5.50 for surface-adsorbed and solution aggregates correlates closely to that previously reported. Differences in these pKa values are rationalized in terms of the pH- and concentration-dependent aggregation behavior of rhamnolipids in aqueous solution.

  7. Detection analysis of surface hydroxyl active sites and simulation calculation of the surface dissociation constants of aqueous diatomite suspensions

    NASA Astrophysics Data System (ADS)

    Ma, Shu-Cui; Wang, Zhi-Gang; Zhang, Ji-Lin; Sun, De-Hui; Liu, Gui-Xia

    2015-02-01

    The surface properties of the diatomite were investigated using nitrogen adsorption/deadsorption isotherms, TG-DSC, FTIR, and XPS, and surface protonation-deprotonation behavior was determined by continuous acid-base potentiometric titration technique. The diatomite sample with porous honeycomb structure has a BET specific surface area of 10.21 m2/g and large numbers of surface hydroxyl functional groups (i.e. tbnd Si-OH, tbnd Fe-OH, and tbnd Al-OH). These surface hydroxyls can be protonated or deprotonated depending on the pH of the suspension. The experimental potentiometric data in two different ionic strength solutions (0.1 and 0.05 mol/L NaCl) were fitted using ProtoFit GUI V2.1 program by applying diffuse double layer model (DLM) with three amphoteric sites and minimizing the sum of squares between a dataset derivative function and a model derivative function. The optimized surface parameters (i.e. surface dissociation constants (log K1, log K2) and surface site concentrations (log C)) of the sample were obtained. Based on the optimized surface parameters, the surface species distribution was calculated using Program-free PHREEQC 3.1.2. Thus, this work reveals considerable new information about surface protonation-deprotonation processes and surface adsorptive behaviors of the diatomite, which helps us to effectively use the cheap and cheerful diatomite clay adsorbent.

  8. Multiwavelength spectrophotometric determination of acid dissociation constants part V: microconstants and tautomeric ratios of diprotic amphoteric drugs.

    PubMed

    Takács-Novák, K; Tam, K Y

    2000-01-01

    The acid-base equilibria of several diprotic amphoteric drugs, namely, niflumic acid, norfloxacin, piroxicam, pyridoxine and 2-methyl-4-oxo-3H-quinazoline-3-acetic acid have been characterized in terms of microconstants and tautomeric ratios. A multiwavelength spectrophotometric (WApH) titration method for determination of acid dissociation constants (pKa values) of ionizable compounds developed previously was applied for this purpose. Microspeciation was investigated by three approaches: (1) selective monitoring of ionizable group by spectrophotometry, (2) deductive method and (3) k(z) method for determination of tautomeric ratio from co-solvent mixtures. The formulation for (3) has been derived and found to invoke fewer assumptions than a reported procedure (K. Takács-Novák, A. Avdeef, K.J Box, B. Podányi, G. Szász, J. Pharm. Biomed. Anal., 12 (1994) 1369-1377). It has been shown that the WApH technique, for such types of ampholytes, is able to deduce the microconstants and tautomeric ratios which are in good agreement with literature data.

  9. Electrostatically-driven fast association and perdeuteration allow detection of transferred cross-relaxation for G protein-coupled receptor ligands with equilibrium dissociation constants in the high-to-low nanomolar range.

    PubMed

    Catoire, Laurent J; Damian, Marjorie; Baaden, Marc; Guittet, Eric; Banères, Jean-Louis

    2011-07-01

    The mechanism of signal transduction mediated by G protein-coupled receptors is a subject of intense research in pharmacological and structural biology. Ligand association to the receptor constitutes a critical event in the activation process. Solution-state NMR can be amenable to high-resolution structure determination of agonist molecules in their receptor-bound state by detecting dipolar interactions in a transferred mode, even with equilibrium dissociation constants below the micromolar range. This is possible in the case of an inherent ultra-fast diffusive association of charged ligands onto a highly charged extracellular surface, and by slowing down the (1)H-(1)H cross-relaxation by perdeuterating the receptor. Here, we demonstrate this for two fatty acid molecules in interaction with the leukotriene BLT2 receptor, for which both ligands display a submicromolar affinity.

  10. Dissociation Rate Constants of Human Fibronectin Binding to Fibronectin-binding Proteins on Living Staphylococcus aureus Isolated from Clinical Patients*

    PubMed Central

    Casillas-Ituarte, Nadia N.; Lower, Brian H.; Lamlertthon, Supaporn; Fowler, Vance G.; Lower, Steven K.

    2012-01-01

    Staphylococcus aureus is part of the indigenous microbiota of humans. Sometimes, S. aureus bacteria enter the bloodstream, where they form infections on implanted cardiovascular devices. A critical, first step in such infections is a bond that forms between fibronectin-binding protein (FnBP) on S. aureus and host proteins, such as fibronectin (Fn), that coat the surface of implants in vivo. In this study, native FnBPs on living S. aureus were shown to form a mechanically strong conformational structure with Fn by atomic force microscopy. The tensile acuity of this bond was probed for 46 bloodstream isolates, each from a patient with a cardiovascular implant. By analyzing the force spectra with the worm-like chain model, we determined that the binding events were consistent with a multivalent, cluster bond consisting of ∼10 or ∼80 proteins in parallel. The dissociation rate constant (koff, s−1) of each multibond complex was determined by measuring strength as a function of the loading rate, normalized by the number of bonds. The bond lifetime (1/koff) was two times longer for bloodstream isolates from patients with an infected device (1.79 or 69.47 s for the 10- or 80-bond clusters, respectively; n = 26 isolates) relative to those from patients with an uninfected device (0.96 or 34.02 s; n = 20 isolates). This distinction could not be explained by different amounts of FnBP, as confirmed by Western blots. Rather, amino acid polymorphisms within the Fn-binding repeats of FnBPA explain, at least partially, the statistically (p < 0.05) longer bond lifetime for isolates associated with an infected cardiovascular device. PMID:22219202

  11. Reliability of dissociation constants and resolution capability of SQUAD(84) and SPECFIT/32 in the regression of multiwavelength spectrophotometric pH-titration data

    NASA Astrophysics Data System (ADS)

    Meloun, Milan; Ferenčíková, Zuzana; Javůrek, Milan

    2012-02-01

    The resolving power of multicomponent spectral analysis and the computation reliability of the stability constants and molar absorptivities determined for five variously protonated anions of physostigmine salicylate by the SQUAD(84) and SPECFIT/32 programs has been examined with the use of simulated and experimental spectra containing overlapping spectral bands. The reliability of the dissociation constants of drug was proven with goodness-of-fit tests and by examining the influence of pre-selected noise level sinst( A) in synthetic spectra regarding the precision s(p K) and also accuracy of the estimated dissociation constants. Precision was examined as the linear regression model s(p K) = β0 + β1 sinst( A). In all cases the intercept β0 was statistically insignificant. When an instrumental error sinst( A) is small and less than 0.5 mAU, the parameters' estimates are nearly the same as the bias Δp K = p Ka,calc - p Ka,true is quite negligible. In all four dissociation constants the bias seems to be quite small even though for p Ka4 it is a little bit higher, i.e., +0.05 for sinst( A) about 1.0 mAU. In the interval of sinst( A) from 0.1 to 1.0 mAU all four dissociation constants p Ki are accurate enough. Of the various regression diagnostics considered, the goodness-of-fit is the most efficient criterion of whether the parameters found adequately represent the data. The magnitude of instrumental error sinst( A) only slightly affects the shape of a Cattel's scree graph sk( A) = f( k) to determine the true number of light-absorbing species in the equilibrium mixture.

  12. The proton dissociation constant of additive effect on self-assembly of poly(3-hexyl-thiophene) for organic solar cells

    NASA Astrophysics Data System (ADS)

    Wang, Po-Hsun; Lee, Hsu-Feng; Huang, Yi-Chiang; Jung, Yi-Jiun; Gong, Fang-Lin; Huang, Wen-Yao

    2014-07-01

    In the decision on the pros and cons of the optical and electrical properties of organic solar cells, the morphology has proven to be very important. Easy to change the morphology via adding a small amount of additive, because proton dissociation constant is the main reason for their application. In this study, the use of poly(3-hexylthiophene) and [6,6]-phenyl C 61-butyric acid methyl ester as the donor and acceptor materials, and were subsequently doped with different quantity of 4,4'-sulfonyldiphenol, 4,4'-dihydroxybiphenyl, biphenyl-4,4'-dithiol. When the proton dissociation constant is higher and lower respectively, the morphology reveals earthworms-like and fiber-like. For the reason that when the additive is biphenyl-4,4'-dithiol, it can improve the power conversion efficiency of about 27% and the incident photon-to-current conversion efficiency of about 12%.

  13. The dissociation constants of the cytostatic bosutinib by nonlinear least-squares regression of multiwavelength spectrophotometric and potentiometric pH-titration data.

    PubMed

    Meloun, Milan; Nečasová, Veronika; Javůrek, Milan; Pekárek, Tomáš

    2016-02-20

    Potentiometric and spectrophotometric pH-titration of the multiprotic cytostatics bosutinib for dissociation constants determination were compared. Bosutinib treats patients with positive chronic myeloid leukemia. Bosutinib exhibits four protonatable sites in a pH range from 2 to 11, where two pK are well separated (ΔpK>3), while the other two are near dissociation constants. In the neutral medium, bosutinib occurs in the slightly water soluble form LH that can be protonated to the soluble cation LH4(3+). The molecule LH can be dissociated to still difficultly soluble anion L(-). The set of spectra upon pH from 2 to 11 in the 239.3-375.0nm was divided into two absorption bands: the first one from 239.3 to 290.5nm and the second from 312.3 to 375.0nm, which differ in sensitivity of chromophores to a pH change. Estimates of pK of the entire set of spectra were compared with those of both absorption bands. Due to limited solubility of bosutinib the protonation in a mixed aqueous-methanolic medium was studied. In low methanol content of 3-6% three dissociation constants can be reliably determined with SPECFIT/32 and SQUAD(84) and after extrapolation to zero content of methanol they lead to pKc1=3.43(12), pKc2=4.54(10), pKc3=7.56(07) and pKc4=11.04(05) at 25°C and pKc1=3.44(06), pKc2=5.03(08) pKc3=7.33(05) and pKc4=10.92(06) at 37°C. With an increasing content of methanol in solvent the dissociation of bosutinib is suppressed and the percentage of LH3(2+) decreases and LH prevails. From the potentiometric pH-titration at 25°C the concentration dissociation constants were estimated with ESAB pKc1=3.51(02), pKc2=4.37(02), pKc3=7.97(02) and pKc4=11.05(03) and with HYPERQUAD: pKc1=3.29(12), pKc2=4.24(10), pKc3=7.95(07) and pKc4=11.29(05).

  14. Mapping of the high affinity Fc epsilon receptor binding site to the third constant region domain of IgE.

    PubMed Central

    Nissim, A; Jouvin, M H; Eshhar, Z

    1991-01-01

    Identification of the precise region(s) on the IgE molecule that take part in the binding of IgE to its high affinity receptor (Fc epsilon RI) may lead to the design of IgE analogues able to block the allergic response. To localize the Fc epsilon RI-binding domain of mouse IgE, we attempted to confer on human IgE, which normally does not bind to the rodent receptor, the ability to bind to the rat Fc epsilon RI. Employing exon shuffling, we have expressed chimeric epsilon-heavy chain genes composed of a mouse (4-hydroxy-3-nitrophenyl)acetic acid (NP)-binding VH domain, and human C epsilon in which various domains were replaced by their murine counterparts. This has enabled us to test the Fc epsilon RI-binding of each mouse IgE domain while maintaining the overall conformation of the molecule. All of the chimeric IgE molecules which contain the murine C epsilon 3, bound equally to both the rodent and human receptor, as well as to monoclonal antibodies recognizing a site on IgE which is identical or very close to the Fc epsilon RI binding site. Deletion of the second constant region domain did not impair either the binding capacity of the mutated IgE or its ability to mediate mast cell degradation. These results assign the third epsilon domain of IgE as the principal region involved in the interaction with the Fc epsilon RI. Images PMID:1824934

  15. Site-Specific Polymer Attachment to HR2 Peptide Fusion Inhibitors against HIV-1 Decreases Binding Association Rates and Dissociation Rates Rather Than Binding Affinity.

    PubMed

    Danial, Maarten; Stauffer, Angela N; Wurm, Frederik R; Root, Michael J; Klok, Harm-Anton

    2017-03-15

    A popular strategy for overcoming the limited plasma half-life of peptide heptad repeat 2 (HR2) fusion inhibitors against HIV-1 is conjugation with biocompatible polymers such as poly(ethylene glycol) (PEG). However, despite improved resistance to proteolysis and reduced renal elimination, covalent attachment of polymers often causes a loss in therapeutic potency. In this study, we investigated the molecular origins of the loss in potency upon conjugation of linear, midfunctional, and hyperbranched PEG-like polymers to peptides that inhibit HIV-1-host cell membrane fusion. Fluorescence binding assays revealed that polymer conjugation imparted mass transport limitations that manifested as coexistent slower association and dissociation rates from the gp41 target on HIV-1. Furthermore, reduced association kinetics rather than affinity disruption was responsible for the loss in antiviral potency. Finally, the binding assays indicated that the unmodified HR2-derived peptide demonstrated diffusion-limited binding. The observed high potency of the unmodified peptide in HIV-1 inhibition assays was therefore attributed to rapid peptide conformational changes upon binding to the gp41 prehairpin structure. This study emphasizes that the view in which polymer ligation to therapeutic peptides inadvertently leads to loss in potency due to a loss in binding affinity requires scientific verification on a case-by-case basis and that high peptide potency may be due to rapid target-binding events.

  16. Lattice-constant and electron-affinity effects on negative-ion conversion in atom-ionic-crystal-surface grazing scattering

    NASA Astrophysics Data System (ADS)

    Zhou, Wang; Zhou, Hu; Zhang, Meixiao; Zhou, Lihua; Li, Yuan; Li, Bowen; Chen, Ximeng

    2016-11-01

    The effects of the lattice constant and electron affinity on the negative-ion conversion of a neutral atom undergoing grazing scattering on an ionic-crystal surface over the complete velocity range were investigated. Here, a comparison of negative-ion conversion of neutral O0-KCl (100 ) , F0-KCl (100 ) , and O0-KI (100 ) surface systems shows that the pronounced difference in the efficiency of negative-ion formation between F0-KCl (100 ) and O0-KCl (100 ) is caused by the large difference in their projectile electron affinities, whereas the difference between O0-KI (100 ) and O0-KCl (100 ) is caused by the difference in their lattice constants.

  17. Formation and reactivity of a porphyrin iridium hydride in water: acid dissociation constants and equilibrium thermodynamics relevant to Ir-H, Ir-OH, and Ir-CH2- bond dissociation energetics.

    PubMed

    Bhagan, Salome; Wayland, Bradford B

    2011-11-07

    Aqueous solutions of group nine metal(III) (M = Co, Rh, Ir) complexes of tetra(3,5-disulfonatomesityl)porphyrin [(TMPS)M(III)] form an equilibrium distribution of aquo and hydroxo complexes ([(TMPS)M(III)(D(2)O)(2-n)(OD)(n)]((7+n)-)). Evaluation of acid dissociation constants for coordinated water show that the extent of proton dissociation from water increases regularly on moving down the group from cobalt to iridium, which is consistent with the expected order of increasing metal-ligand bond strengths. Aqueous (D(2)O) solutions of [(TMPS)Ir(III)(D(2)O)(2)](7-) react with dihydrogen to form an iridium hydride complex ([(TMPS)Ir-D(D(2)O)](8-)) with an acid dissociation constant of 1.8(0.5) × 10(-12) (298 K), which is much smaller than the Rh-D derivative (4.3 (0.4) × 10(-8)), reflecting a stronger Ir-D bond. The iridium hydride complex adds with ethene and acetaldehyde to form organometallic derivatives [(TMPS)Ir-CH(2)CH(2)D(D(2)O)](8-) and [(TMPS)Ir-CH(OD)CH(3)(D(2)O)](8-). Only a six-coordinate carbonyl complex [(TMPS)Ir-D(CO)](8-) is observed for reaction of the Ir-D with CO (P(CO) = 0.2-2.0 atm), which contrasts with the (TMPS)Rh-D analog which reacts with CO to produce an equilibrium with a rhodium formyl complex ([(TMPS)Rh-CDO(D(2)O)](8-)). Reactivity studies and equilibrium thermodynamic measurements were used to discuss the relative M-X bond energetics (M = Rh, Ir; X = H, OH, and CH(2)-) and the thermodynamically favorable oxidative addition of water with the (TMPS)Ir(II) derivatives.

  18. Determination of the dissociation constants (pKa) of secondary and tertiary amines in organic media by capillary electrophoresis and their role in the electrophoretic mobility order inversion.

    PubMed

    Cantu, Marcelo Delmar; Hillebranda, Sandro; Carrilho, Emanuel

    2005-03-11

    Non-aqueous capillary electrophoresis (NACE) may provide a selectivity enhancement in separations since the analyte dissociation constants (pKa) in organic media are different from those in aqueous solutions. In this work, we have studied the inversion in mobility order observed in the separation of tertiary (imipramine (IMI) and amitryptiline (AMI)) and secondary amines (desipramine (DES) and nortryptiline (NOR)) in water, methanol, and acetonitrile. We have determined the pKa values in those solvents and the variation of dissociation constants with the temperature. From these data, and applying the Van't Hoff equation, we have calculated the thermodynamic parameters deltaH and deltaS. The pKa values found in methanol for DES, NOR, IMI, and AMI were 10.80, 10.79, 10.38, and 10.33, respectively. On the other hand, in acetonitrile an opposite relation was found since the values were 20.60, 20.67, 20.74, and 20.81 for DES, NOR, IMI, and AMI. This is the reason why a migration order inversion is observed in NACE for these solvents. The thermodynamic parameters were evaluated and presented a tendency that can be correlated with that observed for pKa values.

  19. Equivalence of Mg2+ and Na+ ions in salt dependence of the equilibrium binding and dissociation rate constants of Escherichia coli RNA polymerase open complex.

    PubMed

    Loziński, Tomasz; Bolewska, Krystyna; Wierzchowski, Kazimierz L

    2009-06-01

    Conflicting experimental data on the influence of Mg(2+) ions on the salt dependence of formation/dissociation of open transcription complex (RPo) of Escherichia coli RNA polymerase led us to carry systematic measurements of the dissociation rate constant (k(d)) and thermodynamic stability of complexes at lambdaP(R) and Pa promoters in a broad range of [NaCl] and [MgCl(2)] at 25, 31 and 37 degrees C, using fluorescence detected abortive transcription assay. Values of k(d) determined in MgCl(2) in the presence of heparin, as a commonly used anionic competitor, were shown to depend on heparin concentration whereas in NaCl this effect was not observed. Kinetics of dissociation was therefore determined in the course of salt-induced down-shift of the binding equilibrium. Salt derivatives of k(d)'s (n(d)) appeared to be similar in NaCl (approximately 8.5) and MgCl(2) (approximately 10) for both complexes. Isotherms of fractional occupancy of promoters by RNAP as a function of ln [salt] were shown to conform to a sigmoid Boltzman function parameterized to include binding constant of RPo and a net change (n(obs)) in the number of electrolyte ions associated with complex components upon its formation/dissociation. The fitted values of n(obs) appeared also similar in NaCl and in MgCl(2): approximately 18 for RPo/lambdaP(R) and approximately 20 for RPo/Pa, respectively. Overall unfavorable vant'Hoff enthalpy (DeltaH(obs)) of RPo proved to be much higher in MgCl(2) than in NaCl by ca. 20 kcal/mol for both complexes, rendering them profoundly less stable in the former salt. In both salts, DeltaH(obs) was higher by approximately 30 kcal/mol for RPo/Pa relative to RPo/lambdaP(R). Similarity of n(obs) and n(d) values for the two salts indicates thermodynamic equivalence of Mg(2+) and Na(+) in [salt]-controlled binding equilibrium of RPo. This finding remains in disagreement with earlier data and suggests that salt effects on open complex stability should be sought in global

  20. Spectral, structural and quantum chemical computational and dissociation constant studies of a novel azo-enamine tautomer

    NASA Astrophysics Data System (ADS)

    Gözel, Asuman; Kose, Muhammet; Karakaş, Duran; Atabey, Hasan; McKee, Vickie; Kurtoglu, Mukerrem

    2014-09-01

    We report here the synthesis of (6Z)-4-[(E)-(4-ethylphenyl)diazenyl]-6-{[(2-hydroxy-5-methylphenyl)amino]methylidene}cyclohexa-2,4-dien-1-one by the condensation reaction between 2-amino-4-methylphenol and 5-[(E)-(4-ethylphenyl)diazenyl]-2-hydroxybenzaldehyde in equimolar ratio in MeOH and characterized by elemental analyses, infrared, electronic, mass,1H and 13C NMR spectroscopy. Molecular structure of the azo-enamine dye was also determined by single crystal X-ray diffraction technique. X-ray investigation of the dye showed that azo-enamine tautomer is favoured in the solid state. There is an intramolecular hydrogen bonding (N3-H···O1) in the molecule forming a S(6) graph set motif. Additionally, there is an intermolecular O2-H···O1 hydrogen bonding in the structure. The same intermolecular hydrogen bonding contacts are extended between the other symmetry-related molecules in their respective planes to form a 1D hydrogen bond chain. Self-isomerisation via intramolecular proton transfer was investigated by UV-Vis. spectra and theoretical calculations. Effects of polarity and temperature on UV-Vis. spectra were examined in detail. Moreover, acid dissociation properties of the polydentate compound was investigated at 25 ± 0.1 °C.

  1. Estimation of the receptor-state affinity constants of ligands in functional studies using wild type and constitutively active mutant receptors: Implications for estimation of agonist bias.

    PubMed

    Ehlert, Frederick J; Stein, Richard S L

    We describe a method for estimating the affinities of ligands for active and inactive states of a G protein-coupled receptor (GPCR). Our protocol involves measuring agonist-induced signaling responses of a wild type GPCR and a constitutively active mutant of it under control conditions and after partial receptor inactivation or reduced receptor expression. Our subsequent analysis is based on the assumption that the activating mutation increases receptor isomerization into the active state without affecting the affinities of ligands for receptor states. A means of confirming this assumption is provided. Global nonlinear regression analysis yields estimates of 1) the active (Kact) and inactive (Kinact) receptor-state affinity constants, 2) the isomerization constant of the unoccupied receptor (Kq-obs), and 3) the sensitivity constant of the signaling pathway (KE-obs). The latter two parameters define the output response of the receptor, and hence, their ratio (Kq-obs/KE) is a useful measure of system bias. If the cellular system is reasonably stable and the Kq-obs and KE-obs values of the signaling pathway are known, the Kact and Kinact values of additional agonists can be estimated in subsequent experiments on cells expressing the wild type receptor. We validated our method through computer simulation, an analytical proof, and analysis of previously published data. Our approach provides 1) a more meaningful analysis of structure-activity relationships, 2) a means of validating in silico docking experiments on active and inactive receptor structures and 3) an absolute, in contrast to relative, measure of agonist bias.

  2. Label-Free Determination of the Dissociation Constant of Small Molecule-Aptamer Interaction by Isothermal Titration Calorimetry.

    PubMed

    Vogel, Marc; Suess, Beatrix

    2016-01-01

    Isothermal titration calorimetry (ITC) is a powerful label-free technique to determine the binding constant as well as thermodynamic parameters of a binding reaction and is therefore well suited for the analysis of small molecule-RNA aptamer interaction. We will introduce you to the method and present a protocol for sample preparation and the calorimetric measurement. A detailed note section will point out useful tips and pitfalls.

  3. 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.

    PubMed

    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.

  4. N-Terminal Derivatization with Structures Having High Proton Affinity for Discrimination between Leu and Ile Residues in Peptides by High-Energy Collision-Induced Dissociation.

    PubMed

    Kitanaka, Atsushi; Miyashita, Masahiro; Kubo, Ayumi; Satoh, Takaya; Toyoda, Michisato; Miyagawa, Hisashi

    2016-01-01

    De novo sequencing is still essential in the identification of peptides and proteins from unexplored organisms whose sequence information is not available. One of the remaining problems in de novo sequencing is discrimination between Leu and Ile residues. The discrimination is possible based on differences in side chain fragmentation between Leu and Ile under high-energy collision-induced dissociation (HE-CID) conditions. However, this is observed only when basic residues, such as Arg and Lys, are present near the N- or C-terminal end. It has been shown that the charge derivatization at the N-terminal end by a quarternary ammonium or phosphonium moiety facilitates the side chain fragmentation by HE-CID. However, the effective backbone fragmentation by low-energy CID (LE-CID) is often hampered in those derivatives with a fixed charge. Previously, we demonstrated that the N-terminal charge derivatization with the structures having high proton affinity induced the preferential formation of b-ions under LE-CID conditions, allowing straightforward interpretation of product ion spectra. In the present study, we further investigated whether the same derivatization approach is also effective for discrimination between Leu and Ile under HE-CID conditions. Consequently, the side chain fragmentation of Leu and Ile residues was most effectively enhanced by the N-terminal derivatization with 4-(guanidinomethyl)benzoic acid among the tested structures. This derivatization approach, which is compatible with both HE- and LE-CID analysis, offers a straightforward and unambiguous de novo peptide sequencing method.

  5. N-Terminal Derivatization with Structures Having High Proton Affinity for Discrimination between Leu and Ile Residues in Peptides by High-Energy Collision-Induced Dissociation

    PubMed Central

    Kitanaka, Atsushi; Miyashita, Masahiro; Kubo, Ayumi; Satoh, Takaya; Toyoda, Michisato; Miyagawa, Hisashi

    2016-01-01

    De novo sequencing is still essential in the identification of peptides and proteins from unexplored organisms whose sequence information is not available. One of the remaining problems in de novo sequencing is discrimination between Leu and Ile residues. The discrimination is possible based on differences in side chain fragmentation between Leu and Ile under high-energy collision-induced dissociation (HE-CID) conditions. However, this is observed only when basic residues, such as Arg and Lys, are present near the N- or C-terminal end. It has been shown that the charge derivatization at the N-terminal end by a quarternary ammonium or phosphonium moiety facilitates the side chain fragmentation by HE-CID. However, the effective backbone fragmentation by low-energy CID (LE-CID) is often hampered in those derivatives with a fixed charge. Previously, we demonstrated that the N-terminal charge derivatization with the structures having high proton affinity induced the preferential formation of b-ions under LE-CID conditions, allowing straightforward interpretation of product ion spectra. In the present study, we further investigated whether the same derivatization approach is also effective for discrimination between Leu and Ile under HE-CID conditions. Consequently, the side chain fragmentation of Leu and Ile residues was most effectively enhanced by the N-terminal derivatization with 4-(guanidinomethyl)benzoic acid among the tested structures. This derivatization approach, which is compatible with both HE- and LE-CID analysis, offers a straightforward and unambiguous de novo peptide sequencing method. PMID:27900234

  6. Dissociation constants and speciation in aqueous Li 2SO 4 and K 2SO 4 from measurements of electrical conductance to 673 K and 29 MPa

    NASA Astrophysics Data System (ADS)

    Sharygin, Andrei V.; Grafton, Brian K.; Xiao, Caibin; Wood, Robert H.; Balashov, Victor N.

    2006-10-01

    The electrical conductivities of aqueous solutions of Li 2SO 4 and K 2SO 4 have been measured at 523-673 K at 20-29 MPa in dilute solutions for molalities up to 2 × 10 -2 mol kg -1. These conductivities have been fitted to the conductance equation of Turq, Blum, Bernard, and Kunz with a consensus mixing rule and mean spherical approximation activity coefficients. In the temperature interval 523-653 K, where the dielectric constant, ɛ, is greater than 14, the electrical conductance data can be fitted by a solution model which includes ion association to form MSO4-, MSO40, and HSO4-, where M is Li or K. The adjustable parameters of this model are the first and second dissociation constants of the M 2SO 4. For the 673 K and 300 kg m -3 state point where the Coulomb interactions are the strongest (dielectric constant, ɛ = 5), models with more extensive association give good fits to the data. In the case of the Li 2SO 4 model, including the multi-ion associate, Li(SO)80, gave an extremely good fit to the conductance data.

  7. Fermion Interactions, Cosmological Constant and Space-Time Dimensionality in a Unified Approach Based on Affine Geometry

    NASA Astrophysics Data System (ADS)

    Capozziello, Salvatore; Cirilo-Lombardo, Diego Julio; Dorokhov, Alexander E.

    2014-11-01

    One of the main features of unified models, based on affine geometries, is that all possible interactions and fields naturally arise under the same standard. Here, we consider, from the effective Lagrangian of the theory, the torsion induced 4-fermion interaction. In particular, how this interaction affects the cosmological term, supposing that a condensation occurs for quark fields during the quark-gluon/hadron phase transition in the early universe. We explicitly show that there is no parity-violating pseudo-scalar density, dual to the curvature tensor (Holst term) and the spinor-bilinear scalar density has no mixed couplings of A-V form. On the other hand, the space-time dimensionality cannot be constrained from multidimensional phenomenological models admitting torsion.

  8. Analysis of agonism and inverse agonism in functional assays with constitutive activity: estimation of orthosteric ligand affinity constants for active and inactive receptor states.

    PubMed

    Ehlert, Frederick J; Suga, Hinako; Griffin, Michael T

    2011-08-01

    We describe a modification of receptor theory for the estimation of observed affinities (K(obs)) and relative efficacies of orthosteric ligands in functional assays that exhibit constitutive activity. Our theory includes parameters for the fractions of the occupied receptor population in the active (intrinsic efficacy, ε) and inactive (ε(i)) states and analogous parameters for the fractions of the free receptor population in the active (ε(sys)) and inactive (ε(i-sys)) states. The total stimulus represents the summation of the active states of the free and occupied receptor populations. A modified operational model is developed that expresses the response as a logistic function of the total stimulus. This function includes the standard parameters related to affinity and efficacy (K(obs) and τ) as well as a parameter proportional to the activity of the free receptor complex, τ(sys). Two related parameters are proportional to the fraction of the free (τ(i-sys)) and occupied (τ(i)) receptor populations in the inactive state. We show that the estimates of the affinity constants of orthosteric ligands for the active (K(b)) and inactive (K(a)) states of the receptor are equivalent to τK(obs)/τ(sys) and τ(i)K(obs)/τ(i-sys), respectively. We verify our method with computer simulation techniques and apply it to the analysis of M(2) and M(3) muscarinic receptors. Our method is applicable in the analysis of ligand bias in drug discovery programs.

  9. Bovine β-lactoglobulin is dimeric under imitative physiological conditions: dissociation equilibrium and rate constants over the pH range of 2.5-7.5.

    PubMed

    Mercadante, Davide; Melton, Laurence D; Norris, Gillian E; Loo, Trevor S; Williams, Martin A K; Dobson, Renwick C J; Jameson, Geoffrey B

    2012-07-18

    The oligomerization of β-lactoglobulin (βLg) has been studied extensively, but with somewhat contradictory results. Using analytical ultracentrifugation in both sedimentation equilibrium and sedimentation velocity modes, we studied the oligomerization of βLg variants A and B over a pH range of 2.5-7.5 in 100 mM NaCl at 25°C. For the first time, to our knowledge, we were able to estimate rate constants (k(off)) for βLg dimer dissociation. At pH 2.5 k(off) is low (0.008 and 0.009 s(-1)), but at higher pH (6.5 and 7.5) k(off) is considerably greater (>0.1 s(-1)). We analyzed the sedimentation velocity data using the van Holde-Weischet method, and the results were consistent with a monomer-dimer reversible self-association at pH 2.5, 3.5, 6.5, and 7.5. Dimer dissociation constants K(D)(2-1) fell close to or within the protein concentration range of ∼5 to ∼45 μM, and at ∼45 μM the dimer predominated. No species larger than the dimer could be detected. The K(D)(2-1) increased as |pH-pI| increased, indicating that the hydrophobic effect is the major factor stabilizing the dimer, and suggesting that, especially at low pH, electrostatic repulsion destabilizes the dimer. Therefore, through Poisson-Boltzmann calculations, we determined the electrostatic dimerization energy and the ionic charge distribution as a function of ionic strength at pH above (pH 7.5) and below (pH 2.5) the isoelectric point (pI∼5.3). We propose a mechanism for dimer stabilization whereby the added ionic species screen and neutralize charges in the vicinity of the dimer interface. The electrostatic forces of the ion cloud surrounding βLg play a key role in the thermodynamics and kinetics of dimer association/dissociation.

  10. Asymmetry in inward- and outward-affinity constant of transport explain unidirectional lysine flux in Saccharomyces cerevisiae

    PubMed Central

    Bianchi, Frans; Klooster, Joury S. van ‘t; Ruiz, Stephanie J.; Luck, Katja; Pols, Tjeerd; Urbatsch, Ina L.; Poolman, Bert

    2016-01-01

    The import of basic amino acids in Saccharomyces cerevisiae has been reported to be unidirectional, which is not typical of how secondary transporters work. Since studies of energy coupling and transport kinetics are complicated in vivo, we purified the major lysine transporter (Lyp1) of yeast and reconstituted the protein into lipid vesicles. We show that the Michaelis constant (KM) of transport from out-to-in is well in the millimolar range and at least 3 to 4-orders of magnitude higher than that of transport in the opposite direction, disfavoring the efflux of solute via Lyp1. We also find that at low values of the proton motive force, the transport by Lyp1 is comparatively slow. We benchmarked the properties of eukaryotic Lyp1 to that of the prokaryotic homologue LysP and find that LysP has a similar KM for transport from in-to-out and out-to-in, consistent with rapid influx and efflux. We thus explain the previously described unidirectional nature of lysine transport in S. cerevisiae by the extraordinary kinetics of Lyp1 and provide a mechanism and rationale for previous observations. The high asymmetry in transport together with secondary storage in the vacuole allow the cell to accumulate basic amino acids to very high levels. PMID:27550794

  11. Analysis of fast and slow acid dissociation equilibria of 3',3″,5',5″-tetrabromophenolphthalein and determination of its equilibrium constants by capillary zone electrophoresis.

    PubMed

    Takayanagi, Toshio

    2013-01-01

    Acid dissociation constants of 3',3″,5',5″-tetrabrompohenolphthalein (TBPP) were determined in an aqueous solution by capillary zone electrophoresis at an ionic strength of 0.01 mol/L. Two steps of the fast acid-dissociation equilibria including precipitable species of H2TBPP were analyzed at a weakly acidic pH region by using the change in effective electrophoretic mobility of TBPP with the pH of the separation buffer. On the other hand, an acid-dissociation reaction of TBPP at an alkaline pH region was reversible, but very slow to reach its equilibrium; the two TBPP species concerned with the equilibrium were detected as distinct signals in the electropherograms. After reaching its equilibrium, the acid-dissociation constant was determined with the signal height corresponding to its dianion form. Thus, three steps of the acid dissociation constants of TBPP were determined in an aqueous solution as pKa1 = 5.29 ± 0.06, pKa2 = 6.35 ± 0.02, and pKa3 = 11.03 ± 0.04.

  12. Effects of deuterium oxide on the rate and dissociation constants for saxitoxin and tetrodotoxin action. Voltage-clamp studies on frog myelinated nerve

    PubMed Central

    1981-01-01

    The actions of tetrodotoxin (TTX) and saxitoxin (STX) in normal water and in deuterium oxide (D2O) have been studied in frog myelinated nerve. Substitution of D2O for H2O in normal Ringer's solution has no effect on the potency of TTX in blocking action potentials but increases the potency of STX by approximately 50%. Under voltage clamp, the steady-state inhibition of sodium currents by 1 nM STX is doubled in D2O as a result of a halving of the rate of dissociation of STX from the sodium channel; the rate of block by STX is not measurably changed by D2O. Neither steady-state inhibition nor the on- or off-rate constants of TTX are changed by D2O substitution. The isotopic effects on STX binding are observed less than 10 min after the toxin has been added to D2O, thus eliminating the possibility that slow-exchange (t 1/2 greater than 10 h) hydrogen-binding sites on STX are involved. The results are consistent with a hypothesis that attributes receptor-toxin stabilization to isotopic changes of hydrogen bonding; this interpretation suggests that hydrogen bonds contribute more to the binding of STX than to that of TTX at the sodium channel. PMID:6268735

  13. Determination of the microscopic equilibrium dissociation constants for risedronate and its analogues reveals two distinct roles for the nitrogen atom in nitrogen-containing bisphosphonate drugs.

    PubMed

    Hounslow, Andrea M; Carran, John; Brown, Richard J; Rejman, Dominik; Blackburn, G Michael; Watts, Donald J

    2008-07-24

    Microscopic equilibrium dissociation constants, k as, were determined for four nitrogen-containing bisphosphonates (N-BP): risedronate and its analogues 2-(2-aminophenyl)-1-hydroxyethylidene-1,1-bisphosphonate, NE 11807, and NE 97220. The proportion of each and of analogues 2-(3'-( N-ethyl)pyridinium)-ethylidenebisphosphonate and 2-(3-piperinidyl)-1-hydroxyethylidene-1,1-bisphosphonate, having a positively charged nitrogen and three negative charges on the bisphosphonate group ("carbocation analogue") at pH 7.5, was calculated. When set in order of increasing potency at inhibiting farnesyl diphosphate (FDP) synthase (their intracellular target), the N-BPs are also ranked in order of decreasing mole fraction of carbocation analogue. However, only a weak correlation exists between potency for inhibiting FDP synthase and potency for inhibiting Dictyostelium discoideum growth. It is concluded that, although high potency for inhibiting FDP synthase is favored when the nitrogen atom in a N-BP is uncharged, N-BPs having a positively charged nitrogen can still be potent inhibitors of Dictyostelium growth owing to favorable interaction with a second, unidentified target.

  14. Modification of the wettability of a polymeric substrate by pH effect. Determination of the surface acid dissociation constant by contact angle measurements.

    PubMed

    Badre, Chantal; Mayaffre, Alain; Letellier, Pierre; Turmine, Mireille

    2006-09-26

    The wetting properties of a substrate can be changed by chemical reaction. Here, we studied simple materials with acid-base properties, by preparing poly(vinyl chloride) films containing lauric acid. These substrates constitute simple polymeric surfaces the wettability of which can be easily controlled by the acid-base equilibrium. The roughness of the material was then varied by adding Aerosil (hydrophobic fumed silica). We then studied the wettability of these materials toward aqueous buffer solutions between pH 2 and 12 from contact angle measurements. The variation of the contact angle of a droplet of buffer solution with the pH of the solution was described by a simple thermodynamic model requiring only two parameters. Thus, we could characterize the acid polymer by an effective surface acid dissociation constant the value of which was consistent with those obtained with a similar surface. We showed that the behavior of any substrate could be described even if the surface geometry was not well-known.

  15. Application of chemometrics in determination of the acid dissociation constants (pKa) of several benzodiazepine derivatives as poorly soluble drugs in the presence of ionic surfactants.

    PubMed

    Shayesteh, Tavakol Heidary; Radmehr, Moojan; Khajavi, Farzad; Mahjub, Reza

    2015-03-10

    In this study, the acid dissociation constants (pKa) of some benzodiazepine derivatives including chlordiazepoxide, clonazepam, lorazepam, and oxazepam in aqueous micellar solution were determined spectrophotometrically at an ionic strength of 0.1M at 25°C. The effect of cetyl trimethylammonium bromide (CTAB) as a cationic and sodium n-dodecyl sulfate(SDS) as an anionic surfactant on the absorption spectra of benzodiazepine drugs at different pH values were studied. The acidity constants of all related species are estimated by considering the surfactant concept and the application of chemometric methods using the whole spectral fitting of the collected data to an established factor analysis model. DATAN® software (Ver. 5.0, Multid Analyses AB, and Goteborg, Sweden) was applied to determine the acidity constants. In this study, a simple and fast method to determine the ionization constant (pKa) of poorly soluble drugs was developed using surfactants. The acidity constant (i.e. pKa) for chlordiazepoxide, clonazepam, lorazepam, and oxazepam were reported as 4.62, pKa1 value of 1.52 and pKa2 value of 10.51, pKa1 value of 1.53 and pKa2 value of 10.92 and pKa1 value 1.63 and pKa2 value of 11.21 respectively. The results showed that the peak values in the spectrophotometric absorption spectra of drugs are influenced by the presence of anionic and cationic surfactants. According to the results, by changing the SDS concentration from 0 to 0.05M, the pKa of chlordiazepoxide was increased to 5.9, the pKa1 of lorazepam was decreased to 0.1 while the pKa2 was increased to 11.5. Increase in SDS concentration has not shown significant alteration in pKa of clonazepam and oxazepam. Results indicate that by Changing the CTAB concentration from 0 to 0.05M, the pKa of chlordiazepoxide was reduced to 4.4, the pKa1 of clonazepam was decreased to 0.1 and the pKa2 was decreased to 9.1, the pKa1 of lorazepam was decreased to 0.4 and the pKa2 was decreased to 9.4, the pKa1 of oxazepam was

  16. Revision of the affinity constant for perchlorate binding to the sodium-iodide symporter based on in vitro and human in vivo data.

    PubMed

    Schlosser, Paul M

    2016-12-01

    A series of previously published physiologically based pharmacokinetic (PBPK) models describe the effect of perchlorate on iodide uptake by the thyroid, with the mechanism being competitive inhibition of iodide transport by the sodium-iodide symporter (NIS). Hence a key parameter of these models is the affinity of perchlorate for the NIS, characterized as the Michaelis-Menten kinetic constant, Km . However, when model predictions were compared to published results of a human study measuring radio-iodide uptake (RAIU) inhibition after controlled perchlorate exposures, it was found to only fit the lowest exposure level and underpredicted RAIU inhibition at higher levels. Published in vitro data, in which perchlorate-induced inhibition of iodide uptake via the NIS was measured, were re-analyzed. Km for binding of perchlorate to the NIS originally derived from these data, 1.5 μm, had been obtained using Lineweaver-Burk plots, which allow for linear regression but invert the signal-noise of the data. Re-fitting these data by non-linear regression of the non-inverted data yielded a 60% lower value for the Km , 0.59 μm. Substituting this value into the PBPK model for an average adult human significantly improved model agreement with the human RAIU data for exposures <100 μg kg(-1)  day(-1) . Thus, this lower Km value both fits the in vitro NIS kinetics and provides better predictions of human in vivo RAIU data. This change in Km increases the predicted sensitivity of humans to perchlorate over twofold for low-level exposures. Published 2016. This article is a U.S. Government work and is in the public domain in the USA. Published 2016. This article is a U.S. Government work and is in the public domain in the USA.

  17. Ultrasensitive characterization of site-specific glycosylation of affinity-purified haptoglobin from lung cancer patient plasma using 10 μm i.d. porous layer open tubular liquid chromatography-linear ion trap collision-induced dissociation/electron transfer dissociation mass spectrometry.

    PubMed

    Wang, Dongdong; Hincapie, Marina; Rejtar, Tomas; Karger, Barry L

    2011-03-15

    Site-specific analysis of protein glycosylation is important for biochemical and clinical research efforts. Glycopeptide analysis using liquid chromatography-collision-induced dissociation/electron transfer dissociation mass spectrometry (LC-CID/ETD-MS) allows simultaneous characterization of the glycan structure and attached peptide site. However, due to the low ionization efficiency of glycopeptides during electrospray ionization, 200-500 fmol of sample per injection is needed for a single LC-MS run, which makes it challenging for the analysis of limited amounts of glycoprotein purified from biological matrixes. To improve the sensitivity of LC-MS analysis for glycopeptides, an ultranarrow porous layer open tubular (PLOT) LC column (2.5 m × 10 μm i.d.) was coupled to a linear ion trap (LTQ) collision-induced dissociation/electron transfer dissociation mass spectrometer to provide sensitive analysis of N-linked protein glycosylation heterogeneity. The potential of the developed method is demonstrated by the characterization of site-specific glycosylation using haptoglobin (Hpt) as a model protein. To limit the amount of haptoglobin to low picomole amounts of protein, we affinity purified it from 1 μL of pooled lung cancer patient plasma. A total of 26 glycoforms/glycan compositions on three Hpt tryptic glycopeptides were identified and quantified from 10 LC-MS runs with a consumption of 100 fmol of Hpt digest (13 ng of protein, 10 fmol per injection). Included in this analysis was the determination of the glycan occupancy level. At this sample consumption level, the high sensitivity of the PLOT LC-LTQ-CID/ETD-MS system allowed glycopeptide identification and structure determination, along with relative quantitation of glycans presented on the same peptide backbone, even for low abundant glycopeptides at the ∼100 amol level. The PLOT LC-MS system is shown to have sufficient sensitivity to allow characterization of site-specific protein glycosylation from trace

  18. Oxygen dissociation curves for whole blood, recorded with an instrument that continuously measures pO2 and sO2 independently at constant t, pCO2, and pH.

    PubMed

    Zwart, A; Kwant, G; Oeseburg, B; Zijlstra, W G

    1982-06-01

    We describe a method for recording oxygen dissociation curves for whole-blood specimens. The blood sample is placed in a thermostated measuring chamber, and pO2 and SO2 are measured continuously by polarography and by reflectometry, respectively. During the recording of an oxygen dissociation curve, the pO2 and SO2 signals are stored in a data-acquisition system, while pH, pCO2, and temperature are kept constant. Determination of precision and error discussion indicated that the coefficient of variation (CV) of the determination of the oxygen dissociation curve is mainly determined by the error in the measurement of SO2. The overall CV of pO2 values belonging to the lower, mid-, and upper parts of the SO2 range is estimated to be about 2.6, 3.1, and 2.1%, respectively. In practice the measurements are about 30% more precise than estimated. With our method, the fixed-acid-induced Bohr effect (H+ factor) can be determined over the entire SO2 range with much greater precision than hitherto.

  19. The lowest-lying electronic singlet and triplet potential energy surfaces for the HNO-NOH system: energetics, unimolecular rate constants, tunneling and kinetic isotope effects for the isomerization and dissociation reactions.

    PubMed

    Bozkaya, Uğur; Turney, Justin M; Yamaguchi, Yukio; Schaefer, Henry F

    2012-04-28

    The lowest-lying electronic singlet and triplet potential energy surfaces (PES) for the HNO-NOH system have been investigated employing high level ab initio quantum chemical methods. The reaction energies and barriers have been predicted for two isomerization and four dissociation reactions. Total energies are extrapolated to the complete basis set limit applying focal point analyses. Anharmonic zero-point vibrational energies, diagonal Born-Oppenheimer corrections, relativistic effects, and core correlation corrections are also taken into account. On the singlet PES, the (1)HNO → (1)NOH endothermicity including all corrections is predicted to be 42.23 ± 0.2 kcal mol(-1). For the barrierless decomposition of (1)HNO to H + NO, the dissociation energy is estimated to be 47.48 ± 0.2 kcal mol(-1). For (1)NOH → H + NO, the reaction endothermicity and barrier are 5.25 ± 0.2 and 7.88 ± 0.2 kcal mol(-1). On the triplet PES the reaction energy and barrier including all corrections are predicted to be 7.73 ± 0.2 and 39.31 ± 0.2 kcal mol(-1) for the isomerization reaction (3)HNO → (3)NOH. For the triplet dissociation reaction (to H + NO) the corresponding results are 29.03 ± 0.2 and 32.41 ± 0.2 kcal mol(-1). Analogous results are 21.30 ± 0.2 and 33.67 ± 0.2 kcal mol(-1) for the dissociation reaction of (3)NOH (to H + NO). Unimolecular rate constants for the isomerization and dissociation reactions were obtained utilizing kinetic modeling methods. The tunneling and kinetic isotope effects are also investigated for these reactions. The adiabatic singlet-triplet energy splittings are predicted to be 18.45 ± 0.2 and 16.05 ± 0.2 kcal mol(-1) for HNO and NOH, respectively. Kinetic analyses based on solution of simultaneous first-order ordinary-differential rate equations demonstrate that the singlet NOH molecule will be difficult to prepare at room temperature, while the triplet NOH molecule is viable with respect to isomerization and dissociation reactions up to

  20. Dissociative Disorders

    MedlinePlus

    ... of continuity between thoughts, memories, surroundings, actions and identity. People with dissociative disorders escape reality in ways ... at bay. Symptoms — ranging from amnesia to alternate identities — depend in part on the type of dissociative ...

  1. A Small, Dilute-Cytoplasm, High-Affinity, Novel Bacterium Isolated by Extinction Culture and Having Kinetic Constants Compatible with Growth at Ambient Concentrations of Dissolved Nutrients in Seawater

    PubMed Central

    Button, D. K.; Robertson, Betsy R.; Lepp, Paul W.; Schmidt, Thomas M.

    1998-01-01

    Dilutions of raw seawater produced a bacterial isolate capable of extended growth in unamended seawater. Its 2.9-Mb genome size and 40-fg dry mass were similar to values for many naturally occurring aquatic organotrophs, but water and DNA comprised a large portion of this small chemoheterotroph, as compared to Escherichia coli. The isolate used only a few aromatic hydrocarbons and acetate, and glucose and amino acid incorporation were entirely absent, although many membrane and cytoplasmic proteins were inducible; it was named Cycloclasticus oligotrophus. A general rate equation that incorporates saturation phenomena into specific affinity theory is derived. It is used to relate the kinetic constants for substrate uptake by the isolate to its cellular proteins. The affinity constant KA for toluene was low at 1.3 μg/liter under optimal conditions, similar to those measured in seawater, and the low value was ascribed to an unknown slow step such as limitation by a cytoplasmic enzyme; KA increased with increasing specific affinities. Specific affinities, a°s, were protocol sensitive, but under optimal conditions were 47.4 liters/mg of cells/h, the highest reported in the literature and a value sufficient for growth in seawater at concentrations sometimes found. Few rRNA operons, few cytoplasmic proteins, a small genome size, and a small cell size, coupled with a high a°s and a low solids content and the ability to grow without intentionally added substrate, are consistent with the isolation of a marine bacterium with properties typical of the bulk of those present. PMID:9797308

  2. Association-dissociation of the flavoprotein hog kidney D-amino acid oxidase. Determination of the monomer-dimer equilibrium constant and the energetics of subunit association.

    PubMed

    Horiike, K; Shiga, K; Nishina, Y; Isomoto, A; Yamano, T

    1977-11-01

    The enzyme concentration dependence of spectrophotometric titrations of hog kidney D-amino acid oxidase [EC 1.4.3.3] with p-aminobenzoate was studied. The monomer-dimer equilibrium constant of the oxidized holoenzyme at 25 degrees C was estimated to be 7 X 10(5)M-1 at pH 7.5 and 4X 10(6)M-1 at pH 8.3. The energetics of subunit association are discussed.

  3. Predicting pressure-dependent unimolecular rate constants using variational transition state theory with multidimensional tunneling combined with system-specific quantum RRK theory: a definitive test for fluoroform dissociation.

    PubMed

    Bao, Junwei Lucas; Zhang, Xin; Truhlar, Donald G

    2016-06-22

    Understanding the falloff in rate constants of gas-phase unimolecular reaction rate constants as the pressure is lowered is a fundamental problem in chemical kinetics, with practical importance for combustion, atmospheric chemistry, and essentially all gas-phase reaction mechanisms. In the present work, we use our recently developed system-specific quantum RRK theory, calibrated by canonical variational transition state theory with small-curvature tunneling, combined with the Lindemann-Hinshelwood mechanism, to model the dissociation reaction of fluoroform (CHF3), which provides a definitive test for falloff modeling. Our predicted pressure-dependent thermal rate constants are in excellent agreement with experimental values over a wide range of pressures and temperatures. The present validation of our methodology, which is able to include variational transition state effects, multidimensional tunneling based on the directly calculated potential energy surface along the tunneling path, and torsional and other vibrational anharmonicity, together with state-of-the-art reaction-path-based direct dynamics calculations, is important because the method is less empirical than models routinely used for generating full mechanisms, while also being simpler in key respects than full master equation treatments and the full reduced falloff curve and modified strong collision methods of Troe.

  4. Dissociative disorders.

    PubMed

    Kihlstrom, John F

    2005-01-01

    The dissociative disorders, including "psychogenic" or "functional" amnesia, fugue, dissociative identity disorder (DID, also known as multiple personality disorder), and depersonalization disorder, were once classified, along with conversion disorder, as forms of hysteria. The 1970s witnessed an "epidemic" of dissociative disorder, particularly DID, which may have reflected enthusiasm for the diagnosis more than its actual prevalence. Traditionally, the dissociative disorders have been attributed to trauma and other psychological stress, but the existing evidence favoring this hypothesis is plagued by poor methodology. Prospective studies of traumatized individuals reveal no convincing cases of amnesia not attributable to brain insult, injury, or disease. Treatment generally involves recovering and working through ostensibly repressed or dissociated memories of trauma; at present, there are few quantitative or controlled outcome studies. Experimental studies are few in number and have focused largely on state-dependent and implicit memory. Depersonalization disorder may be in line for the next "epidemic" of dissociation.

  5. Dissociative amnesia.

    PubMed

    Staniloiu, Angelica; Markowitsch, Hans J

    2014-08-01

    Dissociative amnesia is one of the most enigmatic and controversial psychiatric disorders. In the past two decades, interest in the understanding of its pathophysiology has surged. In this report, we review new data about the epidemiology, neurobiology, and neuroimaging of dissociative amnesia and show how advances in memory research and neurobiology of dissociation inform proposed pathogenetic models of the disorder. Dissociative amnesia is characterised by functional impairment. Additionally, preliminary data suggest that affected people have an increased and possibly underestimated suicide risk. The prevalence of dissociative amnesia differs substantially across countries and populations. Symptoms and disease course also vary, indicating a possibly heterogeneous disorder. The accompanying clinical features differ across cultural groups. Most dissociative amnesias are retrograde, with memory impairments mainly involving the episodic-autobiographical memory domain. Anterograde dissociative amnesia occurring without significant retrograde memory impairments is rare. Functional neuroimaging studies of dissociative amnesia with prevailing retrograde memory impairments show changes in the network that subserves autobiographical memory. At present, no evidence-based treatments are available for dissociative amnesia and no broad framework exists for its rehabilitation. Further research is needed into its neurobiology, course, treatment options, and strategies to improve differential diagnoses.

  6. The role of the medium on the acid dissociation constants of some azo dyes in view of experimental and theoretical data

    NASA Astrophysics Data System (ADS)

    Ebead, Y. H.

    2010-10-01

    In this study, the p Ka values of some azo dyes derived from cyclohexane-1,3-dione in different organic solvent + water mixtures have been determined spectrophotometrically. The organic solvents used are ethanol, acetone and N, N-dimethylformamide (DMF). Thus, the nature and the proportion of the organic cosolvent effects as well as the molecular structure on acidity constants were evaluated. Furthermore, the proton-donating ability of N 10H reflected by thermodynamic parameters - Δ f,298Ho, Δ 298So and Δ f,298Go - for compounds 1- 4 has been estimated using AM1and PM6 methods in the gas and aqueous phases. The data obtained provides a complete and accurate picture of the acid-base properties of the compounds under study. The p Ka values predicted in aqueous phase by the previous methods were compared with the experimental values and the best agreement with the experimental data was obtained by AM1.

  7. A novel strategy of profiling the mechanism of herbal medicines by combining network pharmacology with plasma concentration determination and affinity constant measurement.

    PubMed

    Chen, Langdong; Lv, Diya; Wang, Dongyao; Chen, Xiaofei; Zhu, Zhenyu; Cao, Yan; Chai, Yifeng

    2016-10-18

    Herbal medicines have long been widely used in the treatment of various complex diseases in China. However, the active constituents and therapeutic mechanisms of many herbal medicines remain undefined. Therefore, the identification of the active components and target proteins in these herbal medicines is a formidable task in herbal medicine research. In this study, we proposed a strategy, which integrates network pharmacology with biomedical analysis and surface plasmon resonance (SPR) to predict the active ingredients and potential targets of herbal medicine Sophora flavescens or Kushen in Chinese, and evaluate its anti-fibrosis activity. First, we applied a virtual HTDocking platform to predict the potential targets of Kushen related to liver fibrosis by selecting five crucial protein targets based on network parameters and text mining. Then, we identified nine components in mice plasma after oral administration of Kushen extract and determined the plasma concentration of each compound. Binding affinities between the nine potential active compounds and five target proteins were detected by SPR assays. Finally, we constructed a multi-parameter network model on the basis of three important parameters to tentatively explain the anti-fibrosis mechanism of Kushen. The results not only provide evidence for the therapeutic mechanism of Kushen but also shed new light on the activity-based analysis of other Chinese herbal medicines.

  8. Sodium chloride in supercritical water as a function of density: potentials of mean force and an equation for the dissociation constant from 723 to 1073 K and from 0 to 0.9 g/cm(3).

    PubMed

    Liu, Wenbin; Wood, Robert H; Doren, Douglas J

    2008-06-19

    The potential of mean force (PMF) of sodium chloride in water has been calculated by using the ab initio classical free-energy perturbation method at five state points: at 973 K with densities of 0.2796, 0.0935, and 0.0101 g/cm (3) and at 723 K with densities of 0.0897 and 0.0098 g/cm (3). The method is based on a QM-MM model in which Na-H 2O, Cl-H 2O, and Na-Cl interactions are calculated by ab initio methods. The water-water interactions are from the polarizable TIP4P-FQ model. The logarithm of the dissociation constant (log K c) has been calculated from the PMF. These predictions, together with experimental measurements, were used to derive an equation for log K c at densities from 0 to 0.9 g/cm (3) and temperatures from 723 to 1073 K, as well as from 600 to 1073 K for densities from 0.29 g/cm (3) to 0.9 g/cm (3). Extrapolation of the present equation below 723 K for densities less than 0.29 g/cm (3) does not fit the experimental results. This is attributed to long-range changes in the local dielectric constant due to the high compressibility. Comparisons with previous predictions and simulations are presented.

  9. Use of an exchange method to estimate the association and dissociation rate constants of cadmium complexes formed with low-molecular-weight organic acids commonly exuded by plant roots.

    PubMed

    Schneider, André; Nguyen, Christophe

    2011-01-01

    Organic acids released from plant roots can form complexes with cadmium (Cd) in the soil solution and influence metal bioavailability not only due to the nature and concentration of the complexes but also due to their lability. The lability of a complex influences its ability to buffer changes in the concentration of free ions (Cd); it depends on the association (, m mol s) and dissociation (, s) rate constants. A resin exchange method was used to estimate and (m mol s), which is the conditional estimate of depending on the calcium (Ca) concentration in solution. The constants were estimated for oxalate, citrate, and malate, three low-molecular-weight organic acids commonly exuded by plant roots and expected to strongly influence Cd uptake by plants. For all three organic acids, the and estimates were around 2.5 10 m mol s and 1.3 × 10 s, respectively. Based on the literature, these values indicate that the Cd- low-molecular-weight organic acids complexes formed between Cd and low-molecular-weight organic acids may be less labile than complexes formed with soil soluble organic matter but more labile than those formed with aminopolycarboxylic chelates.

  10. Calculation of the standard partial molal thermodynamic properties and dissociation constants of aqueous HCl{sup 0} and HBr{sup 0} at temperatures to 1000 C and pressures to 5 kbar

    SciTech Connect

    Pokrovskii, V.A.

    1999-04-01

    Dissociation constants of aqueous ion pairs HCl{sup 0} and HBr{sup 0} derived in the literature from vapor pressure and supercritical conductance measurements (Quist and Marshall, 1968b; Frantz and Marshall, 1984) were used to calculate the standard partial molal thermodynamic properties of the species at 25 C and 1 bar. Regression of the data with the aid of revised Helgeson-Kirkham-Flowers equations of state (Helgeson et al., 1981; Tanger and Helgeson, 1988; Shock et al., 1989) resulted in a set of equations-of-state parameters that permits accurate calculation of the thermodynamic properties of the species at high temperatures and pressures. These properties and parameters reproduce generally within 0.1 log unit (with observed maximum deviation of 0.23 log unit) the log K values for HBr{sup 0} and HCl{sup 0} given by Quist and Marshall (1968b) and Frantz and Marshall (1984), respectively, at temperatures to 800 C and pressures to 5 kbar.

  11. Km as an Apparent Dissociation Constant.

    ERIC Educational Resources Information Center

    Cohlberg, Jeffrey A.

    1979-01-01

    An approach to enzyme kinetics which emphasizes similarities between equilibrium binding and steady state kinetics is outlined. It is intended for use in teaching biochemistry to beginning students. (Author/SA)

  12. Importin {beta}-type nuclear transport receptors have distinct binding affinities for Ran-GTP

    SciTech Connect

    Hahn, Silvia; Schlenstedt, Gabriel

    2011-03-18

    Highlights: {yields} Determination of binding properties of nuclear transport receptor/Ran-GTP complexes. {yields} Biosensor measurements provide constants for dissociation, on-rates, and off-rates. {yields} The affinity of receptors for Ran-GTP is widely divergent. {yields} Dissociation constants differ for three orders of magnitude. {yields} The cellular concentration of yeast Ran is not limiting. -- Abstract: Cargos destined to enter or leave the cell nucleus are typically transported by receptors of the importin {beta} family to pass the nuclear pore complex. The yeast Saccharomyces cerevisiae comprises 14 members of this protein family, which can be divided in importins and exportins. The Ran GTPase regulates the association and dissociation of receptors and cargos as well as the transport direction through the nuclear pore. All receptors bind to Ran exclusively in its GTP-bound state and this event is restricted to the nuclear compartment. We determined the Ran-GTP binding properties of all yeast transport receptors by biosensor measurements and observed that the affinity of importins for Ran-GTP differs significantly. The dissociation constants range from 230 pM to 270 nM, which is mostly based on a variability of the off-rate constants. The divergent affinity of importins for Ran-GTP suggests the existence of a novel mode of nucleocytoplasmic transport regulation. Furthermore, the cellular concentration of {beta}-receptors and of other Ran-binding proteins was determined. We found that the number of {beta}-receptors altogether about equals the amounts of yeast Ran, but Ran-GTP is not limiting in the nucleus. The implications of our results for nucleocytoplasmic transport mechanisms are discussed.

  13. [Cell-ELA-based determination of binding affinity of DNA aptamer against U87-EGFRvIII cell].

    PubMed

    Tan, Yan; Liang, Huiyu; Wu, Xidong; Gao, Yubo; Zhang, Xingmei

    2013-05-01

    A15, a DNA aptamer with binding specificity for U87 glioma cells stably overexpressing the epidermal growth factor receptor variant III (U87-EGFRvIII), was generated by cell systematic evolution of ligands by exponential enrichment (cell-SELEX) using a random nucleotide library. Subsequently, we established a cell enzyme-linked assay (cell-ELA) to detect the affinity of A15 compared to an EGFR antibody. We used A15 as a detection probe and cultured U87-EGFRvIII cells as targets. Our data indicate that the equilibrium dissociation constants (K(d)) for A15 were below 100 nmol/L and had similar affinity compared to an EGFR antibody for U87-EGFRvIII. We demonstrated that the cell-ELA was a useful method to determine the equilibrium dissociation constants (K(d)) of aptamers generated by cell-SELEX.

  14. Sperm in poor quality semen from bulls during heat stress have a lower affinity for binding hydrogen-3 heparin

    SciTech Connect

    Ax, R.L.; Gilbert, G.R.; Shook, G.E.

    1987-01-01

    Binding assays with (/sup 3/H) heparin were performed using spermatozoa collected prior to, during, and following summer heat stress to dairy bulls. Ejaculates collected in August 1983 after a period of ambient temperatures exceeding 29.4/sup 0/C exhibited a high frequency of abnormal sperm, and motility was reduced in some samples. Sperm in samples collected during heat stress possessed dissociation constants for binding (/sup 3/H) heparin ranging from 134.5 to 163.2 nmol. In contrast, sperm in semen collected prior to and after heat stress had significantly lower dissociation constants (higher affinity) for (/sup 3/H)heparin, 12.9 to 56.4 nmol. The number of binding sites for (/sup 3/H) heparin on sperm did not change among collection periods. It was concluded that the binding affinity for (/sup 3/H) heparin may reflect membrane integrity of bull sperm.

  15. Direct detection of antibody concentration and affinity in human serum using microscale thermophoresis.

    PubMed

    Lippok, Svenja; Seidel, Susanne A I; Duhr, Stefan; Uhland, Kerstin; Holthoff, Hans-Peter; Jenne, Dieter; Braun, Dieter

    2012-04-17

    The direct quantification of both the binding affinity and absolute concentration of disease-related biomarkers in biological fluids is particularly beneficial for differential diagnosis and therapy monitoring. Here, we extend microscale thermophoresis to target immunological questions. Optically generated thermal gradients were used to deplete fluorescently marked antigens in 2- and 10-fold-diluted human serum. We devised and validated an autocompetitive strategy to independently fit the concentration and dissociation constant of autoimmune antibodies against the cardiac β1-adrenergic receptor related to dilated cardiomyopathy. As an artificial antigen, the peptide COR1 was designed to mimic the second extracellular receptor loop. Thermophoresis resolved antibody concentrations from 2 to 200 nM and measured the dissociation constant as 75 nM. The approach quantifies antibody binding in its native serum environment within microliter volumes and without any surface attachments. The simplicity of the mix and probe protocol minimizes systematic errors, making thermophoresis a promising detection method for personalized medicine.

  16. Recurrent Episodes of Dissociative Fugue

    PubMed Central

    Angothu, Hareesh; Pabbathi, Lokeswar Reddy

    2016-01-01

    Dissociative fugue is rare entity to encounter with possible differentials of epilepsy and malingering. It is one of the dissociative disorders rarely seen in clinical practice more often because of the short lasting nature of this condition. This might also be because of organized travel of the individuals during the episodes and return to their families after the recovery from episodes. This is a case description of a patient who has experienced total three episodes of dissociative fugue. The patient has presented during the third episode and two prior episodes were diagnosed as fugue episodes retrospectively based on the history. Planned travel in this case by the patient to a distant location was prevented because of early diagnosis and constant vigilance till the recovery. As in this case, it may be more likely that persons with Dissociative fugue may develop similar episodes if they encounter exceptional perceived stress. However, such conclusions may require follow-up studies. PMID:27114633

  17. The Measurements of Affinity Constants of Osmptrophs For P and Release Rates of P From Microbial Food Web From Late Summer To Early Winter In The Bay of Villefranche, The NW Mediterranean

    NASA Astrophysics Data System (ADS)

    Tanaka, T.; Rassoulzadegan, F.; Thingstad, T. F.

    Orthophosphate uptake by bacteria and phytoplankton was measured with 32P for wa- ter from the Bay of Villefranche (NW Mediterranean) from September to December 2001. The 32P release rate was measured for waters with different size of micro- bial community. During this period, the thermocline was gradually declined and the vertical mixing started, showing the transition from nutrient depleted to replete pe- riods. Before the vertical mixing started, orthophosphate turnover time ranged from 1 to 5 h. The fraction of orthophosphate uptake was dominated (mean 70%) by the 0.6-2 µm fraction where cyanobacteria biomass was dominant. Based on the theo- retical considerations proposed by Thingstad and Rassoulzadegan (1999), the affinity constants were estimated for bacteria, cyanobacteria and autotrophic nanoflagellates, respectively. They ranged from 0.001 to 0.028 L/nM-P/h for bacteria, from 0.047 to 0.103 L/nM-P/h for cyanobacteria and from 0.002 to 0.032 L/nM-P/h for autotrophic nanoflagellates when 32P turnover time was relatively shorter (<5 h). This may sup- port the previous report in which cyanobacteria were dominant in the primary pro- duction in autumn at the study site. The 32P release rates after the addition of cold chase were always less than 0.5%/h. This release appeared to be from the 0.2-0.6 or 0.6-2 µm fractions. The increase of 32P in the >2 µm fraction suggests P transfer by predation. Virus appeared to not contribute to 32P release from bacteria during the study period. These results indicate long P turnover times within particulate fraction and thus less contribution of P release from microbial food web.

  18. Calorimetric investigation of proton linkage by monitoring both the enthalpy and association constant of binding: application to the interaction of the Src SH2 domain with a high-affinity tyrosyl phosphopeptide.

    PubMed

    Bradshaw, J M; Waksman, G

    1998-11-03

    The binding of Src homology 2 (SH2) domains to tyrosyl phosphopeptides depends on electrostatic interactions between the phosphotyrosine and its binding site. To probe the role of these interactions, we have used isothermal titration calorimetry to study the pH dependence of the binding of the SH2 domain of the Src kinase to a high-affinity tyrosyl phosphopeptide. Two independent approaches were employed. In a first series of experiments that focused on determining the peptide's association constant between pH 5.0 and 9.0, two ionizable groups were characterized. One group, with free and bound pKas of 6.2 and 4.4, respectively, could be identified as the phosphate in the phosphotyrosine while the other group, with free and bound pKas of 8.2 and 8.5, respectively, could be only tentatively assigned to a cysteine in the phosphotyrosine binding pocket. Further information on the linkage between peptide binding and protonation of the phosphotyrosine was obtained from a second series of experiments, which focused on determining the peptide binding enthalpy at low values of pH in several buffers with different ionization enthalpies. These data provided free and bound pKa values for the phosphotyrosine identical to those derived from the first series of experiments, and hence demonstrated for the first time that the two approaches provide identical information regarding proton linkage. In addition, the second series of experiments also determined the intrinsic enthalpy of binding of both the protonated and deprotonated phosphate forms of the peptide. These two sets of experiments provided a complete energetic profile of the linkage between phosphate ionization and peptide binding. From this profile, it was determined that the PO32- form of the peptide binds 2.3 kcal mol-1 more favorably than the PO3H1- form due entirely to a more favorable entropy of binding.

  19. Reversible immobilization of proteins with streptavidin affinity tags on a surface plasmon resonance biosensor chip.

    PubMed

    Li, Yong-Jin; Bi, Li-Jun; Zhang, Xian-En; Zhou, Ya-Feng; Zhang, Ji-Bin; Chen, Yuan-Yuan; Li, Wei; Zhang, Zhi-Ping

    2006-11-01

    Dissociation of biotin from streptavidin is very difficult due to their high binding affinity. The re-use of streptavidin-modified surfaces is therefore almost impossible, making devices containing them (e.g. surface plasmon resonance (SPR) sensor chips) expensive. This paper describes a new protocol for reversible and site-directed immobilization of proteins with streptavidin affinity tags on the streptavidin-coated SPR biosensor chip (SA chip). Two streptavidin affinity tags, nano-tag and streptavidin-binding peptide (SBP tag), were applied. They both can specifically interact with streptavidin but have weaker binding force compared to the biotin-streptavidin system, thus allowing association and dissociation under controlled conditions. The SA chip surface could be regenerated repeatedly without loss of activity by injection of 50 mM NaOH solution. The fusion construct of a SBP tag and a single-chain antibody to mature bovine prion protein (scFv-Z186-SBP) interacts with the SA chip, resulting in a single-chain-antibody-modified surface. The chip showed kinetic response to the prion antigen with equilibrium dissociation constant K (D) approximately equal to 4.01 x 10(-7). All results indicated that the capture activity of the SA chip has no irreversible loss after repeated immobilization and regeneration cycles. The method should be of great benefit to various biosensors, biochips and immunoassay applications based on the streptavidin capture surface.

  20. Scapholunate Dissociation.

    PubMed

    Ramponi, Denise; McSwigan, Tara

    2016-01-01

    Wrist injuries are a common complaint in the emergency setting. Any disruption of the anatomy of the carpal bones can impair hand function, leading to pain, weakness, and complications. One of the most common forms of carpal bone instability is scapholunate dissociation. This injury can lead to significant morbidity including avascular necrosis, impaired healing, limited function, and arthritis. These diagnostic findings may be subtle, thus identifying high-risk mechanisms of injury, and clinical manifestations will assist the emergency practitioner with early diagnosis and treatment of this high-risk injury.

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

    PubMed

    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.

  2. Oxygen affinity and amino acid sequence of myoglobins from endothermic and ectothermic fish.

    PubMed

    Marcinek, D J; Bonaventura, J; Wittenberg, J B; Block, B A

    2001-04-01

    Myoglobin (Mb) buffers intracellular O2 and facilitates diffusion of O2 through the cell. These functions of Mb will be most effective when intracellular PO2 is near the partial pressure of oxygen at which Mb is half saturated (P50) of the molecule. We test the hypothesis that Mb oxygen affinity has evolved such that it is conserved when adjusted for body temperature among closely related animals. We measure oxygen P50s tonometrically and oxygen dissociation rate constants with stopped flow and generate amino acid sequence from cDNA of Mbs from fish with different body temperatures. P50s for the endothermic bluefin tuna, skipjack tuna, and blue marlin at 20 degrees C were 0.62 +/- 0.02, 0.59 +/- 0.01, 0.58 +/- 0.04 mmHg, respectively, and were significantly lower than those for ectothermic bonito (1.03 +/- 0.07 mmHg) and mackerel (1.39 +/- 0.03 mmHg). Because the oxygen affinity of Mb decreases with increasing temperature, the above differences in oxygen affinity between endothermic and ectothermic fish are reduced when adjusted for the in vivo muscle temperature of the animal. Oxygen dissociation rate constants at 20 degrees C for the endothermic species ranged from 34.1 to 49.3 s(-1), whereas those for mackerel and bonito were 102 and 62 s(-1), respectively. Correlated with the low oxygen affinity and fast dissociation kinetics of mackerel Mb is a substitution of alanine for proline that would likely result in a more flexible mackerel protein.

  3. Rate constants of agonist binding to muscarinic receptors in rat brain medulla. Evaluation by competition kinetics

    SciTech Connect

    Schreiber, G.; Henis, Y.I.; Sokolovsky, M.

    1985-07-25

    The method of competition kinetics, which measures the binding kinetics of an unlabeled ligand through its effect on the binding kinetics of a labeled ligand, was employed to investigate the kinetics of muscarinic agonist binding to rat brain medulla pons homogenates. The agonists studied were acetylcholine, carbamylcholine, and oxotremorine, with N-methyl-4-(TH)piperidyl benzilate employed as the radiolabeled ligand. Our results suggested that the binding of muscarinic agonists to the high affinity sites is characterized by dissociation rate constants higher by 2 orders of magnitude than those of antagonists, with rather similar association rate constants. Our findings also suggest that isomerization of the muscarinic receptors following ligand binding is significant in the case of antagonists, but not of agonists. Moreover, it is demonstrated that in the medulla pons preparation, agonist-induced interconversion between high and low affinity bindings sites does not occur to an appreciable extent.

  4. High-throughput fragment screening by affinity LC-MS.

    PubMed

    Duong-Thi, Minh-Dao; Bergström, Maria; Fex, Tomas; Isaksson, Roland; Ohlson, Sten

    2013-02-01

    Fragment screening, an emerging approach for hit finding in drug discovery, has recently been proven effective by its first approved drug, vemurafenib, for cancer treatment. Techniques such as nuclear magnetic resonance, surface plasmon resonance, and isothemal titration calorimetry, with their own pros and cons, have been employed for screening fragment libraries. As an alternative approach, screening based on high-performance liquid chromatography separation has been developed. In this work, we present weak affinity LC/MS as a method to screen fragments under high-throughput conditions. Affinity-based capillary columns with immobilized thrombin were used to screen a collection of 590 compounds from a fragment library. The collection was divided into 11 mixtures (each containing 35 to 65 fragments) and screened by MS detection. The primary screening was performed in <4 h (corresponding to >3500 fragments per day). Thirty hits were defined, which subsequently entered a secondary screening using an active site-blocked thrombin column for confirmation of specificity. One hit showed selective binding to thrombin with an estimated dissociation constant (K (D)) in the 0.1 mM range. This study shows that affinity LC/MS is characterized by high throughput, ease of operation, and low consumption of target and fragments, and therefore it promises to be a valuable method for fragment screening.

  5. Comparative Study of Three Methods for Affinity Measurements: Capillary Electrophoresis Coupled with UV Detection and Mass Spectrometry, and Direct Infusion Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Mironov, Gleb G.; Logie, Jennifer; Okhonin, Victor; Renaud, Justin B.; Mayer, Paul M.; Berezovski, Maxim V.

    2012-07-01

    We present affinity capillary electrophoresis and mass spectrometry (ACE-MS) as a comprehensive separation technique for label-free solution-based affinity analysis. The application of ACE-MS for measuring affinity constants between eight small molecule drugs [ibuprofen, s-flurbiprofen, diclofenac, phenylbutazone, naproxen, folic acid, resveratrol, and 4,4'-(propane-1,3-diyl) dibenzoic acid] and β-cyclodextrin is described. We couple on-line ACE with MS to combine the separation and kinetic capability of ACE together with the molecular weight and structural elucidation of MS in one system. To understand the full potential of ACE-MS, we compare it with two other methods: Direct infusion mass spectrometry (DIMS) and ACE with UV detection (ACE-UV). After the evaluation, DIMS provides less reliable equilibrium dissociation constants than separation-based ACE-UV and ACE-MS, and cannot be used solely for the study of noncovalent interactions. ACE-MS determines apparent dissociation constants for all reacting small molecules in a mixture, even in cases when drugs overlap with each other during separation. The ability of ACE-MS to interact, separate, and rapidly scan through m/z can facilitate the simultaneous affinity analysis of multiple interacting pairs, potentially leading to the high-throughput screening of drug candidates.

  6. A global benchmark study using affinity-based biosensors

    PubMed Central

    Rich, Rebecca L.; Papalia, Giuseppe A.; Flynn, Peter J.; Furneisen, Jamie; Quinn, John; Klein, Joshua S.; Katsamba, Phini S.; Waddell, M. Brent; Scott, Michael; Thompson, Joshua; Berlier, Judie; Corry, Schuyler; Baltzinger, Mireille; Zeder-Lutz, Gabrielle; Schoenemann, Andreas; Clabbers, Anca; Wieckowski, Sebastien; Murphy, Mary M.; Page, Phillip; Ryan, Thomas E.; Duffner, Jay; Ganguly, Tanmoy; Corbin, John; Gautam, Satyen; Anderluh, Gregor; Bavdek, Andrej; Reichmann, Dana; Yadav, Satya P.; Hommema, Eric; Pol, Ewa; Drake, Andrew; Klakamp, Scott; Chapman, Trevor; Kernaghan, Dawn; Miller, Ken; Schuman, Jason; Lindquist, Kevin; Herlihy, Kara; Murphy, Michael B.; Bohnsack, Richard; Andrien, Bruce; Brandani, Pietro; Terwey, Danny; Millican, Rohn; Darling, Ryan J.; Wang, Liann; Carter, Quincy; Dotzlaf, Joe; Lopez-Sagaseta, Jacinto; Campbell, Islay; Torreri, Paola; Hoos, Sylviane; England, Patrick; Liu, Yang; Abdiche, Yasmina; Malashock, Daniel; Pinkerton, Alanna; Wong, Melanie; Lafer, Eileen; Hinck, Cynthia; Thompson, Kevin; Primo, Carmelo Di; Joyce, Alison; Brooks, Jonathan; Torta, Federico; Bagge Hagel, Anne Birgitte; Krarup, Janus; Pass, Jesper; Ferreira, Monica; Shikov, Sergei; Mikolajczyk, Malgorzata; Abe, Yuki; Barbato, Gaetano; Giannetti, Anthony M.; Krishnamoorthy, Ganeshram; Beusink, Bianca; Satpaev, Daulet; Tsang, Tiffany; Fang, Eric; Partridge, James; Brohawn, Stephen; Horn, James; Pritsch, Otto; Obal, Gonzalo; Nilapwar, Sanjay; Busby, Ben; Gutierrez-Sanchez, Gerardo; Gupta, Ruchira Das; Canepa, Sylvie; Witte, Krista; Nikolovska-Coleska, Zaneta; Cho, Yun Hee; D’Agata, Roberta; Schlick, Kristian; Calvert, Rosy; Munoz, Eva M.; Hernaiz, Maria Jose; Bravman, Tsafir; Dines, Monica; Yang, Min-Hsiang; Puskas, Agnes; Boni, Erica; Li, Jiejin; Wear, Martin; Grinberg, Asya; Baardsnes, Jason; Dolezal, Olan; Gainey, Melicia; Anderson, Henrik; Peng, Jinlin; Lewis, Mark; Spies, Peter; Trinh, Quyhn; Bibikov, Sergei; Raymond, Jill; Yousef, Mohammed; Chandrasekaran, Vidya; Feng, Yuguo; Emerick, Anne; Mundodo, Suparna; Guimaraes, Rejane; McGirr, Katy; Li, Yue-Ji; Hughes, Heather; Mantz, Hubert; Skrabana, Rostislav; Witmer, Mark; Ballard, Joshua; Martin, Loic; Skladal, Petr; Korza, George; Laird-Offringa, Ite; Lee, Charlene S.; Khadir, Abdelkrim; Podlaski, Frank; Neuner, Phillippe; Rothacker, Julie; Rafique, Ashique; Dankbar, Nico; Kainz, Peter; Gedig, Erk; Vuyisich, Momchilo; Boozer, Christina; Ly, Nguyen; Toews, Mark; Uren, Aykut; Kalyuzhniy, Oleksandr; Lewis, Kenneth; Chomey, Eugene; Pak, Brian J.; Myszka, David G.

    2013-01-01

    To explore the variability in biosensor studies, 150 participants from 20 countries were given the same protein samples and asked to determine kinetic rate constants for the interaction. We chose a protein system that was amenable to analysis using different biosensor platforms as well as by users of different expertise levels. The two proteins (a 50-kDa Fab and a 60-kDa glutathione S-transferase [GST] antigen) form a relatively high-affinity complex, so participants needed to optimize several experimental parameters, including ligand immobilization and regeneration conditions as well as analyte concentrations and injection/dissociation times. Although most participants collected binding responses that could be fit to yield kinetic parameters, the quality of a few data sets could have been improved by optimizing the assay design. Once these outliers were removed, the average reported affinity across the remaining panel of participants was 620 pM with a standard deviation of 980 pM. These results demonstrate that when this biosensor assay was designed and executed appropriately, the reported rate constants were consistent, and independent of which protein was immobilized and which biosensor was used. PMID:19133223

  7. Dissociative phenomenology of dissociative identity disorder.

    PubMed

    Dell, Paul F

    2002-01-01

    The goal of this study was to investigate the dissociative phenomenology of dissociative identity disorder (DID). The Multidimensional Inventory of Dissociation (MID) was administered to 34 patients with DID, 23 patients with dissociative disorder not otherwise specified (DDNOS), 52 patients with mixed psychiatric disorders, and 58 normal individuals. DID patients obtained significantly higher scores than the other three groups on 27 dissociation-related variables. DDNOS patients had significantly higher scores than normals and mixed psychiatric patients on 17 and 15 dissociation-related variables, respectively. The findings of the present study are virtually identical to a large body of replicated findings about the dissociative phenomenology of DID. This broad range of dissociation-related phenomena, which routinely occurs in individuals with DID, is largely absent from the DSM-IV-TR account of DID. Factor analysis of the 11 dimensions of dissociation that are measured by the MID extracted only one factor that accounted for 85% of the variance. It was concluded that dissociation is a unifactorial taxon or natural type that has different aspects or epiphenomena (i.e., amnesia, depersonalization, voices, trance, etc.).

  8. Ab initio molecular orbital study of substituent effects in vaska type complexes (trans-IrL{sub 2}(CO)X): Electron affinities, ionization potentials, carbonyl stretch frequencies, and the thermodynamics of H{sub 2} dissociative addition

    SciTech Connect

    Abu-Hasanayn, F.; Goldman, A.S.; Krogh-Jespersen, K.

    1994-10-26

    Ab initio electronic structure calculations are used to study substituent effects in Vaska-type complexes, trans-IrL{sub 2}(CO)X (1-X) (X = F, Cl, Br, I, CN, H, CH{sub 3}, SiH{sub 3}, OH, and SH; L = PH{sub 3}). Both the electron affinity and the ionization potential of 1-X are computed to increase upon descending the halogen series of complexes, which indicates, surprisingly, that the complexes with more electronegative halogens are more difficult to reduce and easier to oxidize. The computed electron affinity trend is consistent with the half-wave reduction potential trend known for 1-X (L = PPh{sub 3}; X = F, Cl, Br, and I). Computed carbonyl stretch frequencies for 1-X are greater than experimental values (L = PPh{sub 3}), but observed trends are well reproduced. The redox and spectroscopic trends are discussed in terms of the substituent effects on the electronic structure of 1-X, particularly as revealed in the molecular orbital energy level diagrams of these complexes. The reaction energy for H{sub 2} addition to 1-X, leading to the cis,trans-(H){sub 2}IrL{sub 2}(CO)X (2-X) product, has been computed. After electron correlation effects are included (MP4(SDTQ)), the reaction enthalpy computed for 1-CI is {minus}18.4 kcal/mol (L = PH{sub 3}) as compared to a reported experimental value of {minus}14 kcal/mol (L = PPh{sub 3}). Compared with available experimental data, the electronic effects of L(L = PH{sub 3}, NH{sub 3}, or AsH{sub 3}) and X on the thermodynamics of the H{sub 2} addition reaction are accurately reproduced by the model calculations at all levels of theory (HF and MPn). Formation of the hypothetical products cis,trans- and trans,trans-(H){sub 2}IrL{sub 2}(CO)X(2-X and 3-X) (X = BH{sub 2}, NH{sub 2}, and PH{sub 2}) is used to demonstrate that {pi}-acceptor substituents promote the H{sub 2} addition reaction to 1-X while {pi}-donor substituents disfavor addition.

  9. Effect of Copper and Zinc on the Single Molecule Self-Affinity of Alzheimer's Amyloid-β Peptides.

    PubMed

    Hane, Francis T; Hayes, Reid; Lee, Brenda Y; Leonenko, Zoya

    2016-01-01

    The presence of trace concentrations of metallic ions, such as copper and zinc, has previously been shown to drastically increase the aggregation rate and neurotoxicity of amyloid-β (Aβ), the peptide implicated in Alzheimer's disease (AD). The mechanism of why copper and zinc accelerate Aβ aggregation is poorly understood. In this work, we use single molecule force spectroscopy (SMFS) to probe the kinetic and thermodynamic parameters (dissociation constant, Kd, kinetic dissociation rate, koff, and free energy, ΔG) of the dissociation of an Aβ dimer, the amyloid species which initiates the amyloid cascade. Our results show that nanomolar concentrations of copper do not change the single molecule affinity of Aβ to another Aβ peptide in a statistically significant way, while nanomolar concentrations of zinc decrease the affinity of Aβ-Aβ by an order of magnitude. This suggests that the binding of zinc ion to Aβ may interfere with the binding of Aβ-Aβ, leading to a lower self-affinity.

  10. Effect of Copper and Zinc on the Single Molecule Self-Affinity of Alzheimer’s Amyloid-β Peptides

    PubMed Central

    Hane, Francis T.; Hayes, Reid; Lee, Brenda Y.; Leonenko, Zoya

    2016-01-01

    The presence of trace concentrations of metallic ions, such as copper and zinc, has previously been shown to drastically increase the aggregation rate and neurotoxicity of amyloid-β (Aβ), the peptide implicated in Alzheimer’s disease (AD). The mechanism of why copper and zinc accelerate Aβ aggregation is poorly understood. In this work, we use single molecule force spectroscopy (SMFS) to probe the kinetic and thermodynamic parameters (dissociation constant, Kd, kinetic dissociation rate, koff, and free energy, ΔG) of the dissociation of an Aβ dimer, the amyloid species which initiates the amyloid cascade. Our results show that nanomolar concentrations of copper do not change the single molecule affinity of Aβ to another Aβ peptide in a statistically significant way, while nanomolar concentrations of zinc decrease the affinity of Aβ-Aβ by an order of magnitude. This suggests that the binding of zinc ion to Aβ may interfere with the binding of Aβ-Aβ, leading to a lower self-affinity. PMID:26808970

  11. Definition of the affinity of binding between human von Willebrand factor and coagulation factor VIII.

    PubMed

    Ganz, P R; Atkins, J S; Palmer, D S; Dudani, A K; Hashemi, S; Luison, F

    1991-10-15

    Factor VIII and von Willebrand factor are two plasma proteins essential for effective hemostasis. In vivo, they form a non-covalent complex whose association appears to be metal ion dependent. However, a precise definition of the nature of the molecular forces governing their association remains to be defined, as does their binding affinity. In this paper we have determined the dissociation constant and stoichiometry for Factor VIII binding to immobilized von Willebrand factor. The data demonstrate that these proteins interact saturably and with relatively high affinity. Computer assisted analyses of the Scatchard data favour a two site binding model. The higher affinity site was found to have a Kd of 62 (+/- 13) x 10(-12) M while that of the lower affinity site was 380 (+/- 92) x 10(-12) M. The density of Factor VIII binding sites (Bmax) present on von Willebrand factor was 31 (+/- 3) pM for the high affinity binding site and 46 (+/- 6) pM for the lower site, corresponding to a calculated Factor VIII: von Willebrand factor binding ratio of 1:33 and 1:23, respectively.

  12. On the binding affinity of macromolecular interactions: daring to ask why proteins interact

    PubMed Central

    Kastritis, Panagiotis L.; Bonvin, Alexandre M. J. J.

    2013-01-01

    Interactions between proteins are orchestrated in a precise and time-dependent manner, underlying cellular function. The binding affinity, defined as the strength of these interactions, is translated into physico-chemical terms in the dissociation constant (Kd), the latter being an experimental measure that determines whether an interaction will be formed in solution or not. Predicting binding affinity from structural models has been a matter of active research for more than 40 years because of its fundamental role in drug development. However, all available approaches are incapable of predicting the binding affinity of protein–protein complexes from coordinates alone. Here, we examine both theoretical and experimental limitations that complicate the derivation of structure–affinity relationships. Most work so far has concentrated on binary interactions. Systems of increased complexity are far from being understood. The main physico-chemical measure that relates to binding affinity is the buried surface area, but it does not hold for flexible complexes. For the latter, there must be a significant entropic contribution that will have to be approximated in the future. We foresee that any theoretical modelling of these interactions will have to follow an integrative approach considering the biology, chemistry and physics that underlie protein–protein recognition. PMID:23235262

  13. Affinity improvement by fine tuning of single-chain variable fragment against aflatoxin B1.

    PubMed

    Min, Won-Ki; Na, Kang-In; Yoon, Jung-Hyun; Heo, Yoon-Jee; Lee, Daesang; Kim, Sung-Gun; Seo, Jin-Ho

    2016-10-15

    Aflatoxin B1 (AFB1) produced in Aspergillus flavus is a major hepatocarcinogen found in foods and feed. For effective immunological detection of AFB1 at low concentrations, the development of high affinity antibody for AFB1 is required. Previously, an affinity-maturated single-chain variable fragment containing 6 mutations (scFv-M37) was isolated from an artificial mutagenic library, which showed a 9-fold higher affinity than its wild type scFv. In this study, the effect of the 6 mutated residues on the affinity improvement was characterized using surface plasmon resonance analysis, which identified a deleterious mutation (VH-A110T) located on a framework region of the scFv-M37. The back mutation of VH-A110T resulted in a 3.2-fold affinity improvement, which was attributed to decrease of dissociation rate constant (kd) in interaction between AFB1 and the back mutant scFv. The biophysical analyses using circular dichroism and gel filtration revealed that the back mutation of VH-A110T caused a subtle conformational change of the scFv toward tighter binding to AFB1.

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

    PubMed

    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.

  15. Direct simulation with vibration-dissociation coupling

    NASA Technical Reports Server (NTRS)

    Hash, David B.; Hassan, H. A.

    1992-01-01

    The majority of implementations of the Direct Simulation Monte Carlo (DSMC) method of Bird do not account for vibration-dissociation coupling. Haas and Boyd have proposed the vibrationally-favored dissociation model to accomplish this task. This model requires measurements of induction distance to determine model constants. A more general expression has been derived that does not require any experimental input. The model is used to calculate one-dimensional shock waves in nitrogen and the flow past a lunar transfer vehicle (LTV). For the conditions considered in the simulation, the influence of vibration-dissociation coupling on heat transfer in the stagnation region of the LTV can be significant.

  16. Direct detection of thrombin binding to 8-bromodeoxyguanosine-modified aptamer: effects of modification on affinity and kinetics.

    PubMed

    Goji, Shou; Matsui, Jun

    2011-01-01

    The affinity of an 8-bromodeoxyguanosine- (8-BrdG-) substituted thrombin-binding aptamer (TBA-Br), which has the 1st and 10th guanosine residues replaced with 8-BrdG, was estimated using reflectometric interference spectroscopy (RIfS). When comparing TBA-Br with unmodified TBA (TBA-H), it was demonstrated that the modification effectively improved the affinity of TBA; dissociation constants (K(D)) of TBA-H and TBA-Br were 45.4 nM and 1.99 nM, respectively. These values, which were obtained by direct observation of thrombin binding using RIfS, have the same order of magnitude as those obtained in our previous study utilizing conformational changes in TBA to detect thrombin binding, thus confirming the validity of the obtained K(D) values. RIfS measurements also revealed that the 8-BrdG modification resulted in a lower dissociation rate constant (k(d)), which suggests that the enhancement of affinity can be attributed to the stabilization of the G-quadruplex structure on introduction of 8-BrdG.

  17. Metal-Mediated Affinity and Orientation Specificity in a Computationally Designed Protein Homodimer

    SciTech Connect

    Der, Bryan S.; Machius, Mischa; Miley, Michael J.; Mills, Jeffrey L.; Szyperski, Thomas; Kuhlman, Brian

    2015-10-15

    Computationally designing protein-protein interactions with high affinity and desired orientation is a challenging task. Incorporating metal-binding sites at the target interface may be one approach for increasing affinity and specifying the binding mode, thereby improving robustness of designed interactions for use as tools in basic research as well as in applications from biotechnology to medicine. Here we describe a Rosetta-based approach for the rational design of a protein monomer to form a zinc-mediated, symmetric homodimer. Our metal interface design, named MID1 (NESG target ID OR37), forms a tight dimer in the presence of zinc (MID1-zinc) with a dissociation constant <30 nM. Without zinc the dissociation constant is 4 {micro}M. The crystal structure of MID1-zinc shows good overall agreement with the computational model, but only three out of four designed histidines coordinate zinc. However, a histidine-to-glutamate point mutation resulted in four-coordination of zinc, and the resulting metal binding site and dimer orientation closely matches the computational model (C{alpha} rmsd = 1.4 {angstrom}).

  18. Protein separation using affinity-based reversed micelles

    PubMed

    Sun; Gu; Tong; Bai; Ichikawa; Furusaki

    1999-05-01

    Reversed micellar two-phase extraction is a developing technique for protein separation. Introduction of an affinity ligand is considered to be an effective approach to increase the selectivity and capacity of reversed micelles. In this article, Cibacron Blue F3G-A (CB) as an affinity ligand was immobilized to reversed micelles composed of soybean lecithin by a two-phase reaction. The affinity partitioning of lysozyme and bovine serum albumin (BSA) to the CB-lecithin micelles was studied. Formation of mixed micelles by additionally introducing a nonionic surfactant, Tween 85, to the CB-lecithin micelles was effective to increase the solubilization of lysozyme due to the increase of W0 (water/surfactant molar ratio)/micellar size. The partitioning isotherms of lysozyme to the CB-lecithin micelles with and without Tween 85 were expressed by the Langmuir equation. The dissociation constants in the Langmuir equation decreased on addition of Tween 85, indicating the increase of the effectiveness of lysozyme binding to the immobilized CB. On addition of 20 g/L Tween 85 to 50 g/L lecithin/hexane micellar phase containing 0.1 mmol/L CB, the extraction capacity for lysozyme could be increased by 42%. Moreover, the CB-lecithin micelles with or without Tween 85 showed significant size exclusion for BSA due to its high molecular weight. Thus, lysozyme and BSA were separated from artificial solutions containing the two proteins. In addition, the affinity-based reversed micellar phase containing Tween 85 was recycled three times for lysozyme purification from crude egg-white solutions. Lysozyme purity increased by 16-18-fold, reaching 60-70% in the recycled use.

  19. Phosphatidylserine Reversibly Binds Cu2+ with Extremely High Affinity

    PubMed Central

    Monson, Christopher F.; Cong, Xiao; Robison, Aaron; Pace, Hudson P.; Liu, Chunming; Poyton, Matthew F.; Cremer, Paul S.

    2012-01-01

    Phosphatidylserine (PS) embedded within supported lipid bilayers (SLBs) was found to bind Cu2+ from solution with extraordinarily high affinity. In fact, the equilibrium dissociation constant was in the femtomolar range. The resulting complex formed in a 1:2 Cu2+ to PS ratio and quenches a broad spectrum of lipid-bound fluorophores in a reversible and pH-dependent fashion. At acidic pH values, the fluorophores were almost completely unquenched, while at basic pH values significant quenching (85–90%) was observed. The pH at which the transition occurred was dependent on the PS concentration and ranged from approximately pH 5 to 8. The quenching kinetics was slow at low Cu2+ concentrations and basic values pH (up to several hours), while the unquenching reaction was orders of magnitude more rapid upon lowering the pH. This was consistent with diffusion limited complex formation at basic pH, but rapid dissociation under acidic conditions. The tight binding of Cu2+ to PS may have physiological consequences under certain circumstances. PMID:22548290

  20. Spectroscopic constants and potential energy curve of the iodine weakly bound 1u state correlating with the I(2P1/2) + I(2P1/2) dissociation limit

    NASA Astrophysics Data System (ADS)

    Akopyan, M. E.; Baturo, V. V.; Lukashov, S. S.; Poretsky, S. A.; Pravilov, A. M.

    2015-01-01

    The stepwise three-step three-color laser population of the I2(β1g, νβ, Jβ) rovibronic states via the B0u+, νB, JB rovibronic states and rovibronic levels of the 1u(bb) and 0g+(bb) states mixed by hyperfine interaction is used for determination of rovibronic level energies of the weakly bound I2(1u(bb)) state. Dunham coefficients of the state, Yi0 (i = 0-3), Yi1 (i = 0-2), Y02 and Y12 for the {{v}{{1u}}} = 1-5, 8, 10, 15 and {{J}{{1u}}} ≈ 9-87 ranges, the dissociation energy of the state, De, and equilibrium I-I distance, Re, as well as the potential energy curve are determined. There are aperiodicities in the excitation spectrum corresponding to the β, νβ = 23, Jβ ← 1u(bb), ν1u = 4, 5, J1u progressions in the I2 + Rg = He, Ar mixture, namely, a great number of lines which do not coincide with the R or P line progressions. Their positions conflict with the ΔJ-even selection rule. Furthermore, they do not correspond to the ΔJ-odd progression.

  1. In vitro and in vivo properties of (/sup 125/I) (R,S) 4IQNB: A lower affinity diastereomeric muscarinic receptor radiotracer

    SciTech Connect

    Gibson, R.E.; Schneidau, T.A.; Rzeszotarski, W.J.; Cohen, V.I.; Eckelman, W.C.; Reba, R.C.

    1985-05-01

    The (R,R) diastereomer of 3-Quinuclidinyl 4-Iodobenzilate (4IQNB) is a high affinity muscarinic acetylcholine receptor radiotracer which has provided images of receptor distribution in the CNS of man. The radiotracer is of such high affinity that dissociation in vivo is not evident in man after 6-half-lives I-123. Since the dissociation kinetics of radiotracer may be helpful for receptor quantitation, the authors have prepared (/sup 125/I) (R,S) 4IQNB: a diastereomer of 4IQNB which as a lower affinity for the m-AChR than the (R,R) isomer. The equilibrium association constant for the (R,S) diastereomer is 1.10 x 10/sup 9/ M/sup -1/, which is 4-fold lower in affinity than (/sup 3/H) (R) QNB and 2-fold lower than that of the (R,R) 4IQNB. Of more interest, the dissociation rate constant of (R,S) 4IQNB is 0.099 (+0.01)/min., 15-fold more rapid than that of the (R,R) isomer. The systemic distribution of (R,S) 4IQNB is similar to that of (R,R) 4IQNB except localization in the myocardium is 2-fold lower, reflecting the lower affinity. Nonreceptor interactions are the same since the compounds differ only as optical isomers around the carbinol chiral center. In the CNS peak activities are obtained in the corpus striatum (and other M/sub 1/-receptor rich structures) which are the same as obtained with (R,R) 4IQNB. However, no washout of (R,R) 4IQNB is observed after 4 hrs and only 60% in 24 hrs. By contrast, 65% of (R,S) 4IQNB washes out in 4 hrs and no significant activity is detected after 24 hrs. The increased washout kinetics should provide a better radiotracer for determining muscarinic receptor concentrations in the CNS of man.

  2. Protein A affinity precipitation of human immunoglobulin G.

    PubMed

    Janoschek, Lars; Freiherr von Roman, Matthias; Berensmeier, Sonja

    2014-08-15

    The potential of protein A affinity precipitation as an alternative method for traditional antibody purification techniques was investigated. Recombinant produced protein A from Staphylococcus aureus (SpA) was covalently linked to the pH-responsive copolymer Eudragit(®) S-100 and used for purification of human immunoglobulin G (hIgG). The Eudragit-SpA conjugate had a static binding capacity of 93.9 ± 2.8 mg hIgG per g conjugate and a dissociation constant of 787 ± 67 nM at 7 ± 1°C. The antibody was adsorbed rapidly onto Eudragit-SpA and reached equilibrium within 5 min. An excess of hIgG binding sites, provided by the conjugate, as well as adjusted elution conditions resulted in an appropriate hIgG purification performance. In summary, Eudragit-SpA was successfully applied to capture hIgG from a protein mixture with 65% antibody yield in the elution step. Nearly 96% purity and a purification factor of 12.4 were achieved. The Eudragit-SpA conjugate showed a stable ligand density over several cycles, which enabled reusability for repeated precipitation of hIgG. According to this, pH induced affinity precipitation can be seen as a potential alternative for protein A chromatography in antibody purification processes.

  3. Guilt by dissociation: guilt primes augment the relationship between dissociative tendencies and state dissociation.

    PubMed

    Rugens, Alex; Terhune, Devin Blair

    2013-03-30

    We examined the influence of guilt on the relationship between dissociative tendencies and state dissociation during mirror-gazing in a non-clinical sample. Dissociative tendencies correlated with state dissociation following guilt primes, but not after negative or neutral primes. This suggests that guilt augments the relationship between dissociative tendencies and state dissociation.

  4. High affinity truncated DNA aptamers for the development of fluorescence based progesterone biosensors.

    PubMed

    Alhadrami, Hani A; Chinnappan, Raja; Eissa, Shimaa; Rahamn, Anas Abdel; Zourob, Mohammed

    2017-02-24

    Aptamers have shown a number of potential applications in sensing and therapeutic due to the high affinity and specificity towards their target molecules. Not all the nucleotides in the full length aptamers are involved in the binding with their targets. The non-binding domain of the aptamer may affect the binding affinity of the aptamer-target complex. Mapping the aptamer binding region could increase the affinity and the specificity. In this paper, we designed aptamer-based fluorescence sensors from a truncated progesterone (P4) aptamer. Then, fluorescein and quencher labelled aptamer complementary oligonucleotide sequences were hybridized to the truncated aptamer at different sites to form duplex structures. We used fluorescence-quencher pair displacement assay upon progesterone binding for the determination of P4. One of the truncated sequences has shown high binding affinity with 16 fold increase in the dissociation constant, Kd (2.1 nM) compared to the original aptamer. The aptasensor was highly selective for P4 against similar compounds such as 17-β estradiol, bisphenol-A and vitamin D. The sensor has been applied for the detection of P4 in spiked tap water and in urine samples showing good recovery. This new developed aptamer-based fluorescence biosensor can be applied in food, pharmaceutical, and clinical industries.

  5. Are Fundamental Constants Really Constant?

    ERIC Educational Resources Information Center

    Swetman, T. P.

    1972-01-01

    Dirac's classical conclusions, that the values of e2, M and m are constants and the quantity of G decreases with time. Evoked considerable interest among researchers and traces historical development by which further experimental evidence points out that both e and G are constant values. (PS)

  6. Conformational Destabilization of Immunoglobulin G Increases the Low pH Binding Affinity with the Neonatal Fc Receptor*

    PubMed Central

    Walters, Benjamin T.; Jensen, Pernille F.; Larraillet, Vincent; Lin, Kevin; Patapoff, Thomas; Schlothauer, Tilman; Rand, Kasper D.; Zhang, Jennifer

    2016-01-01

    Crystallographic evidence suggests that the pH-dependent affinity of IgG molecules for the neonatal Fc receptor (FcRn) receptor primarily arises from salt bridges involving IgG histidine residues, resulting in moderate affinity at mildly acidic conditions. However, this view does not explain the diversity in affinity found in IgG variants, such as the YTE mutant (M252Y,S254T,T256E), which increases affinity to FcRn by up to 10×. Here we compare hydrogen exchange measurements at pH 7.0 and pH 5.5 with and without FcRn bound with surface plasmon resonance estimates of dissociation constants and FcRn affinity chromatography. The combination of experimental results demonstrates that differences between an IgG and its cognate YTE mutant vary with their pH-sensitive dynamics prior to binding FcRn. The conformational dynamics of these two molecules are nearly indistinguishable upon binding FcRn. We present evidence that pH-induced destabilization in the CH2/3 domain interface of IgG increases binding affinity by breaking intramolecular H-bonds and increases side-chain adaptability in sites that form intermolecular contacts with FcRn. Our results provide new insights into the mechanism of pH-dependent affinity in IgG-FcRn interactions and exemplify the important and often ignored role of intrinsic conformational dynamics in a protein ligand, to dictate affinity for biologically important receptors. PMID:26627822

  7. Theoretical proton affinity and fluoride affinity of nerve agent VX.

    PubMed

    Bera, Narayan C; Maeda, Satoshi; Morokuma, Keiji; Viggiano, Al A

    2010-12-23

    Proton affinity and fluoride affinity of nerve agent VX at all of its possible sites were calculated at the RI-MP2/cc-pVTZ//B3LYP/6-31G* and RI-MP2/aug-cc-pVTZ//B3LYP/6-31+G* levels, respectively. The protonation leads to various unique structures, with H(+) attached to oxygen, nitrogen, and sulfur atoms; among which the nitrogen site possesses the highest proton affinity of -ΔE ∼ 251 kcal/mol, suggesting that this is likely to be the major product. In addition some H(2), CH(4) dissociation as well as destruction channels have been found, among which the CH(4) + [Et-O-P(═O)(Me)-S-(CH(2))(2)-N(+)(iPr)═CHMe] product and the destruction product forming Et-O-P(═O)(Me)-SMe + CH(2)═N(+)(iPr)(2) are only 9 kcal/mol less stable than the most stable N-protonated product. For fluoridization, the S-P destruction channel to give Et-O-P(═O)(Me)(F) + [S-(CH(2))(2)-N-(iPr)(2)](-) is energetically the most favorable, with a fluoride affinity of -ΔE ∼ 44 kcal. Various F(-) ion-molecule complexes are also found, with the one having F(-) interacting with two hydrogen atoms in different alkyl groups to be only 9 kcal/mol higher than the above destruction product. These results suggest VX behaves quite differently from surrogate systems.

  8. Omega-conotoxin GVIA binding to a high-affinity receptor in brain: characterization, calcium sensitivity, and solubilization

    SciTech Connect

    Wagner, J.A.; Snowman, A.M.; Biswas, A.; Olivera, B.M.; Snyder, S.H.

    1988-09-01

    We describe unique, high-affinity binding sites for omega(/sup 125/I)conotoxin GVIA in membranes from rat brain and rabbit sympathetic ganglia which appear to be primarily associated with N-type voltage-dependent calcium channels. The dissociation constant (KD) for the toxin in rat brain membranes is 60 pM. Physiologic extracellular concentrations of calcium inhibit toxin binding noncompetitively (IC50 = 0.2 mM). The regional distribution of the binding sites in rat brain differs markedly from that of dihydropyridine calcium antagonist receptors associated with L-type calcium channels. In detergent-solubilized brain membranes, toxin binding retains the same affinity, specificity, and ionic sensitivity as in particulate preparations.

  9. Automatic determination of ligand purity and apparent dissociation constant (K(app)) in Ca(2+)/Mg(2+) buffer solutions and the K(app) for Ca(2+)/Mg(2+) anion binding in physiological solutions from Ca(2+)/Mg(2+)-macroelectrode measurements.

    PubMed

    Kay, James W; Steven, Rachel J; McGuigan, John A S; Elder, Hugh Y

    2008-01-01

    Calibration of Ca(2+)/Mg(2+) macroelectrodes and flurochromes in the nmolar and mumolar range, respectively, require the use of buffer solutions. In these buffers the apparent dissociation constant (K(app)) has to be measured since calculation based on tabulated constants gives variable results. The ligand concentration [Ligand](T) has also to be estimated. The most accurate and general method for measuring both is the ligand optimisation method based on macroelectrode potential measurements, but this iterative method is time consuming, thus limiting its application. This paper describes an automatic program based on the method, which on entering the measured macroelectrode data calculates K(app), [Ligand](T) and the ionised concentration [X(2+)] within minutes. This optimisation method cannot be used at K(app) values greater than 0.1mM, but can be extended into this region if the anion concentration is known. The program has been modified to cover this eventuality. Ca(2+)/Mg(2+) macroelectrodes in conjunction with these programs offer an accurate, routine method for determining K(app) and [Ligand](T) in buffer solutions at the appropriate ionic strength, temperature and pH and the K(app) for divalent cations binding to physiological anions under experimental conditions.

  10. Zero-Point Energy Constraint for Unimolecular Dissociation Reactions. Giving Trajectories Multiple Chances To Dissociate Correctly.

    PubMed

    Paul, Amit K; Hase, William L

    2016-01-28

    A zero-point energy (ZPE) constraint model is proposed for classical trajectory simulations of unimolecular decomposition and applied to CH4* → H + CH3 decomposition. With this model trajectories are not allowed to dissociate unless they have ZPE in the CH3 product. If not, they are returned to the CH4* region of phase space and, if necessary, given additional opportunities to dissociate with ZPE. The lifetime for dissociation of an individual trajectory is the time it takes to dissociate with ZPE in CH3, including multiple possible returns to CH4*. With this ZPE constraint the dissociation of CH4* is exponential in time as expected for intrinsic RRKM dynamics and the resulting rate constant is in good agreement with the harmonic quantum value of RRKM theory. In contrast, a model that discards trajectories without ZPE in the reaction products gives a CH4* → H + CH3 rate constant that agrees with the classical and not quantum RRKM value. The rate constant for the purely classical simulation indicates that anharmonicity may be important and the rate constant from the ZPE constrained classical trajectory simulation may not represent the complete anharmonicity of the RRKM quantum dynamics. The ZPE constraint model proposed here is compared with previous models for restricting ZPE flow in intramolecular dynamics, and connecting product and reactant/product quantum energy levels in chemical dynamics simulations.

  11. Determination of the constants of affinity of FeCl3, CuCl2, and ZnCl2 for a nitrogen-containing organosilane bonded on Al2O3-cellulose acetate hybrid material surface from ethanol solution.

    PubMed

    Lazarin, Angélica M; Landers, Richard; Kholin, Yuriy V; Gushikem, Yoshitaka

    2002-10-01

    This work describes the preparation and characterization of a cellulose acetate fiber coated with Al(2)O(3), resulting in the organic-inorganic hybrid Cel/Al(2)O(3). Furthermore, the hybrid was modified by attaching organofunctional groups by reaction with the precursor reagents (RO)(3)Si(CH(2))(3)L (L=NH(2), NH(CH(2))(2)NH(2), NH(CH(2))(2)NH(CH(2))(2)NH(2), and N(2)C(3)H(3) (imidazole)), resulting in Cel/Al(2)O(3)/Si(CH(2))(3)NH(2) (1), Cel/Al(2)O(3)/Si(CH(2))(3)NH(CH(2))(2)NH(2) (2), Cel/Al(2)O(3)/Si(CH(2))(3)NH(CH(2))(2)NH(CH(2))(2)NH(2) (3), and Cel/Al(2)O(3)/Si(CH(2))(3)N(2)C(3)H(3) (4). The amounts of attached organofunctional groups were (in mmol per gram of the material) 1=1.90, 2=1.89, 3=1.66, and 4=1.35. The isotherms of adsorption of FeCl(3), CuCl(2), and ZnCl(2) by Cel/Al(2)O(3)/Si(CH(2))(3)L from ethanol solutions were obtained at 298 K. Accurate estimates of the specific sorption capacities and the heteregeneous stability constants of the immobilized metal complexes were determined with the aid of several computational procedures. It is shown that the sorptional capacities are much less than the concentrations of the attached organofunctional groups. As all sorption isotherms are fitted properly with the Langmuir isotherm equation, the effects of the energetic heterogeneity and the lateral interactions do not affect the chemisorption equilibria. The heterogeneous stability constants of the immobilized complexes are fairly high, which provides efficient removal of the metal ions from solutions by the hybrid materials.

  12. Tuning the Binding Affinities and Reversion Kinetics of a Light Inducible Dimer Allows Control of Transmembrane Protein Localization.

    PubMed

    Zimmerman, Seth P; Hallett, Ryan A; Bourke, Ashley M; Bear, James E; Kennedy, Matthew J; Kuhlman, Brian

    2016-09-20

    Inducible dimers are powerful tools for controlling biological processes through colocalizing signaling molecules. To be effective, an inducible system should have a dissociation constant in the "off" state that is greater (i.e., weaker affinity) than the concentrations of the molecules that are being controlled, and in the "on" state a dissociation constant that is less (i.e., stronger affinity) than the relevant protein concentrations. Here, we reengineer the interaction between the light inducible dimer, iLID, and its binding partner SspB, to better control proteins present at high effective concentrations (5-100 μM). iLID contains a light-oxygen-voltage (LOV) domain that undergoes a conformational change upon activation with blue light and exposes a peptide motif, ssrA, that binds to SspB. The new variant of the dimer system contains a single SspB point mutation (A58V), and displays a 42-fold change in binding affinity when activated with blue light (from 3 ± 2 μM to 125 ± 40 μM) and allows for light-activated colocalization of transmembrane proteins in neurons, where a higher affinity switch (0.8-47 μM) was less effective because more colocalization was seen in the dark. Additionally, with a point mutation in the LOV domain (N414L), we lengthened the reversion half-life of iLID. This expanded suite of light induced dimers increases the variety of cellular pathways that can be targeted with light.

  13. A novel technique for measurement of thermal rate constants and temperature dependences of dissociative recombination: CO2+, CF3+, N2O+, C7H8+, C7H7+, C6H6+, C6H5+, C5H6+, C4H4+, and C3H3+

    NASA Astrophysics Data System (ADS)

    Fournier, Joseph A.; Shuman, Nicholas S.; Melko, Joshua J.; Ard, Shaun G.; Viggiano, Albert A.

    2013-04-01

    A novel technique using a flowing afterglow-Langmuir probe apparatus for measurement of temperature dependences of rate constants for dissociative recombination (DR) is presented. Low (˜1011 cm-3) concentrations of a neutral precursor are added to a noble gas/electron afterglow plasma thermalized at 300-500 K. Charge exchange yields one or many cation species, each of which may undergo DR. Relative ion concentrations are monitored at a fixed reaction time while the initial plasma density is varied between 109 and 1010 cm-3. Modeling of the decrease in concentration of each cation relative to the non-recombining noble gas cation yields the rate constant for DR. The technique is applied to several species (O2+, CO2+, CF3+, N2O+) with previously determined 300 K values, showing excellent agreement. The measurements of those species are extended to 500 K, with good agreement to literature values where they exist. Measurements are also made for a range of CnHm+ (C7H7+, C7H8+, C5H6+, C4H4+, C6H5+, C3H3+, and C6H6+) derived from benzene and toluene neutral precursors. CnHm+ DR rate constants vary from 8-12 × 10-7 cm3 s-1 at 300 K with temperature dependences of approximately T-0.7. Where prior measurements exist these results are in agreement, with the exception of C3H3+ where the present results disagree with a previously reported flat temperature dependence.

  14. A novel technique for measurement of thermal rate constants and temperature dependences of dissociative recombination: CO2(+), CF3(+), N2O(+), C7H8(+), C7H7(+), C6H6(+), C6H5(+), C5H6(+), C4H4(+), and C3H3(+).

    PubMed

    Fournier, Joseph A; Shuman, Nicholas S; Melko, Joshua J; Ard, Shaun G; Viggiano, Albert A

    2013-04-21

    A novel technique using a flowing afterglow-Langmuir probe apparatus for measurement of temperature dependences of rate constants for dissociative recombination (DR) is presented. Low (~10(11) cm(-3)) concentrations of a neutral precursor are added to a noble gas∕electron afterglow plasma thermalized at 300-500 K. Charge exchange yields one or many cation species, each of which may undergo DR. Relative ion concentrations are monitored at a fixed reaction time while the initial plasma density is varied between 10(9) and 10(10) cm(-3). Modeling of the decrease in concentration of each cation relative to the non-recombining noble gas cation yields the rate constant for DR. The technique is applied to several species (O2(+), CO2(+), CF3(+), N2O(+)) with previously determined 300 K values, showing excellent agreement. The measurements of those species are extended to 500 K, with good agreement to literature values where they exist. Measurements are also made for a range of CnHm(+) (C7H7(+), C7H8(+), C5H6(+), C4H4(+), C6H5(+), C3H3(+), and C6H6(+)) derived from benzene and toluene neutral precursors. CnHm(+) DR rate constants vary from 8-12 × 10(-7) cm(3) s(-1) at 300 K with temperature dependences of approximately T(-0.7). Where prior measurements exist these results are in agreement, with the exception of C3H3(+) where the present results disagree with a previously reported flat temperature dependence.

  15. Narrative, dialogue, and dissociation.

    PubMed

    Gedo, Paul M

    2014-02-01

    This paper explores dissociative phenomena as disruptions of dialogue between persons, and disruptions of internal narratives. A dissociating patient temporarily loses ability to convey his or her inner experience to the therapist. The disconnection between dialogue and internal experience can mislead both participants, or distract them from underlying connotations. Dissociation also disrupts the patient's sense of internal coherence and internal conversation. Dissociation represents a regression to an early, preverbal mode of (internal and external) communication. The challenge for the dyad is to restore dialogue and then to discern the multiply determined meanings of the dissociative communication. This therapeutic work allows the patient to achieve a more coherent sense of self and of his or her life course.

  16. Dissociative recombination in aeronomy

    NASA Technical Reports Server (NTRS)

    Fox, J. L.

    1989-01-01

    The importance of dissociative recombination in planetary aeronomy is summarized, and two examples are discussed. The first is the role of dissociative recombination of N2(+) in the escape of nitrogen from Mars. A previous model is updated to reflect new experimental data on the electronic states of N produced in this process. Second, the intensity of the atomic oxygen green line on the nightside of Venus is modeled. Use is made of theoretical rate coefficients for production of O (1S) in dissociative recombination from different vibrational levels of O2(+).

  17. Dissociative Identity Disorder

    PubMed Central

    2009-01-01

    A brief description of the controversies surrounding the diagnosis of dissociative identity disorder is presented, followed by a discussion of the proposed similarities and differences between dissociative identity disorder and borderline personality disorder. The phenomenon of autohypnosis in the context of early childhood sexual trauma and disordered attachment is discussed, as is the meaning of alters or alternate personalities. The author describes recent neurosciences research that may relate the symptoms of dissociative identity disorder to demonstrable disordered attention and memory processes. A clinical description of a typical patient presentation is included, plus some recommendations for approaches to treatment. PMID:19724751

  18. Enzyme activity assay of glycoprotein enzymes based on a boronate affinity molecularly imprinted 96-well microplate.

    PubMed

    Bi, Xiaodong; Liu, Zhen

    2014-12-16

    Enzyme activity assay is an important method in clinical diagnostics. However, conventional enzyme activity assay suffers from apparent interference from the sample matrix. Herein, we present a new format of enzyme activity assay that can effectively eliminate the effects of the sample matrix. The key is a 96-well microplate modified with molecularly imprinted polymer (MIP) prepared according to a newly proposed method called boronate affinity-based oriented surface imprinting. Alkaline phosphatase (ALP), a glycoprotein enzyme that has been routinely used as an indicator for several diseases in clinical tests, was taken as a representative target enzyme. The prepared MIP exhibited strong affinity toward the template enzyme (with a dissociation constant of 10(-10) M) as well as superb tolerance for interference. Thus, the enzyme molecules in a complicated sample matrix could be specifically captured and cleaned up for enzyme activity assay, which eliminated the interference from the sample matrix. On the other hand, because the boronate affinity MIP could well retain the enzymatic activity of glycoprotein enzymes, the enzyme captured by the MIP was directly used for activity assay. Thus, additional assay time and possible enzyme or activity loss due to an enzyme release step required by other methods were avoided. Assay of ALP in human serum was successfully demonstrated, suggesting a promising prospect of the proposed method in real-world applications.

  19. Application of Frontal Affinity Chromatography to Study the Biomolecular Interactions with Trypsin.

    PubMed

    Hu, YuanYuan; Qian, Junqing; Guo, Hui; Jiang, ShengLan; Zhang, Zheng

    2015-07-01

    Trypsin is a serine protease that has been proposed as a potential therapeutic target for metabolic disorders and malignancy diseases, thus the identification of biomolecular interactions of compounds to trypsin could be of great therapeutic importance. In this study, trypsin was immobilized on a monolithic silica capillary column via sol-gel. The binding properties of four small molecules (daidzin, genistin, matrine and oxymatrine) to trypsin were examined using the trypsin affinity columns by frontal analysis. The results indicate that the matrine (dissociation constant, Kd = 7.904 μM) has stronger interaction with trypsin than the oxymatrine (Kd = 8.204 μM), whereas daidzin and genistin were nearly have no affinity with trypsin. The results demonstrated that the frontal affinity chromatography can be used for the direct determination of protein-protease inhibitor binding interactions and have several significant advantages, including easy fabricating, reproducible, minimal technological requirements and potential to become a reliable alternative for quantitative studies of biomolecular interactions.

  20. Antiparkinson therapeutic potencies correlate with their affinities at dopamine D2(High) receptors.

    PubMed

    Seeman, Philip

    2007-12-01

    To determine whether antiparkinson dopamine agonists preferentially act on the high-affinity or the low-affinity states of dopamine D1 and D2 receptors, the agonist potencies were obtained by competition against [(3)H]SCH23390 for D1(High) and D1(Low), and against [(3)H]domperidone for D2(High) and D2(Low). N-propylnorapomorphine and cabergoline were the most potent at D2(High), with dissociation constants of 0.18 and 0.36 nM, respectively. Other agonists had D2(High)K(i) values of 0.52 nM for quinagolide, 0.6 nM for (+)PHNO, 0.9 for bromocriptine, 1.8 nM for apomorphine, 2.4 nM for pergolide, 3 nM for quinpirole, and 6.2 nM for lergotrile. There was a clear correlation between the K(i) values at D2(High) and their therapeutic concentrations in the plasma water, as derived from the known concentrations after correction for the fraction bound to the human plasma proteins. The data suggest that D2(High) is the primary and common target for the antiparkinson action of dopamine agonists. Bromocriptine, cabergoline, lergotrile, pergolide, and pramipexole had no affinity for D1(High), consistent with the clinical observations that the D2-selective bromocriptine and pramipexole elicit low levels of dyskinesia.

  1. Conformational kinetics reveals affinities of protein conformational states.

    PubMed

    Daniels, Kyle G; Suo, Yang; Oas, Terrence G

    2015-07-28

    Most biological reactions rely on interplay between binding and changes in both macromolecular structure and dynamics. Practical understanding of this interplay requires detection of critical intermediates and determination of their binding and conformational characteristics. However, many of these species are only transiently present and they have often been overlooked in mechanistic studies of reactions that couple binding to conformational change. We monitored the kinetics of ligand-induced conformational changes in a small protein using six different ligands. We analyzed the kinetic data to simultaneously determine both binding affinities for the conformational states and the rate constants of conformational change. The approach we used is sufficiently robust to determine the affinities of three conformational states and detect even modest differences in the protein's affinities for relatively similar ligands. Ligand binding favors higher-affinity conformational states by increasing forward conformational rate constants and/or decreasing reverse conformational rate constants. The amounts by which forward rate constants increase and reverse rate constants decrease are proportional to the ratio of affinities of the conformational states. We also show that both the affinity ratio and another parameter, which quantifies the changes in conformational rate constants upon ligand binding, are strong determinants of the mechanism (conformational selection and/or induced fit) of molecular recognition. Our results highlight the utility of analyzing the kinetics of conformational changes to determine affinities that cannot be determined from equilibrium experiments. Most importantly, they demonstrate an inextricable link between conformational dynamics and the binding affinities of conformational states.

  2. Have many estimates of efficacy and affinity been misled? Revisiting the operational model of agonism.

    PubMed

    Roche, David; van der Graaf, Piet H; Giraldo, Jesús

    2016-11-01

    The operational model of agonism offers a general equation to account for steep or flat functional curves by including a slope parameter different from 1. However, because this equation is not a Hill equation, those steep or flat experimental curves that follow the Hill model are excluded from the operational framework. This conceptual omission could have significant consequences in the estimation of affinity and efficacy - the operational model tends to overestimate agonist-receptor dissociation constants and operational efficacy parameters to accommodate the shape of theoretical curves to steep or flat experimental Hill curves. To avoid misled parameter estimates for an ample space of pharmacological data a new version of the operational model has been developed.

  3. Determination of acidity constants of acid-base indicators by second-derivative spectrophotometry

    NASA Astrophysics Data System (ADS)

    Kara, Derya; Alkan, Mahir

    2000-12-01

    A method for calculation of acid-base dissociation constants of monoprotic weak organic acids whose acid and base species have overlapping spectra from absorptiometric and pH measurements is described. It has been shown that the second-derivative spectrophotometry can effectively be used for determining the dissociation constants, when dissociation constants obtained for methyl orange and bromothymol blue were compared with the values given in the literature.

  4. Dissociative disorders in medical settings.

    PubMed

    MacPhee, Edward

    2013-10-01

    Despite the challenges of conducting research on dissociation and the dissociative disorders, our understanding has grown greatly over the past three decades, including our knowledge of the often overlooked sensorimotor manifestations of dissociation, more commonly referred to as somatoform dissociation. This article will first review the definitions and presentations of dissociation in general along with recent research on the concept of somatoform dissociation. Then, each of the dissociative disorders and conversion disorder will be discussed in further detail as well as how they might present in a medical setting. Current recommendations for diagnosis and treatment will also be provided.

  5. Protection against anthrax toxin by recombinant antibody fragments correlates with antigen affinity.

    PubMed

    Maynard, Jennifer A; Maassen, Catharina B M; Leppla, Stephen H; Brasky, Kathleen; Patterson, Jean L; Iverson, Brent L; Georgiou, George

    2002-06-01

    The tripartite toxin produced by Bacillus anthracis is the key determinant in the etiology of anthrax. We have engineered a panel of toxin-neutralizing antibodies, including single-chain variable fragments (scFvs) and scFvs fused to a human constant kappa domain (scAbs), that bind to the protective antigen subunit of the toxin with equilibrium dissociation constants (K(d)) between 63 nM and 0.25 nM. The entire antibody panel showed high serum, thermal, and denaturant stability. In vitro, post-challenge protection of macrophages from the action of the holotoxin correlated with the K(d) of the scFv variants. Strong correlations among antibody construct affinity, serum half-life, and protection were also observed in a rat model of toxin challenge. High-affinity toxin-neutralizing antibodies may be of therapeutic value for alleviating the symptoms of anthrax toxin in infected individuals and for medium-term prophylaxis to infection.

  6. Optimal Affinity of a Monoclonal Antibody: Guiding Principles Using Mechanistic Modeling.

    PubMed

    Tiwari, Abhinav; Abraham, Anson K; Harrold, John M; Zutshi, Anup; Singh, Pratap

    2017-03-01

    Affinity optimization of monoclonal antibodies (mAbs) is essential for developing drug candidates with the highest likelihood of clinical success; however, a quantitative approach for setting affinity requirements is often lacking. In this study, we computationally analyzed the in vivo mAb-target binding kinetics to delineate general principles for defining optimal equilibrium dissociation constant ([Formula: see text]) of mAbs against soluble and membrane-bound targets. Our analysis shows that in general [Formula: see text] to achieve 90% coverage for a soluble target is one tenth of its baseline concentration ([Formula: see text]), and is independent of the dosing interval, target turnover rate or the presence of competing ligands. For membrane-bound internalizing targets, it is equal to the ratio of internalization rate of mAb-target complex and association rate constant ([Formula: see text]). In cases where soluble and membrane-bound forms of the target co-exist, [Formula: see text] lies within a range determined by the internalization rate ([Formula: see text]) of the mAb-membrane target complex and the ratio of baseline concentrations of soluble and membrane-bound forms ([Formula: see text]). Finally, to demonstrate practical application of these general rules, we collected target expression and turnover data to project [Formula: see text] for a number of marketed mAbs against soluble (TNFα, RANKL, and VEGF) and membrane-bound targets (CD20, EGFR, and HER2).

  7. Dissociation of diatomic gases

    NASA Technical Reports Server (NTRS)

    Hansen, C. F.

    1991-01-01

    The Landau-Zener theory of reactive cross sections has been applied to diatomic molecules dissociating from a ladder of rotational and vibrational states. Although the preexponential factor of the Arrhenius rate expression is shown to be a complex function of the dimensionless activation energy, the average over all states in the ladder is well represented by a single factor that varies about as T exp (-n), where the coefficient n is the order of unity. This relation agrees very well with experimental data for dissociation of O2 and N2, for example. The results validate previous empirical assignment of a single preexponential factor in the Arrhenius expression and justify the extrapolation of the expression well beyond the range of data. The theory is then used to calculate the effect of vibrational nonequilibrium on dissociation rate. For Morse oscillators the results are about the same as for harmonic oscillators, and the dissociation from a ladder of equilibrium rotational and nonequilibrium vibrational states is close to an analytic approximation provided by Hammerling, Kivel, and Teare for harmonic oscillators all dissociating from the ground rotational state.

  8. Limited proteolysis for assaying ligand binding affinities of nuclear receptors.

    PubMed

    Benkoussa, M; Nominé, B; Mouchon, A; Lefebvre, B; Bernardon, J M; Formstecher, P; Lefebvre, P

    1997-01-01

    The binding of natural or synthetic ligands to nuclear receptors is the triggering event leading to gene transcription activation or repression. Ligand binding to the ligand binding domain of these receptors induces conformational changes that are evidenced by an increased resistance of this domain to proteases. In vitro labeled receptors were incubated with various synthetic or natural agonists or antagonists and submitted to trypsin digestion. Proteolysis products were separated by SDS-PAGE and quantified. The amount of trypsin-resistant fragments was proportional to receptor occupancy by the ligand, and allowed the determination of dissociation constants (kDa). Using the wild-type or mutated human retinoic acid receptor alpha as a model, kDa values determined by classical competition binding assays using tritiated ligands are in agreement with those measured by the proteolytic assay. This method was successfully extended to human retinoic X receptor alpha, glucocorticoid receptor, and progesterone receptor, thus providing a basis for a new, faster assay to determine simultaneously the affinity and conformation of receptors when bound to a given ligand.

  9. Communication: Revised electron affinity of SF6 from kinetic data.

    PubMed

    Troe, Jürgen; Miller, Thomas M; Viggiano, Albert A

    2012-03-28

    Previously determined experimental data for thermal attachment of electrons to SF(6) and thermal detachment from SF(6)(-) over the range 590-670 K are reevaluated by a third-law analysis. Recent high precision calculations of SF(6)(-) harmonic frequences and anharmonicities (for several of the modes) lead to considerable changes in modeled vibrational partition functions which then have to be accommodated for by a smaller value of the derived adiabatic electron affinity EA of SF(6). The previously estimated value of EA = 1.20 (±0.05) eV in this way is reduced to a value of EA = 1.03 (±0.05) eV. In addition, the bond dissociation energy E(0,dis) for SF(6)(-) → SF(5)(-) + F is reduced to E(0,dis) = 1.44 (±0.05) eV. Finally, the consequences for modeled specific rate constants k(det)(E,J) of electron detachment from SF(6)(-) are discussed.

  10. Two high-affinity ligand binding states of uterine estrogen receptor distinguished by modulation of hydrophobic environment

    SciTech Connect

    Hutchens, T.W.; Li, C.M.; Zamah, N.M.; Besch, P.K.

    1987-02-10

    The steroid binding function of soluble (cytosolic) estrogen receptors from calf uteri was evaluated under conditions known to modify the extent of hydrophobic interaction with receptor-associated proteins. Receptor preparations were equilibrated into 6 M urea buffers and control buffers by chromatography through small columns of Sephadex G-25 or by dialysis at 0.6 /sup 0/C. Equilibrium dissociation constants (K/sub d/) and binding capacities (n) of experimental and control receptor preparations were determined by 13-point Scatchard analyses using concentrations of 17..beta..-(/sup 3/H)estradiol from 0.05 to 10 nM. Nonspecific binding was determined at each concentration by parallel incubations with a 200-fold molar excess of the receptor-specific competitor diethylstilbestrol. The control receptor population was consistently found to be a single class of binding sites with a high affinity for estradiol which was unaffected by G-25 chromatography, by dialysis, by dilution, or by the presence of 0.4 M KCl. However, equilibration into 6 M urea induced a discrete (10-fold) reduction in receptor affinity to reveal a second, thermodynamically stable, high-affinity binding state. The presence of 0.4 M KCl did not significantly influence the discrete change in receptor affinity induced by urea. The effects of urea on both receptor affinity and binding capacity were reversible, suggesting a lack of covalent modification. These results demonstrate nonenzymatic means by which not only the binding capacity but also the affinity of receptor for estradiol can be reversibly controlled, suggesting that high concentrations of urea might be more effectively utilized during the physicochemical characterization and purification of steroid receptor proteins.

  11. Psychophysiology of dissociated consciousness.

    PubMed

    Bob, Petr

    2014-01-01

    Recent study of consciousness provides an evidence that there is a limit of consciousness, which presents a barrier between conscious and unconscious processes. This barrier likely is specifically manifested as a disturbance of neural mechanisms of consciousness that through distributed brain processing, attentional mechanisms and memory processes enable to constitute integrative conscious experience. According to recent findings a level of conscious integration may change during certain conditions related to experimental cognitive manipulations, hypnosis, or stressful experiences that can lead to dissociation of consciousness. In psychopathological research the term dissociation was proposed by Pierre Janet for explanation of processes related to splitting of consciousness due to traumatic events or during hypnosis. According to several recent findings dissociation of consciousness likely is related to deficits in global distribution of information and may lead to heightened levels of "neural complexity" that reflects brain integration or differentiation based on numbers of independent neural processes in the brain that may be specifically related to various mental disorders.

  12. The Cutting Edge of Affinity Electrophoresis Technology

    PubMed Central

    Kinoshita, Eiji; Kinoshita-Kikuta, Emiko; Koike, Tohru

    2015-01-01

    Affinity electrophoresis is an important technique that is widely used to separate and analyze biomolecules in the fields of biology and medicine. Both quantitative and qualitative information can be gained through affinity electrophoresis. Affinity electrophoresis can be applied through a variety of strategies, such as mobility shift electrophoresis, charge shift electrophoresis or capillary affinity electrophoresis. These strategies are based on changes in the electrophoretic patterns of biological macromolecules that result from interactions or complex-formation processes that induce changes in the size or total charge of the molecules. Nucleic acid fragments can be characterized through their affinity to other molecules, for example transcriptional factor proteins. Hydrophobic membrane proteins can be identified by means of a shift in the mobility induced by a charged detergent. The various strategies have also been used in the estimation of association/disassociation constants. Some of these strategies have similarities to affinity chromatography, in that they use a probe or ligand immobilized on a supported matrix for electrophoresis. Such methods have recently contributed to profiling of major posttranslational modifications of proteins, such as glycosylation or phosphorylation. Here, we describe advances in analytical techniques involving affinity electrophoresis that have appeared during the last five years. PMID:28248262

  13. Modulating Uranium Binding Affinity in Engineered Calmodulin EF-Hand Peptides: Effect of Phosphorylation

    PubMed Central

    Pardoux, Romain; Sauge-Merle, Sandrine; Lemaire, David; Delangle, Pascale; Guilloreau, Luc; Adriano, Jean-Marc; Berthomieu, Catherine

    2012-01-01

    To improve our understanding of uranium toxicity, the determinants of uranyl affinity in proteins must be better characterized. In this work, we analyzed the contribution of a phosphoryl group on uranium binding affinity in a protein binding site, using the site 1 EF-hand motif of calmodulin. The recombinant domain 1 of calmodulin from A. thaliana was engineered to impair metal binding at site 2 and was used as a structured template. Threonine at position 9 of the loop was phosphorylated in vitro, using the recombinant catalytic subunit of protein kinase CK2. Hence, the T9TKE12 sequence was substituted by the CK2 recognition sequence TAAE. A tyrosine was introduced at position 7, so that uranyl and calcium binding affinities could be determined by following tyrosine fluorescence. Phosphorylation was characterized by ESI-MS spectrometry, and the phosphorylated peptide was purified to homogeneity using ion-exchange chromatography. The binding constants for uranyl were determined by competition experiments with iminodiacetate. At pH 6, phosphorylation increased the affinity for uranyl by a factor of ∼5, from Kd = 25±6 nM to Kd = 5±1 nM. The phosphorylated peptide exhibited a much larger affinity at pH 7, with a dissociation constant in the subnanomolar range (Kd = 0.25±0.06 nM). FTIR analyses showed that the phosphothreonine side chain is partly protonated at pH 6, while it is fully deprotonated at pH 7. Moreover, formation of the uranyl-peptide complex at pH 7 resulted in significant frequency shifts of the νas(P-O) and νs(P-O) IR modes of phosphothreonine, supporting its direct interaction with uranyl. Accordingly, a bathochromic shift in νas(UO2)2+ vibration (from 923 cm−1 to 908 cm−1) was observed upon uranyl coordination to the phosphorylated peptide. Together, our data demonstrate that the phosphoryl group plays a determining role in uranyl binding affinity to proteins at physiological pH. PMID:22870263

  14. Blackbody Infrared Radiative Dissociation of Protonated Oligosaccharides

    NASA Astrophysics Data System (ADS)

    Fentabil, Messele A.; Daneshfar, Rambod; Kitova, Elena N.; Klassen, John S.

    2011-12-01

    The dissociation pathways, kinetics, and energetics of protonated oligosaccharides in the gas phase were investigated using blackbody infrared radiative dissociation (BIRD). Time-resolved BIRD measurements were performed on singly protonated ions of cellohexaose (Cel6), which is composed of β-(1 → 4)-linked glucopyranose rings, and five malto-oligosaccharides (Malx, where x = 4-8), which are composed of α-(1 → 4)-linked glucopyranose units. At the temperatures investigated (85-160 °C), the oligosaccharides dissociate at the glycosidic linkages or by the loss of a water molecule to produce B- or Y-type ions. The Y ions dissociate to smaller Y or B ions, while the B ions yield exclusively smaller B ions. The sequential loss of water molecules from the smallest B ions (B1 and B2) also occurs. Rate constants for dissociation of the protonated oligosaccharides and the corresponding Arrhenius activation parameters (Ea and A) were determined. The Ea and A-factors measured for protonated Malx (x > 4) are indistinguishable within error (~19 kcal mol-1, 1010 s-1), which is consistent with the ions being in the rapid energy exchange limit. In contrast, the Arrhenius parameters for protonated Cel6 (24 kcal mol-1, 1012 s-1) are significantly larger. These results indicate that both the energy and entropy changes associated with the glycosidic bond cleavage are sensitive to the anomeric configuration. Based on the results of this study, it is proposed that formation of B and Y ions occurs through a common dissociation mechanism, with the position of the proton establishing whether a B or Y ion is formed upon glycosidic bond cleavage.

  15. Binding of L-(/sup 3/H)nicotine to a single class of high affinity sites in rat brain membranes

    SciTech Connect

    Lippiello, P.M.; Fernandes, K.G.

    1986-05-01

    The binding of optically pure L-(/sup 3/H)nicotine to rat brain membrane preparations was studied using a rapid filtration method. The binding properties observed depended on the method used for tissue isolation. The most consistent results were obtained with membranes prepared in the presence of protease inhibitors, without divalent cations. Binding was saturable, reversible, and stereospecific. Scatchard analysis revealed a single class of high affinity sites with an average KD of 2 nM and a Bmax of approximately 200 fmol/mg of protein. The Hill coefficient was near unity. The KD calculated from the kinetic rate constants for association (k1 = 0.012 min-1 nM-1) and dissociation (k-1 = 0.04 min-1) was around 3 nM, in good agreement with the dissociation constant determined from equilibrium binding. In competition studies, cholinergic agonists were generally the most effective in inhibiting L-(/sup 3/H)nicotine binding, whereas antagonists were relatively ineffective. The D-isomer of nicotine was about 60-fold less potent than the L-isomer in inhibiting binding. The results were unaffected by temperature, with the exception that Bmax was somewhat lower at 37 degrees. The equilibrium binding properties of these sites were essentially identical in adult male and female brain. However, Bmax was lower in fetal brain tissue. The present findings are consistent with the idea that there is a single class of high affinity nicotinic binding sites in rat brain with cholinoceptive properties.

  16. The dissociative recombination of ?

    NASA Astrophysics Data System (ADS)

    Laubé, S.; Lehfaoui, L.; Rowe, B. R.; Mitchell, J. B. A.

    1998-09-01

    The dissociative recombination rate coefficient for 0953-4075/31/18/016/img2 has been measured at 300 K using a flowing afterglow Langmuir probe-mass spectrometer apparatus. A value of 0953-4075/31/18/016/img3 has been found.

  17. Introduction to dissociative recombination

    NASA Technical Reports Server (NTRS)

    Guberman, Steven L.; Mitchell, J. Brian A.

    1989-01-01

    Dissociative recombination (DR) of molecular ions with electrons has important consequences in many areas of physical science. Ab-initio calculations coupled with resonant scattering theory and multichannel quantum defect studies have produced detailed results illuminating the role of ion vibrational excitation, the quantum yields of the DR products, and the role of Rydberg states. The theoretical and experimental results are discussed.

  18. Dissociative Identity Disorder

    ERIC Educational Resources Information Center

    Schmidt, Tom

    2007-01-01

    Few psychological disorders in the Diagnostic Statistical Manual have generated as much controversy as Dissociative Identity Disorder (DID). For the past 35 years diagnoses of DID, previously referred to as Multiple Personality Disorder (MPD), have increased exponentially, causing various psychological researchers and clinicians to question the…

  19. Functional (dissociative) retrograde amnesia.

    PubMed

    Markowitsch, H J; Staniloiu, A

    2017-01-01

    Retrograde amnesia is described as condition which can occur after direct brain damage, but which occurs more frequently as a result of a psychiatric illness. In order to understand the amnesic condition, content-based divisions of memory are defined. The measurement of retrograde memory is discussed and the dichotomy between "organic" and "psychogenic" retrograde amnesia is questioned. Briefly, brain damage-related etiologies of retrograde amnesia are mentioned. The major portion of the review is devoted to dissociative amnesia (also named psychogenic or functional amnesia) and to the discussion of an overlap between psychogenic and "brain organic" forms of amnesia. The "inability of access hypothesis" is proposed to account for most of both the organic and psychogenic (dissociative) patients with primarily retrograde amnesia. Questions such as why recovery from retrograde amnesia can occur in retrograde (dissociative) amnesia, and why long-term new learning of episodic-autobiographic episodes is possible, are addressed. It is concluded that research on retrograde amnesia research is still in its infancy, as the neural correlates of memory storage are still unknown. It is argued that the recollection of episodic-autobiographic episodes most likely involves frontotemporal regions of the right hemisphere, a region which appears to be hypometabolic in patients with dissociative amnesia.

  20. Dissociative Reactions to Incest.

    ERIC Educational Resources Information Center

    Hall, J. Mark

    In contrast to Freud's later and revised view of the etiology of hysterical, or dissociative, symptoms, it is now known that real, and not fantasized, sexual experiences in childhood are experienced in disociative symptomatology. It is useful to understand that incest involves both traumatic events, that is, incidents of sexual violation per se,…

  1. Pathological Dissociation as Measured by the Child Dissociative Checklist

    ERIC Educational Resources Information Center

    Wherry, Jeffrey N.; Neil, Debra A.; Taylor, Tamara N.

    2009-01-01

    The component structure of the Child Dissociative Checklist was examined among abused children. A factor described as pathological dissociation emerged that was predicted by participants being male. There also were differences in pathological dissociation between groups of sexually abused and physically abused children. Replication of this factor…

  2. Dissociation and psychosis in dissociative identity disorder and schizophrenia.

    PubMed

    Laddis, Andreas; Dell, Paul F

    2012-01-01

    Dissociative symptoms, first-rank symptoms of schizophrenia, and delusions were assessed in 40 schizophrenia patients and 40 dissociative identity disorder (DID) patients with the Multidimensional Inventory of Dissociation (MID). Schizophrenia patients were diagnosed with the Structured Clinical Interview for the DSM-IV Axis I Disorders; DID patients were diagnosed with the Structured Clinical Interview for DSM-IV Dissociative Disorders-Revised. DID patients obtained significantly (a) higher dissociation scores; (b) higher passive-influence scores (first-rank symptoms); and (c) higher scores on scales that measure child voices, angry voices, persecutory voices, voices arguing, and voices commenting. Schizophrenia patients obtained significantly higher delusion scores than did DID patients. What is odd is that the dissociation scores of schizophrenia patients were unrelated to their reports of childhood maltreatment. Multiple regression analyses indicated that 81% of the variance in DID patients' dissociation scores was predicted by the MID's Ego-Alien Experiences Scale, whereas 92% of the variance in schizophrenia patients' dissociation scores was predicted by the MID's Voices Scale. We propose that schizophrenia patients' responses to the MID do not index the same pathology as do the responses of DID patients. We argue that neither phenomenological definitions of dissociation nor the current generation of dissociation instruments (which are uniformly phenomenological in nature) can distinguish between the dissociative phenomena of DID and what we suspect are just the dissociation-like phenomena of schizophrenia.

  3. Stability of flavin semiquinones in the gas phase: the electron affinity, proton affinity, and hydrogen atom affinity of lumiflavin.

    PubMed

    Zhang, Tianlan; Papson, Kaitlin; Ochran, Richard; Ridge, Douglas P

    2013-11-07

    Examination of electron transfer and proton transfer reactions of lumiflavin and proton transfer reactions of the lumiflavin radical anion by Fourier transform ion cyclotron resonance mass spectrometry is described. From the equilibrium constant determined for electron transfer between 1,4-naphthoquinone and lumiflavin the electron affinity of lumiflavin is deduced to be 1.86 ± 0.1 eV. Measurements of the rate constants and efficiencies for proton transfer reactions indicate that the proton affinity of the lumiflavin radical anion is between that of difluoroacetate (331.0 kcal/mol) and p-formyl-phenoxide (333.0 kcal/mol). Combining the electron affinity of lumiflavin with the proton affinity of the lumiflavin radical anion gives a lumiflavin hydrogen atom affinity of 59.7 ± 2.2 kcal/mol. The ΔG298 deduced from these results for adding an H atom to gas phase lumiflavin, 52.1 ± 2.2 kcal/mol, is in good agreement with ΔG298 for adding an H atom to aqueous lumiflavin from electrochemical measurements in the literature, 51.0 kcal/mol, and that from M06-L density functional calculations in the literature, 51.2 kcal/mol, suggesting little, if any, solvent effect on the H atom addition. The proton affinity of lumiflavin deduced from the equilibrium constant for the proton transfer reaction between lumiflavin and 2-picoline is 227.3 ± 2.0 kcal mol(-1). Density functional theory calculations on isomers of protonated lumiflavin provide a basis for assigning the most probable site of protonation as position 1 on the isoalloxazine ring and for estimating the ionization potentials of lumiflavin neutral radicals.

  4. Performance of dye-affinity beads for aluminium removal in magnetically stabilized fluidized bed

    PubMed Central

    Yavuz, Handan; Say, Ridvan; Andaç, Müge; Bayraktar, Necmi; Denizli, Adil

    2004-01-01

    Background Aluminum has recently been recognized as a causative agent in dialysis encephalopathy, osteodystrophy, and microcytic anemia occurring in patients with chronic renal failure who undergo long-term hemodialysis. Only a small amount of Al(III) in dialysis solutions may give rise to these disorders. Methods Magnetic poly(2-hydroxyethyl methacrylate) (mPHEMA) beads in the size range of 80–120 μm were produced by free radical co-polymerization of HEMA and ethylene dimethacrylate (EDMA) in the presence of magnetite particles (Fe3O4). Then, metal complexing ligand alizarin yellow was covalently attached onto mPHEMA beads. Alizarin yellow loading was 208 μmol/g. These beads were used for the removal of Al(III) ions from tap and dialysis water in a magnetically stabilized fluidized bed. Results Al(III) adsorption capacity of the beads decreased with an increase in the flow-rate. The maximum Al(III) adsorption was observed at pH 5.0. Comparison of batch and magnetically stabilized fluidized bed (MSFB) maximum capacities determined using Langmuir isotherms showed that dynamic capacity (17.5 mg/g) was somewhat higher than the batch capacity (11.8 mg/g). The dissociation constants for Al(III) were determined using the Langmuir isotherm equation to be 27.3 mM (MSFB) and 6.7 mM (batch system), indicating medium affinity, which was typical for pseudospecific affinity ligands. Al(III) ions could be repeatedly adsorbed and desorbed with these beads without noticeable loss in their Al(III) adsorption capacity. Conclusions Adsorption of Al(III) demonstrate the affinity of magnetic dye-affinity beads. The MSFB experiments allowed us to conclude that this inexpensive sorbent system may be an important alternative to the existing adsorbents in the removal of aluminium. PMID:15329149

  5. High affinity binding of [3H]-tyramine in the central nervous system.

    PubMed Central

    Vaccari, A.

    1986-01-01

    Optimum assay conditions for the association of [3H]-para-tyramine [( 3H]-pTA) with rat brain membranes were characterized, and a saturable, reversible, drug-specific, and high affinity binding mechanism for this trace amine was revealed. The binding capacity (Bmax) for [3H]-pTA in the corpus striatum was approximately 30 times higher than that in the cerebellum, with similar dissociation constants (KD). The binding process of [3H]-pTA involved the dopamine system, inasmuch as (a) highest binding capacity was associated with dopamine-rich regions; (b) dopamine and pTA equally displaced specifically bound [3H]-pTA; (c) there was a severe loss in striatal binding capacity for [3H]-pTA and, reportedly, for [3H]-dopamine, following unilateral nigrostriatal lesion; (d) acute in vivo reserpine treatment markedly decreased the density of [3H]-pTA and, reportedly, of [3H]-dopamine binding sites. In competition experiments [3H]-pTA binding sites, though displaying nanomolar affinity for dopamine, revealed micromolar affinities for the dopamine agonists apomorphine and pergolide, and for several dopamine antagonists, while having very high affinity for reserpine, a marker for the catecholamine transporter in synaptic vesicles. The binding process of [3H]-pTA was both energy-dependent (ouabain-sensitive), and ATP-Mg2+-insensitive; furthermore, the potencies of various drugs in competing for [3H]-pTA binding to rat striatal membranes correlated well (r = 0.96) with their reported potencies in inhibiting [3H]-dopamine uptake into striatal synaptosomes. It is concluded that [3H]-pTA binds at a site located on/within synaptic vesicles where it is involved in the transport mechanism of dopamine. PMID:3801770

  6. Surface plasmon resonance imaging for affinity analysis of aptamer-protein interactions with PDMS microfluidic chips.

    PubMed

    Wang, Zhuangzhi; Wilkop, Thomas; Xu, Danke; Dong, Yi; Ma, Guangyu; Cheng, Quan

    2007-10-01

    We report on the use of PDMS multichannels for affinity studies of DNA aptamer-human Immunoglobulin E (IgE) interactions by surface plasmon resonance imaging (SPRi). The sensing surface was prepared with thiol-terminated aptamers through a self-assembling process in the PDMS channels defined on a gold substrate. Cysteamine was codeposited with the thiol aptamers to promote proper spatial arrangement of the aptamers and thus maintain their optimal binding efficiencies. Four aptamers with different nucleic acid sequences were studied to test their interaction affinity toward IgE, and the results confirmed that aptamer I (5'-SH-GGG GCA CGT TTA TCC GTC CCT CCT AGT GGC GTG CCC C-3') has the strongest binding affinity. Control experiments were conducted with a PEG-functionalized surface and IgG was used to replace IgE in order to verify the selective binding of aptamer I to the IgE molecules. A linear concentration-dependent relationship between IgE and aptamer I was obtained, and a 2-nM detection limit was achieved. SPRi data were further analyzed by global fitting, and the dissociation constant of aptamer I-IgE complex was found to be 2.7 x 10(-7) M, which agrees relatively well with the values reported in the literature. Aptamer affinity screening by SPR imaging demonstrates marked advantages over competing methods because it does not require labeling, can be used in real-time, and is potentially high-throughput. The ability to provide both qualitative and quantitative results on a multichannel chip further establishes SPRi as a powerful tool for the study of biological interactions in a multiplexed format.

  7. The serotonin transporter: Examination of the changes in transporter affinity induced by ligand binding

    SciTech Connect

    Humphreys, C.J.

    1989-01-01

    The plasmalemmal serotonin transporter uses transmembrane gradients of Na{sup +}, Cl{sup {minus}} and K{sup +} to accumulate serotonin within blood platelets. Transport is competitively inhibited by the antidepressant imipramine. Like serotonin transport, imipramine binding requires Na{sup +}. Unlike serotonin, however, imipramine does not appear to be transported. To gain insight into the mechanism of serotonin transport the author have analyzed the influences of Na{sup +} and Cl{sup {minus}}, the two ions cotransported with serotonin, on both serotonin transport and the interaction of imipramine and other antidepressant drugs with the plasmalemmal serotonin transporter of human platelets. Additionally, the author have synthesized, purified and characterized the binding of 2-iodoimipramine to the serotonin transporter. Finally, the author have conducted a preliminary study of the inhibition of serotonin transport and imipramine binding produced by dicyclohexylcarbodiimide. My results reveal many instances of positive heterotropic cooperativity in ligand binding to the serotonin transporter. Na{sup +} binding enhances the transporters affinity for imipramine and several other antidepressant drugs, and also increases the affinity for Cl{sup {minus}}. Cl{sup {minus}} enhances the transporters affinity for imipramine, as well as for Na{sup +}. At concentrations in the range of its K{sub M} for transport serotonin is a competitive inhibitor of imipramine binding. At much higher concentrations, however, serotonin also inhibits imipramines dissociation rate constant. This latter effect which is Na{sup +}-independent and species specific, is apparently produced by serotonin binding at a second, low affinity site on, or near, the transporter complex. Iodoimipramine competitively inhibit both ({sup 3}H)imipramine binding and ({sup 3}H)serotonin transport.

  8. A dielectric affinity microbiosensor

    NASA Astrophysics Data System (ADS)

    Huang, Xian; Li, Siqi; Schultz, Jerome S.; Wang, Qian; Lin, Qiao

    2010-01-01

    We present an affinity biosensing approach that exploits changes in dielectric properties of a polymer due to its specific, reversible binding with an analyte. The approach is demonstrated using a microsensor comprising a pair of thin-film capacitive electrodes sandwiching a solution of poly(acrylamide-ran-3-acrylamidophenylboronic acid), a synthetic polymer with specific affinity to glucose. Binding with glucose induces changes in the permittivity of the polymer, which can be measured capacitively for specific glucose detection, as confirmed by experimental results at physiologically relevant concentrations. The dielectric affinity biosensing approach holds the potential for practical applications such as long-term continuous glucose monitoring.

  9. Affinity in electrophoresis.

    PubMed

    Heegaard, Niels H H

    2009-06-01

    The journal Electrophoresis has greatly influenced my approaches to biomolecular affinity studies. The methods that I have chosen as my main tools to study interacting biomolecules--native gel and later capillary zone electrophoresis--have been the topic of numerous articles in Electrophoresis. Below, the role of the journal in the development and dissemination of these techniques and applications reviewed. Many exhaustive reviews on affinity electrophoresis and affinity CE have been published in the last few years and are not in any way replaced by the present deliberations that are focused on papers published by the journal.

  10. Evaluation of kinetic constants of biomolecular interaction on optical surface plasmon resonance sensor with Newton Iteration Method

    NASA Astrophysics Data System (ADS)

    Zhao, Yuanyuan; Jiang, Guoliang; Hu, Jiandong; Hu, Fengjiang; Wei, Jianguang; Shi, Liang

    2010-10-01

    In the immunology, there are two important types of biomolecular interaction: antigens-antibodies and receptors-ligands. Monitoring the response rate and affinity of biomolecular interaction can help analyze the protein function, drug discover, genomics and proteomics research. Moreover the association rate constant and dissociation rate constant of receptors-ligands are the important parameters for the study of signal transmission between cells. Recent advances in bioanalyzer instruments have greatly simplified the measurement of the kinetics of molecular interactions. Non-destructive and real-time monitoring the response to evaluate the parameters between antigens and antibodies can be performed by using optical surface plasmon resonance (SPR) biosensor technology. This technology provides a quantitative analysis that is carried out rapidly with label-free high-throughput detection using the binding curves of antigens-antibodies. Consequently, the kinetic parameters of interaction between antigens and antibodies can be obtained. This article presents a low cost integrated SPR-based bioanalyzer (HPSPR-6000) designed by ourselves. This bioanalyzer is mainly composed of a biosensor TSPR1K23, a touch-screen monitor, a microprocessor PIC24F128, a microflow cell with three channels, a clamp and a photoelectric conversion device. To obtain the kinetic parameters, sensorgrams may be modeled using one of several binding models provided with BIAevaluation software 3.0, SensiQ or Autolab. This allows calculation of the association rate constant (ka) and the dissociation rate constant (kd). The ratio of ka to kd can be used to estimate the equilibrium constant. Another kind is the analysis software OriginPro, which can process the obtained data by nonlinear fitting and then get some correlative parameters, but it can't be embedded into the bioanalyzer, so the bioanalyzer don't support the use of OriginPro. This paper proposes a novel method to evaluate the kinetic parameters

  11. Three dimensions of dissociative amnesia.

    PubMed

    Dell, Paul F

    2013-01-01

    Principal axis factor analysis with promax rotation extracted 3 factors from the 42 memory and amnesia items of the Multidimensional Inventory of Dissociation (MID) database (N = 2,569): Discovering Dissociated Actions, Lapses of Recent Memory and Skills, and Gaps in Remote Memory. The 3 factors' shared variance ranged from 36% to 64%. Construed as scales, the 3 factor scales had Cronbach's alpha coefficients of .96, .94, and .93, respectively. The scales correlated strongly with mean Dissociative Experiences Scale scores, mean MID scores, and total scores on the Structured Clinical Interview for DSM-IV Dissociative Disorders-Revised (SCID-D-R). What is interesting is that the 3 amnesia factors exhibited a range of correlations with SCID-D-R Amnesia scores (.52, .63, and .70, respectively), suggesting that the SCID-D-R Amnesia score emphasizes gaps in remote memory over amnesias related to dissociative identity disorder. The 3 amnesia factor scales exhibited a clinically meaningful pattern of significant differences among dissociative identity disorder, dissociative disorder not otherwise specified-1, dissociative amnesia, depersonalization disorder, and nonclinical participants. The 3 amnesia factors may have greater clinical utility for frontline clinicians than (a) amnesia as discussed in the context of the Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition, nosology of the dissociative disorders or (b) P. Janet's (1893/1977 ) 4-fold classification of dissociative amnesia. The author recommends systematic study of the phenomenological differences within specific dissociative symptoms and their differential relationship to specific dissociative disorders.

  12. Affine dynamics with torsion

    NASA Astrophysics Data System (ADS)

    Gültekin, Kemal

    2016-03-01

    In this study, we give a thorough analysis of a general affine gravity with torsion. After a brief exposition of the affine gravities considered by Eddington and Schrödinger, we construct and analyze different affine gravities based on the determinants of the Ricci tensor, the torsion tensor, the Riemann tensor, and their combinations. In each case we reduce equations of motion to their simplest forms and give a detailed analysis of their solutions. Our analyses lead to the construction of the affine connection in terms of the curvature and torsion tensors. Our solutions of the dynamical equations show that the curvature tensors at different points are correlated via non-local, exponential rescaling factors determined by the torsion tensor.

  13. Lectin affinity electrophoresis.

    PubMed

    Kobayashi, Yuka

    2014-01-01

    An interaction or a binding event typically changes the electrophoretic properties of a molecule. Affinity electrophoresis methods detect changes in the electrophoretic pattern of molecules (mainly macromolecules) that occur as a result of biospecific interactions or complex formation. Lectin affinity electrophoresis is a very effective method for the detection and analysis of trace amounts of glycobiological substances. It is particularly useful for isolating and separating the glycoisomers of target molecules. Here, we describe a sensitive technique for the detection of glycoproteins separated by agarose gel-lectin affinity electrophoresis that uses antibody-affinity blotting. The technique is tested using α-fetoprotein with lectin (Lens culinaris agglutinin and Phaseolus vulgaris agglutinin)-agarose gels.

  14. In vitro analysis of human immunodeficiency virus particle dissociation: gag proteolytic processing influences dissociation kinetics.

    PubMed

    Müller, Barbara; Anders, Maria; Reinstein, Jochen

    2014-01-01

    Human immunodeficiency virus particles undergo a step of proteolytic maturation, in which the main structural polyprotein Gag is cleaved into its mature subunits matrix (MA), capsid (CA), nucleocapsid (NC) and p6. Gag proteolytic processing is accompanied by a dramatic structural rearrangement within the virion, which is necessary for virus infectivity and has been proposed to proceed through a sequence of dissociation and reformation of the capsid lattice. Morphological maturation appears to be tightly regulated, with sequential cleavage events and two small spacer peptides within Gag playing important roles by regulating the disassembly of the immature capsid layer and formation of the mature capsid lattice. In order to measure the influence of individual Gag domains on lattice stability, we established Förster's resonance energy transfer (FRET) reporter virions and employed rapid kinetic FRET and light scatter measurements. This approach allowed us to measure dissociation properties of HIV-1 particles assembled in eukaryotic cells containing Gag proteins in different states of proteolytic processing. While the complex dissociation behavior of the particles prevented an assignment of kinetic rate constants to individual dissociation steps, our analyses revealed characteristic differences in the dissociation properties of the MA layer dependent on the presence of additional domains. The most striking effect observed here was a pronounced stabilization of the MA-CA layer mediated by the presence of the 14 amino acid long spacer peptide SP1 at the CA C-terminus, underlining the crucial role of this peptide for the resolution of the immature particle architecture.

  15. Dissociative recombination and the decay of a molecular ultracold plasma

    NASA Astrophysics Data System (ADS)

    Rennick, C. J.; Saquet, N.; Morrison, J. P.; Ortega-Arroyo, J.; Godin, P.; Fu, L.; Schulz-Weiling, M.; Grant, E. R.

    2011-07-01

    Double-resonant photoexcitation of nitric oxide in a molecular beam creates a dense ensemble of 51f(2) Rydberg states, which evolves to form a plasma of free electrons trapped in the potential well of an NO+ spacecharge. The plasma travels at the velocity of the molecular beam, and, on passing through a grounded grid, yields an electron time-of-flight signal that gauges the plasma size and quantity of trapped electrons. This plasma expands at a rate that fits with an electron temperature as low as 5 K. Dissociative recombination of NO+ ions with electrons provides the primary dissipation mechanism for the plasma. We have identified three dissociation pathways, and quantified their relative contributions to the measured rate: Two-body dissociative recombination competes with direct three-body recombination to neutral dissociation products, and with a process in which three-body recombination and electron-impact ionization form an equilibrium population of high-Rydberg states that decays by predissociation. Using available collision-theory rate constants for three-body recombination and ionization, together with quantum mechanical estimates of predissociation rates, we predict that the relaxation of the plasma to a high-Rydberg equilibrium outpaces direct three-body dissociative recombination, and, among second-order processes, the rate of two-body electron-cation dissociative recombination substantially exceeds the rate at which the high-Rydberg equilibrium dissociatively relaxes. The rate constant for dissociative recombination extracted from these data conforms with predictions drawn from theory for isolated electron-ion collisions. Methods based on the dissipation of molecular ultracold plasmas may provide a means for estimating rates of dissociative recombination for a variety of complex molecules.

  16. The second acidic constant of salicylic acid.

    PubMed

    Porto, Raffaella; De Tommaso, Gaetano; Furia, Emilia

    2005-01-01

    The second dissociation constant of salicylic acid (H2L) has been determined, at 25 degrees C, in NaCl ionic media by UV spectrophotometric measurements. The investigated ionic strength values were 0.16, 0.25, 0.50, 1.0, 2.0 and 3.0 M. The protolysis constants calculated at the different ionic strengths yielded, with the Specific Interaction Theory, the infinite dilution constant, log beta1(0) = 13.62 +/- 0.03, for the equilibrium L2- + H+ <==> HL-. The interaction coefficient between Na+ and L2-, b(Na+, L2-) = 0.02 +/- 0.07, has been also calculated.

  17. Crystal structure and ligand affinity of avidin in the complex with 4‧-hydroxyazobenzene-2-carboxylic acid

    NASA Astrophysics Data System (ADS)

    Strzelczyk, Paweł; Bujacz, Grzegorz

    2016-04-01

    Avidin is a protein found in egg white that binds numerous organic compounds with high affinity, especially biotin and its derivatives. Due to its extraordinary affinity for its ligands, avidin is extensively used in biotechnology. X-ray crystallography and fluorescence-based biophysical techniques were used to show that avidin binds the dye 4‧-hydroxyazobenzene-2-carboxylic acid (HABA) with a lower affinity than biotin. The apparent dissociation constant determined for the avidin complex with HABA by microscale thermophoresis (MST) is 4.12 μM. The crystal structure of avidin-HABA complex was determined at a resolution of 2.2 Å (PDB entry 5chk). The crystals belong to a hexagonal system, in the space group P6422. In that structure, the hydrazone tautomer of HABA is bound at the bottom part of the central calyx near the polar residues. We show interactions of the dye with avidin and compare them with the previously reported avidin-biotin complex.

  18. High-affinity interactions between peptides and heat shock protein 70 augment CD8+ T lymphocyte immune responses.

    PubMed

    Flechtner, Jessica B; Cohane, Kenya Prince; Mehta, Sunil; Slusarewicz, Paul; Leonard, Alexis Kays; Barber, Brian H; Levey, Daniel L; Andjelic, Sofija

    2006-07-15

    Exogenously delivered antigenic peptides complexed to heat shock proteins (HSPs) are able to enter the endogenous Ag-processing pathway and prime CD8+ CTL. It was determined previously that a hybrid peptide containing a MHC class I-binding epitope and HSP70-binding sequence Javelin (J0) in complex with HSP70 could induce cytotoxic T cell responses in vivo that were more robust than those induced by the minimal epitope complexed with HSP70. The present study introduces a novel, higher-affinity HSP70-binding sequence (J1) that significantly enhances binding of various antigenic peptides to HSP70. A competition binding assay revealed a dissociation constant that was 15-fold lower for the H2-K(b) OVA epitope SIINFEKL-J1 compared with SIINFEKL-J0, indicating a substantially higher affinity for HSP70. Further, modifying the orientation of the hybrid epitope and introducing a cleavable linker sequence between the Javelin and the epitope results in even greater immunogenicity, presumably by greater efficiency of epitope processing. The enhanced immunogenicity associated with Javelin J1 and the cleavable linker is consistently observed with multiple mouse and human epitopes. Thus, by creating a series of epitopes with uniform, high-affinity binding to HSP70, successful multiple epitope immunizations are possible, with equal delivery of each antigenic epitope to the immune system via HSP70. These modified epitopes have the potential for creating successful multivalent vaccines for immunotherapy of both infectious disease and cancer.

  19. Intensity dependence of multiphoton dissociation in formaldehyde

    NASA Astrophysics Data System (ADS)

    Koren, G.

    1980-01-01

    The paper reports a new intensity-dependent measurement of multiple-photon dissociation (MPD) in H2CO, HDCO, and D2CO gases using an intense pulsed CO2 TEA laser. In this measurement the energy and duration of the laser pulses are constant, and the intensity is varied by irradiating the sample with concave mirrors of different focal lengths. A model calculation is used to analyze and fit the MPD data of HDCO and D2CO which assumes that dissociation is obtained by a repeated mechanism in which coherent multiphoton excitation (CME) of the molecule to high vibration-rotation states is followed by intramolecular transfer of the excitation energy (ITEE) to the other molecule modes. It is concluded that the results are consistent with the absorption of 14 plus or minus 4 and 17 plus or minus 5 photons per molecule of HDCO and D2CO, respectively.

  20. Dissociation: cognitive capacity or dysfunction?

    PubMed

    de Ruiter, Michiel B; Elzinga, Bernet M; Phaf, R Hans

    2006-01-01

    Dissociative experiences are mostly studied as a risk factor for dissociative pathology. Nonpathological dissociation is quite common in the general population, however, and may reflect a constitutionally determined cognitive style rather than a pathological trait acquired through the experience of adverse life events. In a theoretical model, we propose that nonpathological dissociation is characterized by high levels of elaboration learning and reconstructive retrieval, for which enhanced levels of attentional and working memory abilities are a prerequisite. These characteristics, in general, seem to be representative for a higher ability to (re-)construct conscious experiences. We review some of our behavioral as well as neural (i.e., fMRI, ERPs) studies, suggesting that high dissociative individuals are characterized by heightened levels of attention, working memory and episodic memory. In nonpathological conditions a person may benefit from these dissociative abilities, although after adverse (e.g., traumatic) events the disposition may develop into dissociative pathology.

  1. Affinity ligands for glycoprotein purification based on the multi-component Ugi reaction.

    PubMed

    Chen, Chen; Khoury, Graziella El; Lowe, Christopher R

    2014-10-15

    One challenge facing the purification of therapeutic glycoproteins by affinity chromatography is creating ligands specific for the glycan moiety. Affinity chromatography of glycoproteins is currently conducted with immobilized lectins or boronates, although biomimetic ligands could present a more desirable option. This work describes the rational design and combinatorial synthesis of carbohydrate-binding ligands based on the solid phase multi-component Ugi reaction. An aldehyde-functionalized Sepharose™ solid support constitutes one component (aldehyde) in the four-component reaction, while the other three components (a primary/secondary amine, a carboxylic acid and an isocyanide) are varied in a combinatorial fashion to generate a tri-substituted Ugi scaffold which provides a degree of rigidity and is functionally suitable for interacting with the glycan moiety of glycoproteins. An Ugi library containing 48 ligands was initially screened against glucose oxidase (GOx) as the model glycoprotein to identify a candidate ligand, A13C24I8, which showed affinity to GOx through its carbohydrate moiety. Immobilized ligand A13C24I8 demonstrated a static binding capacity of 16.7mg GOx/ml resin and an apparent dissociation constant (Kd) of 1.45×10(-6)M at pH 7.4. The adsorbent can also bind 8.1mg AGP/ml resin and displays an apparent affinity constant Kd=1.44×10(-5)M. The ligand has a sugar specificity in the following sequence: sorbitol>fructose>mannitol>ribose>arabinose>xylose>galactose>mannose>glucose>fructose; however, it did not display any specificity for sialic acid or methyl α-D-glycosides. A control ligand, generated by substitution of C24 (3-carboxyphenylboronic acid) with C7 (4-hydroxyphenyl acetic acid), failed to show affinity to the carbohydrate moiety, supporting the importance of the role that boronic acid group plays in sugar binding. GOx spiked E. coli samples were loaded onto immobilized ligand A13C24I8, 3-aminophenylboronic acid (APBA) and

  2. Hsp70 chaperones are non-equilibrium machines that achieve ultra-affinity by energy consumption.

    PubMed

    De Los Rios, Paolo; Barducci, Alessandro

    2014-05-27

    70-kDa Heat shock proteins are ATP-driven molecular chaperones that perform a myriad of essential cellular tasks. Although structural and biochemical studies have shed some light on their functional mechanism, the fundamental issue of the role of energy consumption, due to ATP-hydrolysis, has remained unaddressed. Here we establish a clear connection between the non-equilibrium nature of Hsp70, due to ATP hydrolysis, and the determining feature of its function, namely its high affinity for its substrates. Energy consumption can indeed decrease the dissociation constant of the chaperone-substrate complex by several orders of magnitude with respect to an equilibrium scenario. We find that the biochemical requirements for observing such ultra-affinity coincide with the physiological conditions in the cell. Our results rationalize several experimental observations and pave the way for further analysis of non-equilibrium effects underlying chaperone functions.DOI: http://dx.doi.org/10.7554/eLife.02218.001.

  3. Isomer-Specific Binding Affinity of Perfluorooctanesulfonate (PFOS) and Perfluorooctanoate (PFOA) to Serum Proteins.

    PubMed

    Beesoon, Sanjay; Martin, Jonathan W

    2015-05-05

    Perfluorooctanesulfonate (PFOS) and perfluorooctanoate (PFOA) are among the most prominent contaminants in human serum, and these were historically manufactured as technical mixtures of linear and branched isomers. The isomers display unique pharmacokinetics in humans and in animal models, but molecular mechanisms underlying isomer-specific PFOS and PFOA disposition have not previously been studied. Here, ultrafiltration devices were used to examine (i) the dissociation constants (Kd) of individual PFOS and PFOA isomers with human serum albumin (HSA) and (ii) relative binding affinity of isomers in technical mixtures spiked to whole calf serum and human serum. Measurement of HSA Kd's demonstrated that linear PFOS (Kd=8(±4)×10(-8) M) was much more tightly bound than branched PFOS isomers (Kd range from 8(±1)×10(-5) M to 4(±2)×10(-4) M). Similarly, linear PFOA (Kd=1(±0.9)×10(-4) M) was more strongly bound to HSA compared to branched PFOA isomers (Kd range from 4(±2)×10(-4) M to 3(±2)×10(-4) M). The higher binding affinities of linear PFOS and PFOA to total serum protein were confirmed when both calf serum and human serum were spiked with technical mixtures. Overall, these data provide a mechanistic explanation for the longer biological half-life of PFOS in humans, compared to PFOA, and for the higher transplacental transfer efficiencies and renal clearance of branched PFOS and PFOA isomers, compared to the respective linear isomer.

  4. Purification and affinity labeling of dihydropyridine receptor from rabbit skeletal muscle membranes

    SciTech Connect

    Kanngiesser, U.; Nalik, P.; Pongs, O.

    1988-05-01

    Undegraded dihydropyridine (DHP)-receptor (putatively a voltage-gated Ca/sup 2 +/ channel) has been purified as a 340-kDa protein complex to approx.80% homogeneity (2.4 nmol of DHP-receptor per mg of protein) from rabbit skeletal muscle by a rapid purification protocol. Transverse-tubule membranes were prepared in high yield by Ribi-press treatment. The DHP-receptor complex was solubilized in 1% digitonin followed by a two step-chromatographic purification procedure. The equilibrium dissociation constant of (/sup 3/H) (+) -PN200-110 binding (K/sub d/; 0.9 nM) was not significantly changed by solubilization or purification. The purified DHP-receptor is composed of two subunits with apparent molecular masses of 148 kDa and 195 kDa migrating in polyacrylamide gels under nonreducing conditions as a single moiety of approx.300 kDa. The 195-kDa subunit was affinity-labeled with (/sup 3/H)azidopine in both transverse-tubule membranes and purified DHP-receptor preparations. The subunit can be degraded by high-energy irradiation to a 26-kDa peptide and by proteolysis to a 32-kDa peptide. Thus, it is probably due to proteolytic cleavage and/or photolysis that neither purification nor affinity-labeling studies have previously identified a DHP-receptor subunit of comparable molecular mass (195 kDa).

  5. Affine group formulation of the Standard Model coupled to gravity

    SciTech Connect

    Chou, Ching-Yi; Ita, Eyo; Soo, Chopin

    2014-04-15

    In this work we apply the affine group formalism for four dimensional gravity of Lorentzian signature, which is based on Klauder’s affine algebraic program, to the formulation of the Hamiltonian constraint of the interaction of matter and all forces, including gravity with non-vanishing cosmological constant Λ, as an affine Lie algebra. We use the hermitian action of fermions coupled to gravitation and Yang–Mills theory to find the density weight one fermionic super-Hamiltonian constraint. This term, combined with the Yang–Mills and Higgs energy densities, are composed with York’s integrated time functional. The result, when combined with the imaginary part of the Chern–Simons functional Q, forms the affine commutation relation with the volume element V(x). Affine algebraic quantization of gravitation and matter on equal footing implies a fundamental uncertainty relation which is predicated upon a non-vanishing cosmological constant. -- Highlights: •Wheeler–DeWitt equation (WDW) quantized as affine algebra, realizing Klauder’s program. •WDW formulated for interaction of matter and all forces, including gravity, as affine algebra. •WDW features Hermitian generators in spite of fermionic content: Standard Model addressed. •Constructed a family of physical states for the full, coupled theory via affine coherent states. •Fundamental uncertainty relation, predicated on non-vanishing cosmological constant.

  6. Increased blood oxygen affinity during digestion in the snake Python molurus.

    PubMed

    Overgaard, Johannes; Wang, Tobias

    2002-11-01

    Many snakes exhibit large increases in metabolic rate during digestion that place extensive demands on efficient oxygen transport. In the present study, we describe blood oxygen affinity following three weeks of fasting and 48 h after feeding in the Burmese python (Python molurus). We also report simultaneous measurements of arterial blood gases and haematological parameters. Arterial blood was obtained from chronically implanted catheters, and blood oxygen-dissociation curves were constructed from oxygen content measurements at known oxygen partial pressure (P(O(2))) values at 2% and 5% CO(2). Arterial pH remained constant at approximately 7.6 after feeding, but digestion was associated with an approximately 6 mmol l(-1) increase in [HCO(3)(-)], while CO(2) partial pressure (P(CO(2))) increased from 2.21+/-0.13 kPa in fasted animals to 2.89+/-0.17 kPa at 48 h after feeding. Blood oxygen affinity in vivo was predicted on the basis of pH in vivo and the blood oxygen-dissociation curves obtained in vitro. The blood oxygen affinity in vivo increased during digestion, with P(50) values decreasing from 4.58+/-0.11 kPa to 3.53+/-0.24 kPa. This increase was associated with a significant decrease in the red blood cell [NTP]/[Hb(4)] ratio (relationship between the concentrations of organic phosphates and total haemoglobin) and a significant decrease in mean cellular haemoglobin content, which is indicative of swelling of the red blood cells. Our data for blood oxygen affinity and arterial oxygen levels, together with previously published values of oxygen uptake and blood flows, allow for a quantitative evaluation of oxygen transport during digestion. This analysis shows that a large part of the increased metabolism during digestion is supported by an increased venous extraction, while arterial P(O(2)) (Pa(O(2))) and haemoglobin saturation do not vary with digestive status. Thus, we predict that venous P(O(2)) (Pv(O(2))) is reduced from a fasting value of 5.2 kPa to 1.6 k

  7. Extraction of haemoglobin from human blood by affinity precipitation using a haptoglobin-based stimuli-responsive affinity macroligand.

    PubMed

    Stocker-Majd, Gisela; Hilbrig, Frank; Freitag, Ruth

    2008-06-13

    Affinity precipitation was compared to affinity chromatography and batch adsorption as the final purification step in a protocol for the isolation of haemoglobin from human blood. Haptoglobin was the affinity ligand. The first steps on the process were realized by traditional methods (lyses of red blood cells followed by ammonium sulphate precipitation). For affinity chromatography (and batch adsorption) the ligand was linked to Sepharose, for affinity precipitation to a thermoresponsive polymer, namely poly(N-isopropylacrylamide). Five haptoglobin-poly(N-isopropylacrylamide) bioconjugates (affinity macroligands) were constructed with different polymer: haptoglobin-coupling ratios. Conjugation of haptoglobin to the soluble poly(N-isopropylacrylamide) apparently does not change the interaction thermodynamics with haemoglobin, as the haemoglobin binding constants calculated by a Scatchard analysis for the affinity macroligand were of the same order of magnitude as those described in the literature for the haemoglobin-haptoglobin complex in solution. Two elution protocols were used for haemoglobin release from the various affinity materials, one at pH 2, the other with 5 M urea at pH 11. Both affinity chromatography and affinity precipitation yielded a pure haemoglobin of high quality. Compared to the affinity chromatography, affinity precipitation showed a significantly higher ligand efficiency (ratio of the experimental capacity to the theoretical one). The method thus makes better use of the expensive affinity ligands. As affinity precipitation only requires small temperature changes to bring about precipitation/redissolution of the affinity complexes and a centrifugation step for recovery of the precipitate, the method in addition has advantages in term of scalability and simplicity.

  8. Determinants of the Differential Antizyme-Binding Affinity of Ornithine Decarboxylase

    PubMed Central

    Liu, Yen-Chin; Hsu, Den-Hua; Huang, Chi-Liang; Liu, Yi-Liang; Liu, Guang-Yaw; Hung, Hui-Chih

    2011-01-01

    Ornithine decarboxylase (ODC) is a ubiquitous enzyme that is conserved in all species from bacteria to humans. Mammalian ODC is degraded by the proteasome in a ubiquitin-independent manner by direct binding to the antizyme (AZ). In contrast, Trypanosoma brucei ODC has a low binding affinity toward AZ. In this study, we identified key amino acid residues that govern the differential AZ binding affinity of human and Trypanosoma brucei ODC. Multiple sequence alignments of the ODC putative AZ-binding site highlights several key amino acid residues that are different between the human and Trypanosoma brucei ODC protein sequences, including residue 119, 124,125, 129, 136, 137 and 140 (the numbers is for human ODC). We generated a septuple human ODC mutant protein where these seven bases were mutated to match the Trypanosoma brucei ODC protein sequence. The septuple mutant protein was much less sensitive to AZ inhibition compared to the WT protein, suggesting that these amino acid residues play a role in human ODC-AZ binding. Additional experiments with sextuple mutants suggest that residue 137 plays a direct role in AZ binding, and residues 119 and 140 play secondary roles in AZ binding. The dissociation constants were also calculated to quantify the affinity of the ODC-AZ binding interaction. The Kd value for the wild type ODC protein-AZ heterodimer ([ODC_WT]-AZ) is approximately 0.22 μM, while the Kd value for the septuple mutant-AZ heterodimer ([ODC_7M]-AZ) is approximately 12.4 μM. The greater than 50-fold increase in [ODC_7M]-AZ binding affinity shows that the ODC-7M enzyme has a much lower binding affinity toward AZ. For the mutant proteins ODC_7M(-Q119H) and ODC_7M(-V137D), the Kd was 1.4 and 1.2 μM, respectively. These affinities are 6-fold higher than the WT_ODC Kd, which suggests that residues 119 and 137 play a role in AZ binding. PMID:22073206

  9. Dissociative absorption: An empirically unique, clinically relevant, dissociative factor.

    PubMed

    Soffer-Dudek, Nirit; Lassri, Dana; Soffer-Dudek, Nir; Shahar, Golan

    2015-11-01

    Research of dissociative absorption has raised two questions: (a) Is absorption a unique dissociative factor within a three-factor structure, or a part of one general dissociative factor? Even when three factors are found, the specificity of the absorption factor is questionable. (b) Is absorption implicated in psychopathology? Although commonly viewed as "non-clinical" dissociation, absorption was recently hypothesized to be specifically associated with obsessive-compulsive symptoms. To address these questions, we conducted exploratory and confirmatory factor analyses on 679 undergraduates. Analyses supported the three-factor model, and a "purified" absorption scale was extracted from the original inclusive absorption factor. The purified scale predicted several psychopathology scales. As hypothesized, absorption was a stronger predictor of obsessive-compulsive symptoms than of general psychopathology. In addition, absorption was the only dissociative scale that longitudinally predicted obsessive-compulsive symptoms. We conclude that absorption is a unique and clinically relevant dissociative tendency that is particularly meaningful to obsessive-compulsive symptoms.

  10. Dissociation of motor maturation.

    PubMed

    DiMario, Francis J

    2003-06-01

    We prospectively acquired clinical data regarding the presentation, evaluation, and developmental progress of all patients identified with dissociated motor maturation to define their clinical outcomes. Children (N = 8) referred for evaluation of suspected cerebral palsy because of delayed sitting or walking and identified to have dissociated motor maturation were followed with serial clinical examination. All displayed the characteristic "sitting on air" posture while held in vertical suspension and had otherwise normal developmental assessments. This posture is composed of the hips held in flexion and abduction with the knees extended and feet plantar or dorsiflexed. Three children were initially evaluated at 10 months of age owing to absence of sitting and five other children were evaluated at a mean of 14 months (range 12-19 months) owing to inability to stand. Follow-up evaluations were conducted over a mean of 10.5 months (range 5-34 months). Five children were born prematurely at 34 to 36 weeks gestation. Denver Developmental Screening Test and general and neurologic examinations were normal except to note hypotonia in six children and the "sitting on air" posture in all of the children. Four children have older siblings or parents who "walked late" (after 15 months). On average, the children attained sitting by 8 months (range 7-10 months). One child did not crawl prior to independent walking, two children scooted rather than crawled, and five children crawled at an average of 13.5 months (range 10-16 months). All children cruised by a mean of 18 months (range 16-21.5 months) and attained independent walking by 20.1 months (range 18-25 months). Neuroimaging and serum creatine kinase enzyme testing were normal in two children who were tested. These eight children conform to the syndrome of dissociated motor maturation. The "sitting on air" posture serves as a diagnostic sign and anticipated excellent prognosis, but follow-up is required to ensure a normal

  11. Affine Sphere Relativity

    NASA Astrophysics Data System (ADS)

    Minguzzi, E.

    2017-03-01

    We investigate spacetimes whose light cones could be anisotropic. We prove the equivalence of the structures: (a) Lorentz-Finsler manifold for which the mean Cartan torsion vanishes, (b) Lorentz-Finsler manifold for which the indicatrix (observer space) at each point is a convex hyperbolic affine sphere centered on the zero section, and (c) pair given by a spacetime volume and a sharp convex cone distribution. The equivalence suggests to describe (affine sphere) spacetimes with this structure, so that no algebraic-metrical concept enters the definition. As a result, this work shows how the metric features of spacetime emerge from elementary concepts such as measure and order. Non-relativistic spacetimes are obtained replacing proper spheres with improper spheres, so the distinction does not call for group theoretical elements. In physical terms, in affine sphere spacetimes the light cone distribution and the spacetime measure determine the motion of massive and massless particles (hence the dispersion relation). Furthermore, it is shown that, more generally, for Lorentz-Finsler theories non-differentiable at the cone, the lightlike geodesics and the transport of the particle momentum over them are well defined, though the curve parametrization could be undefined. Causality theory is also well behaved. Several results for affine sphere spacetimes are presented. Some results in Finsler geometry, for instance in the characterization of Randers spaces, are also included.

  12. Dissociative Electron Attachment

    NASA Astrophysics Data System (ADS)

    Arreola, Esmeralda; Esmeralda Arreola Collaboration; Leigh Hargreaves Collaboration

    Since the pioneering work of Boudiaffa et al., it has been understood that electrons, even with energies near or below the ionization threshold, are capable of initiating strand-breaks in human DNA. This discovery raised important questions for cancer treatments, since sub-ionizing electrons are known to be the most copiously produced secondary product of radiation therapy. But even to date these factors are largely excluded from dosimetry calculations. This lack of inclusion is, at least in part, certainly due to the dearth of fundamental data describing low-energy electron interactions with nucleotide molecules that form the basis of DNA. Understanding of how such slow electrons are able to damage DNA remains incomplete, but the strongly peaked nature of Boudiaffa et al.'s data gives strong hints at resonantly driven collision processes. DNA damage is therefore most likely driven by ``dissociative electron attachment'' (DEA). DEA is a rather complicated process to model due to the coupling of electronic and nuclear degrees of freedom in the molecule. At the California State University Fullerton, we are currently commissioning a new spectrometer to study dissociation channels, reaction rates and orientation effects in DEA collisions between slow electrons and nucleotide molecules. At the meeting we will present design parameters and commissioning data for this new apparatus.

  13. [Dissociative disorders and affective disorders].

    PubMed

    Montant, J; Adida, M; Belzeaux, R; Cermolacce, M; Pringuey, D; Da Fonseca, D; Azorin, J-M

    2014-12-01

    The phenomenology of dissociative disorders may be complex and sometimes confusing. We describe here two cases who were initially misdiagnosed. The first case concerned a 61 year-old woman, who was initially diagnosed as an isolated dissociative fugue and was actually suffering from severe major depressive episode. The second case concerned a 55 year-old man, who was suffering from type I bipolar disorder and polyvascular disease, and was initially diagnosed as dissociative fugue in a mooddestabilization context, while it was finally a stroke. Yet dissociative disorders as affective disorder comorbidity are relatively unknown. We made a review on this topic. Dissociative disorders are often studied through psycho-trauma issues. Litterature is rare on affective illness comorbid with dissociative disorders, but highlight the link between bipolar and dissociative disorders. The later comorbidity often refers to an early onset subtype with also comorbid panic and depersonalization-derealization disorder. Besides, unipolar patients suffering from dissociative symptoms have more often cyclothymic affective temperament. Despite the limits of such studies dissociative symptoms-BD association seems to correspond to a clinical reality and further works on this topic may be warranted.

  14. High-affinity AKAP7δ–protein kinase A interaction yields novel protein kinase A-anchoring disruptor peptides

    PubMed Central

    Hundsrucker, Christian; Krause, Gerd; Beyermann, Michael; Prinz, Anke; Zimmermann, Bastian; Diekmann, Oliver; Lorenz, Dorothea; Stefan, Eduard; Nedvetsky, Pavel; Dathe, Margitta; Christian, Frank; Mcsorley, Theresa; Krause, Eberhard; Mcconnachie, George; Herberg, Friedrich W.; Scott, John D.; Rosenthal, Walter; Klussmann, Enno

    2006-01-01

    PKA (protein kinase A) is tethered to subcellular compartments by direct interaction of its regulatory subunits (RI or RII) with AKAPs (A kinase-anchoring proteins). AKAPs preferentially bind RII subunits via their RII-binding domains. RII-binding domains form structurally conserved amphipathic helices with unrelated sequences. Their binding affinities for RII subunits differ greatly within the AKAP family. Amongst the AKAPs that bind RIIα subunits with high affinity is AKAP7δ [AKAP18δ; Kd (equilibrium dissociation constant) value of 31 nM]. An N-terminally truncated AKAP7δ mutant binds RIIα subunits with higher affinity than the full-length protein presumably due to loss of an inhibitory region [Henn, Edemir, Stefan, Wiesner, Lorenz, Theilig, Schmidtt, Vossebein, Tamma, Beyermann et al. (2004) J. Biol. Chem. 279, 26654–26665]. In the present study, we demonstrate that peptides (25 amino acid residues) derived from the RII-binding domain of AKAP7δ bind RIIα subunits with higher affinity (Kd=0.4±0.3 nM) than either full-length or N-terminally truncated AKAP7δ, or peptides derived from other RII binding domains. The AKAP7δ-derived peptides and stearate-coupled membrane-permeable mutants effectively disrupt AKAP–RII subunit interactions in vitro and in cell-based assays. Thus they are valuable novel tools for studying anchored PKA signalling. Molecular modelling indicated that the high affinity binding of the amphipathic helix, which forms the RII-binding domain of AKAP7δ, with RII subunits involves both the hydrophobic and the hydrophilic faces of the helix. Alanine scanning (25 amino acid peptides, SPOT technology, combined with RII overlay assays) of the RII binding domain revealed that hydrophobic amino acid residues form the backbone of the interaction and that hydrogen bond- and salt-bridge-forming amino acid residues increase the affinity of the interaction. PMID:16483255

  15. A set of robust fluorescent peptide probes for quantification of Cu(ii) binding affinities in the micromolar to femtomolar range.

    PubMed

    Young, Tessa R; Wijekoon, Chathuri J K; Spyrou, Benjamin; Donnelly, Paul S; Wedd, Anthony G; Xiao, Zhiguang

    2015-03-01

    Reliable quantification of copper binding affinities and identification of the binding sites provide a molecular basis for an understanding of the nutritional roles and toxic effects of copper ions. Sets of chromophoric probes are now available that can quantify Cu(i) binding affinities from nanomolar to attomolar concentrations on a unified scale under in vitro conditions. Equivalent probes for Cu(ii) are lacking. This work reports development of a set of four fluorescent dansyl peptide probes (DP1-4) that can quantify Cu(ii) binding affinities from micromolar to femtomolar concentrations, also on a unified scale. The probes were constructed by conjugation of a dansyl group to four short peptides of specific design. Each was characterised by its dissociation constant KD, its pH dependence and the nature of its binding site. One equivalent of Cu(ii) is bound by the individual probes that display different and well-separated affinities at pH 7.4 (log KD = -8.1, -10.1, -12.3 and -14.1, respectively). Intense fluorescence is emitted at λmax ∼ 550 nm upon excitation at ∼330 nm. Binding of Cu(ii) quenches the fluorescence intensity linearly until one equivalent of Cu(ii) is bound. Multiple approaches and multiple affinity standards were employed to ensure reliability. Selected examples of application to well-characterised Cu(ii) binding peptides and proteins are presented. These include Aβ16 peptides, two naturally occurring Cu(ii)-chelating motifs in human serum and cerebrospinal fluid with sequences GHK and DAHK and two copper binding proteins, CopC from Pseudomonas syringae and PcoC from Escherichia coli. Previously reported affinities are reproduced, demonstrating that peptides DP1-4 form a set of robust and reliable probes for Cu(ii) binding to peptides and protein targets.

  16. Kinetic rate constant prediction supports the conformational selection mechanism of protein binding.

    PubMed

    Moal, Iain H; Bates, Paul A

    2012-01-01

    The prediction of protein-protein kinetic rate constants provides a fundamental test of our understanding of molecular recognition, and will play an important role in the modeling of complex biological systems. In this paper, a feature selection and regression algorithm is applied to mine a large set of molecular descriptors and construct simple models for association and dissociation rate constants using empirical data. Using separate test data for validation, the predicted rate constants can be combined to calculate binding affinity with accuracy matching that of state of the art empirical free energy functions. The models show that the rate of association is linearly related to the proportion of unbound proteins in the bound conformational ensemble relative to the unbound conformational ensemble, indicating that the binding partners must adopt a geometry near to that of the bound prior to binding. Mirroring the conformational selection and population shift mechanism of protein binding, the models provide a strong separate line of evidence for the preponderance of this mechanism in protein-protein binding, complementing structural and theoretical studies.

  17. Tityus serrulatus venom contains two classes of toxins. Tityus gamma toxin is a new tool with a very high affinity for studying the Na+ channel.

    PubMed

    Barhanin, J; Giglio, J R; Léopold, P; Schmid, A; Sampaio, S V; Lazdunski, M

    1982-11-10

    The interaction of TiTx gamma, the major toxin in the venom of the scorpion Tityus serrulatus, with its receptor in excitable membranes was studied with the use of 125I-TiTx gamma. This derivative retains biological activity, and its specific binding to both brain synaptosomes and electroplaque membranes from Electrophorus electricus is characterized by a dissociation constant equal to that of the native toxin-receptor complex, about 2 to 5 pM. This very high affinity results mainly from a very slow rate of dissociation, equivalent to a half-life longer than 10 h at 4 degrees C. There is a 1:1 stoichiometry between TiTx gamma binding and tetrodotoxin binding to the membranes, but neither tetrodotoxin nor any of 7 other neurotoxins that are representative of 4 different classes of effectors of the Na+ channel interfere with TiTx gamma binding. Similarly, local anesthetics and other molecules that affect other types of ionic channels or neurotransmitter receptors have no effect on TiTx gamma binding. However, toxin II from Centruroides suffusus suffusus does compete with TiTx gamma, though its affinity for the receptor is much lower. Since the Centruroides toxin II is known to affect Na+ channel function, these two scorpion toxins must be put into a fifth class of Na+ channel effectors.

  18. Characterization of the ER-Targeted Low Affinity Ca2+ Probe D4ER

    PubMed Central

    Greotti, Elisa; Wong, Andrea; Pozzan, Tullio; Pendin, Diana; Pizzo, Paola

    2016-01-01

    Calcium ion (Ca2+) is a ubiquitous intracellular messenger and changes in its concentration impact on nearly every aspect of cell life. Endoplasmic reticulum (ER) represents the major intracellular Ca2+ store and the free Ca2+ concentration ([Ca2+]) within its lumen ([Ca2+]ER) can reach levels higher than 1 mM. Several genetically-encoded ER-targeted Ca2+ sensors have been developed over the last years. However, most of them are non-ratiometric and, thus, their signal is difficult to calibrate in live cells and is affected by shifts in the focal plane and artifactual movements of the sample. On the other hand, existing ratiometric Ca2+ probes are plagued by different drawbacks, such as a double dissociation constant (Kd) for Ca2+, low dynamic range, and an affinity for the cation that is too high for the levels of [Ca2+] in the ER lumen. Here, we report the characterization of a recently generated ER-targeted, Förster resonance energy transfer (FRET)-based, Cameleon probe, named D4ER, characterized by suitable Ca2+ affinity and dynamic range for monitoring [Ca2+] variations within the ER. As an example, resting [Ca2+]ER have been evaluated in a known paradigm of altered ER Ca2+ homeostasis, i.e., in cells expressing a mutated form of the familial Alzheimer’s Disease-linked protein Presenilin 2 (PS2). The lower Ca2+ affinity of the D4ER probe, compared to that of the previously generated D1ER, allowed the detection of a conspicuous, more clear-cut, reduction in ER Ca2+ content in cells expressing mutated PS2, compared to controls. PMID:27598166

  19. KW-3902, a selective high affinity antagonist for adenosine A1 receptors.

    PubMed Central

    Nonaka, H.; Ichimura, M.; Takeda, M.; Kanda, T.; Shimada, J.; Suzuki, F.; Kase, H.

    1996-01-01

    1. We demonstrate that 8-(noradamantan-3-yl)-1,3-dipropylxanthine (KW-3902) is a very potent and selective adenosine A1 receptor antagonist, assessed by radioligand binding and cyclic AMP response in cells. 2. In rat forebrain adenosine A1 receptors labelled with [3H]-cyclohexyladenosine (CHA), KW-3902 had a Ki value of 0.19 nM, whereas it showed a Ki value of 170 nM in rat striatal A2A receptors labelled with [3H]-2-[p-(2-carboxyethyl)-phenethylamino]-5'-N-ethylcarboxamidoad enosine (CGS21680), indicating 890 fold A1 receptor selectivity versus the A2A receptor. KW-3902 at 10 microM showed no effect on recombinant rat A3 receptors expressed on CHO cells. 3. Saturation studies with [3H]-KW-3902 revealed that it bound with high affinity (Kd = 77 pM) and limited capacity (Bmax = 470 fmol mg-1 of protein) to a single class of recognition sites. A high positive correlation was observed between the pharmacological profile of adenosine ligands inhibiting the binding of [3H]-KW-3902 and that of [3H]-CHA. 4. KW-3902 showed potent A1 antagonism against the inhibition of forskolin-induced cyclic AMP accumulation in DDT1 MF-2 cells by the A1-selective agonist, cyclopentyladenosine with a dissociation constant (KB value) of 0.34 nM. KW-3902 antagonized 5'-N-ethylcarboxamidoadenosine-elicited cyclic AMP accumulation via A2B receptors with a KB value of 52 nM. 5. KW-3902 exhibited marked species-dependent differences in the binding affinities. The highest affinity was for the rat A1 receptor (ki = 0.19 nM) and these values for guinea-pig and dog A1 receptors were 1.3 and 10 nM, respectively. PMID:8732272

  20. Isolation, identification and characterisation of starch-interacting proteins by 2-D affinity electrophoresis.

    PubMed

    Kosar-Hashemi, Behjat; Irwin, Jennifer A; Higgins, Jody; Rahman, Sadequr; Morell, Matthew K

    2006-05-01

    A 2-D affinity electrophoretic technique (2-DAE) has been used to isolate proteins that interact with various starch components from total barley endosperm extracts. In the first dimension, proteins are separated by native PAGE. The second-dimensional gel contains polysaccharides such as amylopectin and glycogen. The migration of starch-interacting proteins in this dimension is determined by their affinity towards a particular polysaccharide and these proteins are therefore spatially separated from the bulk of proteins in the crude extract. Four distinct proteins demonstrate significant affinity for amylopectin and have been identified as starch branching enzyme I (SBEI), starch branching enzyme IIa (SBEIIa), SBEIIb and starch phosphorylase using polyclonal antibodies and zymogram activity analysis. In the case of starch phosphorylase, a protein spot was excised from a 2-DAE polyacrylamide gel and analysed using Q-TOF MS/MS, resulting in the alignment of three internal peptide sequences with the known sequence of the wheat plastidic starch phosphorylase isoform. This assignment was confirmed by the determination of the enzyme's function using zymogram analysis. Dissociation constants (Kd) were calculated for the three enzymes at 4 degrees C and values of 0.20, 0.21 and 1.3 g/L were determined for SBEI, SBEIIa and starch phosphorylase, respectively. Starch synthase I could also be resolved from the other proteins in the presence of glycogen and its identity was confirmed using a polyclonal antibody and by activity analysis. The 2-DAE method described here is simple, though powerful, enabling protein separation from crude extracts on the basis of function.

  1. High-affinity antibodies to the 1,4-dihydropyridine Ca2+-channel blockers

    SciTech Connect

    Campbell, K.P.; Sharp, A.; Strom, M.; Kahl, S.D.

    1986-05-01

    Antibodies with high affinity and specificity for the 1,4-dihydropyridine Ca2+-channel blockers have been produced in rabbits by immunization with dihydropyridine-protein conjugates. Anti-dihydropyridine antibodies were found to specifically bind (/sup 3/H)nitrendipine, (/sup 3/H)-nimodipine, (/sup 3/H)nisoldipine, and (/sup 3/H)PN 200-110 (all 1,4-dihydropyridine Ca2+-channel blockers) with high affinity, while (/sup 3/H)verapamil, (/sup 3/H)diltiazem, and (/sup 3/H)trifluoperazine were not recognized. The average dissociation constant of the (/sup 3/H)nitrendipine-antibody complex was 0.06 (+/- 0.02) X 10(-9) M for an antiserum studied in detail and ranged from 0.01 to 0.24 X 10(-9) M for all antisera. Inhibition of (/sup 3/H)nitrendipine binding was specific for the 1,4-dihydropyridine Ca2+-channel modifiers and the concentrations required for half-maximal inhibition ranged between 0.25 and 0.90 nM. Structurally unrelated Ca2+-channel blockers, calmodulin antagonists, inactive metabolites of nitrendipine, and UV-inactivated nisoldipine did not modify (/sup 3/H)nitrendipine binding to the anti-dihydropyridine antibodies. Dihydropyridines without a bulky substituent in the 4-position of the heterocycle were able to displace (/sup 3/H)nitrendipine binding, but the concentrations required for half-maximal inhibition were greater than 800 nM. In summary, anti-dihydropyridine antibodies have been shown to have high affinity and specificity for the 1,4-dihydropyridine Ca2+-channel blockers and to exhibit dihydropyridine binding properties similar to the membrane receptor for the 1,4-dihydropyridine Ca2+-channel blockers.

  2. Hyperglycemia associated dissociative fugue (organic dissociative disorder) in an elderly.

    PubMed

    Ram, Dushad; Ashoka, H G; Gowdappa, Basavnna

    2015-01-01

    Inadequate glycemic control in patients with diabetes is known to be associated with psychiatric disorders such as depression, anxiety disorder, and cognitive impairment. However, dissociative syndrome has not been reported so far. Here we are reporting a case of repeated dissociative fugue associated with hyperglycemia, in an elderly with type II diabetes. Possible neurobiological mechanism has been discussed.

  3. Ionic molal conductivities, activity coefficients, and dissociation constants of HAsO42− and H2AsO4− from 5 to 90°C and ionic strengths from 0.001 up to 3 mol kg−1 and applications in natural systems

    USGS Publications Warehouse

    Zhu, Xiangyu; Nordstrom, D. Kirk; McCleskey, R. Blaine; Wang, Rucheng

    2016-01-01

    Arsenic is known to be one of the most toxic inorganic elements, causing worldwide environmental contamination. However, many fundamental properties related to aqueous arsenic species are not well known which will inhibit our ability to understand the geochemical behavior of arsenic (e.g. speciation, transport, and solubility). Here, the electrical conductivity of Na2HAsO4 solutions has been measured over the concentration range of 0.001–1 mol kg−1 and the temperature range of 5–90°C. Ionic strength and temperature-dependent equations were derived for the molal conductivity of HAsO42−and H2AsO4− aqueous ions. Combined with speciation calculations and the approach used by McCleskey et al. (2012b), these equations can be used to calculate the electrical conductivities of arsenic-rich waters having a large range of effective ionic strengths (0.001–3 mol kg−1) and temperatures (5–90°C). Individual ion activity coefficients for HAsO42− and H2AsO4− in the form of the Hückel equation were also derived using the mean salt method and the mean activity coefficients of K2HAsO4 (0.001–1 mol kg−1) and KH2AsO4 (0.001–1.3 mol kg−1). A check on these activity coefficients was made by calculating mean activity coefficients for Na2HAsO4 and NaH2AsO4 solutions and comparing them to measured values. At the same time Na-arsenate complexes were evaluated. The NaH2AsO40 ion pair is negligible in NaH2AsO4 solutions up to 1.3 mol kg−1. The NaHAsO4− ion pair is important in NaHAsO4 solutions >0.1 mol kg−1 and the formation constant of 100.69 was confirmed. The enthalpy, entropy, free energy and heat capacity for the second and third arsenic acid dissociation reactions were calculated from pH measurements. These properties have been incorporated into a widely used geochemical calculation code WATEQ4F and applied to natural arsenic waters. For arsenic spiked water samples from Yellowstone National Park, the mean difference between the

  4. Nanofluidic Fluorescence Microscopy (NFM) for real-time monitoring of protein binding kinetics and affinity studies.

    PubMed

    Teerapanich, Pattamon; Pugnière, Martine; Henriquet, Corinne; Lin, Yii-Lih; Chou, Chia-Fu; Leïchlé, Thierry

    2017-02-15

    Kinetic monitoring of protein interactions offers insights to their corresponding functions in cellular processes. Surface plasmon resonance (SPR) is the current standard tool used for label-free kinetic assays; however, costly and sophisticated setups are required, decreasing its accessibility to research laboratories. We present a cost-effective nanofluidic-based immunosensor for low-noise real-time kinetic measurement of fluorescent-labeled protein binding. With the combination of fluorescence microscopy and reversed buffer flow operation, association and dissociation kinetics can be accessed in one single experiment without extra buffer loading step, which results in a simplified operation and reduced time of analysis compared to typical microfluidic immunoassays. Kinetic constants of two representative protein-ligand binding pairs (streptavidin/biotin; IgG/anti-IgG) were quantified. The good agreement of extracted rate constants with literature values and analogous SPR measurements indicates that this approach is applicable to study protein interactions of medium- and high-affinities with a limit of detection down to 1 pM, regardless of the analyte size.

  5. Affinity labeling of soybean beta-amylase with 2',3'-epoxypropyl alpha-D-glucopyranoside.

    PubMed

    Isoda, Y; Nitta, Y

    1986-06-01

    The synthesized 2',3'-epoxypropyl alpha-D-glucopyranoside (alpha-EPG) inactivated soybean beta-amylase completely. The incorporation of alpha-EPG into the enzyme at 92% inactivation was 1.1 mol per mol of enzyme, as determined by using 14C-labeled alpha-EPG. The inactivation obeyed saturation kinetics of a two-step mechanism. The dissociation constant of alpha-EPG-enzyme complex and the rate constant of the irreversible inactivation step were estimated to be 119 mM and 1.14 X 10(-3)s-1, respectively. alpha-Cyclodextrin, a competitive inhibitor of this enzyme, protected the enzyme against the inactivation by alpha-EPG in a competitive manner. This suggests that alpha-EPG binds to the active site of the enzyme. The above results indicate that alpha-EPG acts on soybean beta-amylase as an affinity labeling reagent. It was also shown that an essential SH group near the active site, but not the catalytic one, scarcely participated in the inactivation by alpha-EPG.

  6. Four-body atomic potential for modeling protein-ligand binding affinity: application to enzyme-inhibitor binding energy prediction

    PubMed Central

    2013-01-01

    Background Models that are capable of reliably predicting binding affinities for protein-ligand complexes play an important role the field of structure-guided drug design. Methods Here, we begin by applying the computational geometry technique of Delaunay tessellation to each set of atomic coordinates for over 1400 diverse macromolecular structures, for the purpose of deriving a four-body statistical potential that serves as a topological scoring function. Next, we identify a second, independent set of three hundred protein-ligand complexes, having both high-resolution structures and known dissociation constants. Two-thirds of these complexes are randomly selected to train a predictive model of binding affinity as follows: two tessellations are generated in each case, one for the entire complex and another strictly for the isolated protein without its bound ligand, and a topological score is computed for each tessellation with the four-body potential. Predicted protein-ligand binding affinity is then based on an empirically derived linear function of the difference between both topological scores, one that appropriately scales the value of this difference. Results A comparison between experimental and calculated binding affinity values over the two hundred complexes reveals a Pearson's correlation coefficient of r = 0.79 with a standard error of SE = 1.98 kcal/mol. To validate the method, we similarly generated two tessellations for each of the remaining protein-ligand complexes, computed their topological scores and the difference between the two scores for each complex, and applied the previously derived linear transformation of this topological score difference to predict binding affinities. For these one hundred complexes, we again observe a correlation of r = 0.79 (SE = 1.93 kcal/mol) between known and calculated binding affinities. Applying our model to an independent test set of high-resolution structures for three hundred diverse enzyme-inhibitor complexes

  7. Selective induction of high-ouabain-affinity isoform of Na sup + -K sup + -ATPase by thyroid hormone

    SciTech Connect

    Haber, R.S.; Loeb, J.N. )

    1988-12-01

    The administration of thyroid hormone is known to result in an induction of the Na{sup +}-K{sup +}-adenosinetriphosphatase (Na{sup +}-K{sup +}-ATPase) in rat skeletal muscle and other thyroid hormone-responsive tissues. Since the Na{sup +}-K{sup +}-ATPase in a variety of mammalian tissues has recently been reported to exist in at least two forms distinguishable by differing affinities for the inhibitory cardiac glycoside ouabain. The authors have studied the effects of 3,3{prime},5-triiodo-L-thyronine (T{sub 3}) treatment on these two forms of the enzyme in rat diaphragm. The inhibition of Na{sup +}-K{sup +}-ATPase activity in a crude membrane fraction by varying concentrations of ouabain conformed to a biphasic pattern consistent with the presence of two distinct isoforms with inhibition constants (K{sub I}s) for ouabain of {approximately}10{sup {minus}7} and 10{sup {minus}4} M, respectively. Measurement of the specific binding of ({sup 3}H)ouabain to these membranes confirmed the presence of a class of high-affinity ouabain binding sites with a dissociation constant (K{sub d}) of slightly less than 10{sup {minus}7}M, whose maximal binding capacity was increased by T{sub 3} treatment by 185%. Binding studies in unfractionated homogenates of diaphragm similarly demonstrated the presence of high-affinity sites whose maximal binding capacity was increased by T{sub 3} treatment. Quantitation of both the high- and low-ouabain-affinity forms of the Na{sup +}-K{sup +}-ATPase by ouabain-dependent phosphorylation from ({sup 32}P)orthophosphate confirmed that T{sub 3} treatment markedly increased the number of high-affinity sites while having little effect on the number of low-affinity sites. These observations provide, to our knowledge, the first demonstration that these two forms of the Na{sup +}-K{sup +}-ATPase are subject to selective hormonal induction.

  8. Dissociative States and Neural Complexity

    ERIC Educational Resources Information Center

    Bob, Petr; Svetlak, Miroslav

    2011-01-01

    Recent findings indicate that neural mechanisms of consciousness are related to integration of distributed neural assemblies. This neural integration is particularly vulnerable to past stressful experiences that can lead to disintegration and dissociation of consciousness. These findings suggest that dissociation could be described as a level of…

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

    PubMed

    Baldwin, Graham S; George, Graham N; Pushie, M Jake

    2015-01-01

    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.

  10. Binding constants determined from Ca2+ current responses to rapid applications and washouts of nifedipine in frog cardiac myocytes.

    PubMed

    Méry, P F; Hove-Madsen, L; Mazet, J L; Hanf, R; Fischmeister, R

    1996-07-01

    1. A fast perfusion system was used to analyse the kinetics of the response of L-type calcium current (ICa) to rapid applications and washouts of the dihydropyridine antagonist nifedipine in whole-cell patch-clamped frog ventricular myocytes. 2. Both the inhibition of ICa induced by nifedipine and the recovery from inhibition upon washout of the drug behaved as mono-exponential functions of time. 3. During application or washout of 100 nM nifedipine, only the peak amplitude of ICa varied but not its time course of activation or inactivation. 4. The rate constant of the onset of ICa inhibition increased with the concentration of nifedipine. However, the time course of the recovery from inhibition was independent of drug concentration. 5. Both rate constants were strongly sensitive to the holding potential but insensitive to the test potential. 6. Using simple rate equations and a one-binding-site analysis it was possible to determine the rate constants for association (k1) and dissociation (k-1) and the equilibrium dissociation constant (KD) of the reaction between nifedipine and Ca2+ channels. KD values for nifedipine were identical to IC50 values obtained from classical steady-state experiments. 7. With depolarized holding potentials, KD decreased strongly due to a large reduction in k-1 and a modest increase in k1. Assuming that these changes result from the distribution of Ca2+ channels between resting and inactivated states, a low-affinity binding to the resting state (R) and a high-affinity binding to the inactivated state (I) were obtained with the binding constants: k1R = 1.0 x 10(6) M-1 S-1, k-1R = 0.077 S-1, and KDR = 77 nM for the resting state; k1I = 4.47 x 10(6) M-1 S-1, k-1I = 7.7 x 10(-4) S-1, and KDI = 0.17 nM for the inactivated state. 8. Rapid application/washout experiments provide a unique way to determine, in an intact cell and in a relatively short period (2-4 min), the binding rate constants and the KD value of the reaction between a

  11. Quantitative analysis of the high-affinity binding sites for [3H]ouabain in the rat vas deferens and their immunological identification as the alpha 2 isoform of Na+/K(+)-ATPase.

    PubMed

    Noël, F; Quintas, L E; Freitas, E; Caricati-Neto, A; Lafayette, S S; Wanderley, A G; Jurkiewicz, A

    1998-05-01

    Binding assays were performed with [3H]ouabain to investigate the presence of, and to characterize, a Na+/K(+)-ATPase isoform with high affinity for cardiac glycosides in the rat vas deferens. Nonlinear regression analysis of equilibrium experiments carried out with crude preparations in a Mg-Pi medium indicated the presence of high-affinity sites characterized with good precision (individual coefficients of variation = 11-35%) by their density (Bmax = 0.42 to 0.72 pmol/mg protein) and dissociation constant (Kd = 0.069 to 0.136 microM) values. The values of the dissociation rate constant (kappa-1) and the association rate constant (kappa+1) for these sites were 0.151 to 0.267 min-1 and 2.87 to 3.60 microM-1.min-1, respectively. A higher number of low-affinity sites (Kd around 15 microM), supposed to correspond to the alpha 1 isoform, was also identified, but their Kd and Bmax values were not quantified precisely in this crude preparation. Western blot assays indicated hybridization with specific anti-alpha 1 and anti-alpha 2 isoform antibodies but not with anti-alpha 3 isoform antibody. Taken together, the present results indicate the existence of a low proportion of the alpha 2 isoform of Na+/K(+)-ATPase in the rat vas deferens that can be quantified precisely by [3H]ouabain binding even in a crude membrane preparation that is suitable for studies under conditions of plasticity.

  12. Effect of heme modification on oxygen affinity of myoglobin and equilibrium of the acid-alkaline transition in metmyoglobin.

    PubMed

    Shibata, Tomokazu; Nagao, Satoshi; Fukaya, Masashi; Tai, Hulin; Nagatomo, Shigenori; Morihashi, Kenji; Matsuo, Takashi; Hirota, Shun; Suzuki, Akihiro; Imai, Kiyohiro; Yamamoto, Yasuhiko

    2010-05-05

    Functional regulation of myoglobin (Mb) is thought to be achieved through the heme environment furnished by nearby amino acid residues, and subtle tuning of the intrinsic heme Fe reactivity. We have performed substitution of strongly electron-withdrawing perfluoromethyl (CF(3)) group(s) as heme side chain(s) of Mb to obtain large alterations of the heme electronic structure in order to elucidate the relationship between the O(2) affinity of Mb and the electronic properties of heme peripheral side chains. We have utilized the equilibrium constant (pK(a)) of the "acid-alkaline transition" in metmyoglobin in order to quantitatively assess the effects of the CF(3) substitutions for the electron density of heme Fe atom (rho(Fe)) of the protein. The pK(a) value of the protein was found to decrease by approximately 1 pH unit upon the introduction of one CF(3) group, and the decrease in the pK(a) value with decreasing the rho(Fe) value was confirmed by density functional theory calculations on some model compounds. The O(2) affinity of Mb was found to correlate well with the pK(a) value in such a manner that the P(50) value, which is the partial pressure of O(2) required to achieve 50% oxygenation, increases by a factor of 2.7 with a decrease of 1 pK(a) unit. Kinetic studies on the proteins revealed that the decrease in O(2) affinity upon the introduction of an electron-withdrawing CF(3) group is due to an increase in the O(2) dissociation rate. Since the introduction of a CF(3) group substitution is thought to prevent further Fe(2+)-O(2) bond polarization and hence formation of a putative Fe(3+)-O(2)(-)-like species of the oxy form of the protein [Maxwell, J. C.; Volpe, J. A.; Barlow, C. H.; Caughey, W. S. Biochem. Biophys. Res. Commun. 1974, 58, 166-171], the O(2) dissociation is expected to be enhanced by the substitution of electron-withdrawing groups as heme side chains. We also found that, in sharp contrast to the case of the O(2) binding to the protein, the CO

  13. Emergent properties of nanosensor arrays: applications for monitoring IgG affinity distributions, weakly affined hypermannosylation, and colony selection for biomanufacturing.

    PubMed

    Reuel, Nigel F; Grassbaugh, Brittany; Kruss, Sebastian; Mundy, J Zachary; Opel, Cary; Ogunniyi, Adebola O; Egodage, Kamal; Wahl, Ramon; Helk, Bernhard; Zhang, Jingqing; Kalcioglu, Z Ilke; Tvrdy, Kevin; Bellisario, Darin O; Mu, Bin; Blake, Steven S; Van Vliet, Krystyn J; Love, J Christopher; Wittrup, Karl Dane; Strano, Michael S

    2013-09-24

    It is widely recognized that an array of addressable sensors can be multiplexed for the label-free detection of a library of analytes. However, such arrays have useful properties that emerge from the ensemble, even when monofunctionalized. As examples, we show that an array of nanosensors can estimate the mean and variance of the observed dissociation constant (KD), using three different examples of binding IgG with Protein A as the recognition site, including polyclonal human IgG (KD μ = 19 μM, σ(2) = 1000 mM(2)), murine IgG (KD μ = 4.3 nM, σ(2) = 3 μM(2)), and human IgG from CHO cells (KD μ = 2.5 nM, σ(2) = 0.01 μM(2)). Second, we show that an array of nanosensors can uniquely monitor weakly affined analyte interactions via the increased number of observed interactions. One application involves monitoring the metabolically induced hypermannosylation of human IgG from CHO using PSA-lectin conjugated sensor arrays where temporal glycosylation patterns are measured and compared. Finally, the array of sensors can also spatially map the local production of an analyte from cellular biosynthesis. As an example, we rank productivity of IgG-producing HEK colonies cultured directly on the array of nanosensors itself.

  14. Direct labelling of myocardial beta 1-adrenoceptors. Comparison of binding affinity of 3H-(-)-bisoprolol with its blocking potency.

    PubMed

    Kaumann, A J; Lemoine, H

    1985-10-01

    A radioligand that selectively labels beta 1-adrenoceptors, 3H-(-)-bisoprolol (3H-BIS), is introduced. The binding properties of 3H-BIS to membrane particles of kitten heart are compared with the blocking properties of (-)-bisoprolol against stimulant effects of (-)-adrenaline and (-)-noradrenaline in heart preparations of kitten and guinea pig. 1. On kitten heart tissues (-)-bisoprolol antagonized the positive chronotropic and inotropic effects of catecholamines competitively. The effects of (-)-adrenaline were antagonized considerably less by (-)-bisoprolol than the corresponding effects of (-)-noradrenaline on sinoatrial pacemakers. The antagonism was nearly the same against both (-)-adrenaline and (-)-noradrenaline in left atria and papillary muscles. The data were analyzed with a model for 2-receptor subtypes by non-linear regression. Equilibrium dissociation constants KB (mol/l; -log KB = pKB) for a high-affinity beta 1-adrenoceptor of 8.8 and for a low-affinity beta 2-adrenoceptor of 7.0 were estimated in the three classes of tissues. In kitten sinoatrial pacemaker beta 1-adrenoceptors contribute 76% to the stimulus induced by (-)-adrenaline and 97% to the stimulus induced by (-)-noradrenaline. In ventricle and left atrium beta 1-adrenoceptors contribute 97-99% and 100% to the stimulus caused by (-)-adrenaline and (-)-noradrenaline, respectively. 2. Both 3H-BIS and unlabelled (-)-bisoprolol caused competitive blockade of the positive chronotropic effects of (-)-noradrenaline in guinea-pig right atria. pKB-values of 8.7 were estimated for both unlabelled and tritiated (-)-bisoprolol. The positive chronotropic effects of (-)-adrenaline were antagonized considerably less by (-)-bisoprolol than those of (-)-noradrenaline in guinea-pig atria. In the presence of low concentrations of beta 2-selective ICI 118,551, which did not antagonize beta 1-adrenoceptor mediated effects, (-)-bisoprolol antagonized positive chronotropic effects of (-)-adrenaline to the same

  15. Shock induced dissociation of polyethylene

    SciTech Connect

    Morris, C.E.; Loughran, E.D.; Mortensen, G.F.; Gray, G.T. III; Shaw, M.S.

    1989-01-01

    To identify the physical processes occurring on the Hugoniot, shock-recovery experiments were performed. Cylindrical recovery systems were used that enabled a wide range of single-shock Hugoniot states to be examined. Mass spectroscopy was used to examine the gaseous dissociation products. X-ray and TEM measurements were made to characterize the post-shock carbon structures. A dissociation product equation of state is presented to interpret the observed results. Polyethylene (PE) samples that were multiply shocked to their final states dissociated at much higher pressures than single-shocked samples. 5 refs., 2 figs., 1 tab.

  16. Chiral separation of new sulfonamide derivatives and evaluation of their enantioselective affinity for human carbonic anhydrase II by microscale thermophoresis and surface plasmon resonance.

    PubMed

    Rogez-Florent, Tiphaine; Foulon, Catherine; Drucbert, Anne-Sophie; Schifano, Nadège; Six, Perrine; Devassine, Stéphanie; Depreux, Patrick; Danzé, Pierre-Marie; Goossens, Laurence; Danel, Cécile; Goossens, Jean-François

    2017-04-15

    The aim of this study was to develop a method combining chiral separation and biophysical techniques to evaluate the enantioselective affinity of original sulfonamide derivatives towards their therapeutic target, the human carbonic anhydrase II (hACII). The first step consisted in the preparation of the enantiomers by chromatographic separation. The performances of HPLC and Supercritical Fluid Chromatography (SFC) were studied at the analytical scale by optimization of various experimental conditions using adsorbed polysaccharide chiral stationary phases (amylose AD-H and cellulose OD-H). Since SFC allowed obtaining higher enantioresolutions per time unit, it was selected for the semi-preparative scale and successfully used to isolate each enantiomer with a satisfactory enantiomeric purity (>98%). Secondly, microscale thermophoresis (MST) method and surface plasmon resonance (SPR) used as reference method were developed to measure potential enantioselective affinities of these enantiomers towards the hACII. The optimizations of both methods were performed using a reference compound, i.e. acetazolamide, which affinity for hCAII has previously been demonstrated. For all compounds, KD values obtained using MST and SPR were in good agreement, leading to similar affinity scales despite both approaches totally differ (labeling for MST versus immobilization of the protein for SPR). The equilibrium dissociation constants of our original compounds for the hCAII were in the range 100-1000nM and an enantioselectivity was observed using the MST and SPR methods for the diarylpyrazole 2. Finally, by comparing the MST and SPR techniques, MST appears especially adapted for further screening of a series of sulfonamide derivatives due to the lower time required to estimate a binding constant while consuming as little hCAII as SPR.

  17. Familial secondary erythrocytosis due to increased oxygen affinity is caused by destabilization of the T state of hemoglobin Brigham (α2β2Pro100Leu)

    PubMed Central

    Mollan, Todd L; Abraham, Bindu; Strader, Michael Brad; Jia, Yiping; Lozier, Jay N; Olson, John S; Alayash, Abdu I

    2012-01-01

    Hemoglobin Brigham (β Pro100 to Leu) was first reported in a patient with familial erythrocytosis. Erythrocytes of an affected individual from the same family contain both HbA and Hb Brigham and exhibit elevated O2 affinity compared with normal cells (P50 = 23 mm Hg vs. 31 mmHg at pH 7.4 at 37°C). O2 affinities measured for hemolysates were sensitive to changes in pH or chloride concentrations, indicating little change in the Bohr and Chloride effects. Hb Brigham was separated from normal HbA by nondenaturing cation exchange liquid chromatography, and the amino acid substitution was verified by mass spectrometry. The properties of Hb Brigham isolated from the patient's blood were then compared with those of recombinant Hb Brigham expressed in Escherichia coli. Kinetic experiments suggest that the rate constants for ligand binding and release in the high (R) and low (T) affinity quaternary states of Hb Brigham are similar to those of native hemoglobin. However, the Brigham mutation decreases the T to R equilibrium constant (L) which accelerates the switch to the R state during ligand binding to deoxy-Hb, increasing the rate of association by approximately twofold, and decelerates the switch during ligand dissociation from HbO2, decreasing the rate approximately twofold. These kinetic data help explain the high O2 affinity characteristics of Hb Brigham and provide further evidence for the importance of the contribution of Pro100 to intersubunit contacts and stabilization of the T quaternary structure. PMID:22821886

  18. Magneto-nanosensor platform for probing low-affinity protein–protein interactions and identification of a low-affinity PD-L1/PD-L2 interaction

    PubMed Central

    Lee, Jung-Rok; Bechstein, Daniel J. B.; Ooi, Chin Chun; Patel, Ashka; Gaster, Richard S.; Ng, Elaine; Gonzalez, Lino C.; Wang, Shan X.

    2016-01-01

    Substantial efforts have been made to understand the interactions between immune checkpoint receptors and their ligands targeted in immunotherapies against cancer. To carefully characterize the complete network of interactions involved and the binding affinities between their extracellular domains, an improved kinetic assay is needed to overcome limitations with surface plasmon resonance (SPR). Here, we present a magneto-nanosensor platform integrated with a microfluidic chip that allows measurement of dissociation constants in the micromolar-range. High-density conjugation of magnetic nanoparticles with prey proteins allows multivalent receptor interactions with sensor-immobilized bait proteins, more closely mimicking natural-receptor clustering on cells. The platform has advantages over traditional SPR in terms of insensitivity of signal responses to pH and salinity, less consumption of proteins and better sensitivities. Using this platform, we characterized the binding affinities of the PD-1—PD-L1/PD-L2 co-inhibitory receptor system, and discovered an unexpected interaction between the two known PD-1 ligands, PD-L1 and PD-L2. PMID:27447090

  19. Are major dissociative disorders characterized by a qualitatively different kind of dissociation?

    PubMed

    Rodewald, Frauke; Dell, Paul F; Wilhelm-Gossling, Claudia; Gast, Ursula

    2011-01-01

    A total of 66 patients with a major dissociative disorder, 54 patients with nondissociative disorders, and 30 nonclinical controls were administered the Structured Clinical Interview for DSM-IV Dissociative Disorders-Revised, the Dissociative Experiences Scale, the Multidimensional Inventory of Dissociation, and the Symptom Checklist 90-Revised. Dissociative patients reported significantly more dissociative and nondissociative symptoms than did nondissociative patients and nonclinical controls. When general psychopathology was controlled, the dissociation scores of dissociative patients were still significantly higher than those of both other groups, whereas the dissociation scores of nondissociative patients and nonclinical controls no longer differed. These findings appear to be congruent with a typological model of dissociation that distinguishes between 2 qualitatively different kinds of dissociation. Specifically, the results of this study suggest that the dissociation that occurs in major dissociative disorders (i.e., dissociative identity disorder [DID] and dissociative disorder not otherwise specified, Type 1 [DDNOS-1]) is qualitatively different from the dissociation that occurs in persons who do not have a dissociative disorder. In contrast to previous research, the dissociation of persons who do not have a dissociative disorder is not limited to absorption; it covers a much wider range of phenomena. The authors hypothesize that different mechanisms produce the dissociation of persons with DID and DDNOS-1 as opposed to the dissociation of persons who do not have a dissociative disorder.

  20. Distinction between high-affinity (/sup 3/H)phencyclidine binding sites and muscarinic receptors in guinea-pig ileum muscle

    SciTech Connect

    El-Fakahany, E.E.; Triggle, D.J.; Eldefrawi, A.T.; Eldefrawi, M.E.

    1984-05-01

    (/sup 3/H)Phencyclidine ((/sup 3/H)PCP) binding was studied in guinea-pig ileum muscle membranes. Specific binding of (/sup 3/H)PCP was time dependent, reversible and saturable, with an equilibrium dissociation constant of 154 nM and maximum binding of 12.9 pmol/mg of protein at pH 9. Its pH dependency suggests that the un-ionized PCP is the pharmacologically active form. The binding site was on a protein which was sensitive to heat, proteolytic enzymes and the carboxylic group reagent dicyclohexylcarbodiimide, but insensitive to phospholipase A and C, concanavalin A, dithiothreitol and N-ethylmaleimide. Specific (/sup 3/H)PCP binding was displaced effectively by several PCP analogs and Ca/sup + +/ channel antagonists including verapamil, to which these sites had a high affinity. These high-affinity PCP-binding sites were found at a much higher concentration in the same membrane preparation than muscarinic receptor sites identified by their specific binding of (/sup 3/H)quinuclidinyl benzilate. PCP bound to both sites, but with a lower affinity to the muscarinic receptor sites. The PCP and muscarinic receptor sites differed in their sensitivities to pH and drug specifities.

  1. Influence of methylation on the properties of uracil and its noncovalent interactions with alkali metal ions: Threshold collision-induced dissociation and theoretical studies

    NASA Astrophysics Data System (ADS)

    Yang, Zhibo; Rodgers, M. T.

    2005-03-01

    The influence of methylation on the properties of uracil and its noncovalent interactions with alkali metal ions is investigated both experimentally and theoretically. Threshold collision-induced dissociation (CID) of M+(xMeU) with Xe is studied in a guided ion beam mass spectrometer. M+ include the following alkali metal ions: Li+, Na+, and K+. Five methylated uracils are examined, xMeU = 1-methyluracil, 3-methyluracil, 6-methyluracil, 1,3-dimethyluracil, and 5,6-dimethyluracil. In all cases endothermic loss of the intact nucleobase is the dominant reaction pathway, while ligand exchange to produce MXe+ is observed as a minor reaction pathway. The threshold regions of the cross sections are interpreted to extract 0 and 298 K bond dissociation energies (BDEs) for M+xMeU after accounting for the effects of multiple ion-neutral collisions, kinetic and internal energies of the reactants, and dissociation lifetimes. Ab initio calculations at the MP2(full)/6-31G* level of theory are used to determine the structures of these complexes and provide molecular constants required for the thermochemical analysis of the experimental data. Theoretical bond dissociation energies are determined from single point energy calculations at the MP2(full)/6-311+G(2d,2p) level using the MP2(full)/6-31G* geometries. Excellent agreement between theory and experiment is found for the Na+ and K+ systems, while theory systematically underestimates the strength of binding in the Li+ systems. Theoretical calculations are also performed to examine the influence of methylation on the acidities, proton affinities, and Watson-Crick base pairing energies. The present results are compared to earlier studies of uracil and 5-methyluracil to more fully elucidate the influence of methylation on the properties of uracil, its noncovalent interactions with alkali metal ions, and nucleic acid stability.

  2. Solubilization of high affinity corticotropin-releasing factor receptors from rat brain: Characterization of an active digitonin-solubilized receptor complex

    SciTech Connect

    Grigoriadis, D.E.; Zaczek, R.; Pearsall, D.M.; De Souza, E.B. )

    1989-12-01

    The binding characteristics of CRF receptors in rat frontal cerebral cortex membranes solubilized in 1% digitonin were determined. The binding of (125I)Tyro-ovine CRF ((125I)oCRF) to solubilized membrane proteins was dependent on incubation time, temperature, and protein concentration, was saturable and of high affinity, and was absent in boiled tissue. The solubilized receptors retained their high affinity for (125I) oCRF in the solubilized state, exhibiting a dissociation constant (KD) of approximately 200 pM, as determined by direct binding saturation isotherms. Solubilized CRF receptors maintained the rank order of potencies for various related and unrelated CRF peptides characteristic of the membrane CRF receptor: rat/human CRF congruent to ovine CRF congruent to Nle21,38-rat CRF greater than alpha-helical oCRF-(9-41) greater than oCRF-(7-41) much greater than vasoactive intestinal peptide, arginine vasopressin, or the substance-P antagonist. Furthermore, the absolute potencies (Ki values) for the various CRF-related peptides in solubilized receptors were almost identical to those observed in the membrane preparations, indicating that the CRF receptor retained its high affinity binding capacity in the digitonin-solubilized state. Chemical affinity cross-linking of digitonin-solubilized rat cortical membrane proteins revealed a specifically labeled protein with an apparent mol wt of 58,000 which was similar to the labeled protein in native membrane homogenates. Although solubilized CRF receptors retained their high affinity for agonists, their sensitivity for guanine nucleotide was lost. Size exclusion chromatography substantiated these results, demonstrating that in the presence or absence of guanine nucleotides, (125I)oCRF labeled the same size receptor complex.

  3. Somatostatin binding to dissociated cells from rat cerebral cortex

    SciTech Connect

    Colas, B.; Prieto, J.C.; Arilla, E. )

    1990-11-01

    A method has been developed for the study of somatostatin (SS) binding to dissociated cells from rat cerebral cortex. Binding of {sup 125}I (Tyr11)SS to cells obtained by mechanical dissociation of rat cerebral cortex was dependent on time and temperature, saturable, reversible and highly specific. Under conditions of equilibrium, i.e., 60 min at 25 degrees C, native SS inhibited tracer binding in a dose-dependent manner. The Scatchard analysis of binding data was linear and yielded a dissociation constant of 0.60 +/- 0.08 nM with a maximal binding capacity of 160 +/- 16 fmol/mg protein. The binding of {sup 125}I (Tyr11)SS was specific as shown in experiments on tracer displacement by the native peptides, SS analogues, and unrelated peptides.

  4. Single-molecule Imaging Analysis of Binding, Processive Movement, and Dissociation of Cellobiohydrolase Trichoderma reesei Cel6A and Its Domains on Crystalline Cellulose.

    PubMed

    Nakamura, Akihiko; Tasaki, Tomoyuki; Ishiwata, Daiki; Yamamoto, Mayuko; Okuni, Yasuko; Visootsat, Akasit; Maximilien, Morice; Noji, Hiroyuki; Uchiyama, Taku; Samejima, Masahiro; Igarashi, Kiyohiko; Iino, Ryota

    2016-10-21

    Trichoderma reesei Cel6A (TrCel6A) is a cellobiohydrolase that hydrolyzes crystalline cellulose into cellobiose. Here we directly observed the reaction cycle (binding, surface movement, and dissociation) of single-molecule intact TrCel6A, isolated catalytic domain (CD), cellulose-binding module (CBM), and CBM and linker (CBM-linker) on crystalline cellulose Iα The CBM-linker showed a binding rate constant almost half that of intact TrCel6A, whereas those of the CD and CBM were only one-tenth of intact TrCel6A. These results indicate that the glycosylated linker region largely contributes to initial binding on crystalline cellulose. After binding, all samples showed slow and fast dissociations, likely caused by the two different bound states due to the heterogeneity of cellulose surface. The CBM showed much higher specificity to the high affinity site than to the low affinity site, whereas the CD did not, suggesting that the CBM leads the CD to the hydrophobic surface of crystalline cellulose. On the cellulose surface, intact molecules showed slow processive movements (8.8 ± 5.5 nm/s) and fast diffusional movements (30-40 nm/s), whereas the CBM-Linker, CD, and a catalytically inactive full-length mutant showed only fast diffusional movements. These results suggest that both direct binding and surface diffusion contribute to searching of the hydrolysable point of cellulose chains. The duration time constant for the processive movement was 7.7 s, and processivity was estimated as 68 ± 42. Our results reveal the role of each domain in the elementary steps of the reaction cycle and provide the first direct evidence of the processive movement of TrCel6A on crystalline cellulose.

  5. Dissociation Rates of Diatomic Molecules

    DTIC Science & Technology

    1992-12-01

    relatively simple design that we have used extensively in earlier research; its characteristics were recently described in detail. 10 This source is by...the dominant dissociation background when observing electron- impact dissociation products. This background constitutes the single greatest impediment...a3aF) + o(3P) at 11.385 eV. Photoexcitation studies indeed find that O(1S) and CO(a) are the dominant photodissociation products. 32,33 If the energy

  6. Dissociative disorders in DSM-5.

    PubMed

    Spiegel, David; Lewis-Fernández, Roberto; Lanius, Ruth; Vermetten, Eric; Simeon, Daphne; Friedman, Matthew

    2013-01-01

    The rationale, research literature, and proposed changes to the dissociative disorders and conversion disorder in the fifth edition of the Diagnostic and Statistical Manual of Mental Disorders (DSM-5) are presented. Dissociative identity disorder will include reference to possession as well as identity fragmentation, to make the disorder more applicable to culturally diverse situations. Dissociative amnesia will include dissociative fugue as a subtype, since fugue is a rare disorder that always involves amnesia but does not always include confused wandering or loss of personality identity. Depersonalization disorder will include derealization as well, since the two often co-occur. A dissociative subtype of posttraumatic stress disorder (PTSD), defined by the presence of depersonalization or derealization in addition to other PTSD symptoms, is being recommended, based upon new epidemiological and neuroimaging evidence linking it to an early life history of adversity and a combination of frontal activation and limbic inhibition. Conversion disorder (functional neurological symptom disorder) will likely remain with the somatic symptom disorders, despite considerable dissociative comorbidity.

  7. PHARMACEUTICAL AND BIOMEDICAL APPLICATIONS OF AFFINITY CHROMATOGRAPHY: RECENT TRENDS AND DEVELOPMENTS

    PubMed Central

    Hage, David S.; Anguizola, Jeanethe A.; Bi, Cong; Li, Rong; Matsuda, Ryan; Papastavros, Efthimia; Pfaunmiller, Erika; Vargas, John; Zheng, Xiwei

    2012-01-01

    Affinity chromatography is a separation technique that has become increasingly important in work with biological samples and pharmaceutical agents. This method is based on the use of a biologically-related agent as a stationary phase to selectively retain analytes or to study biological interactions. This review discusses the basic principles behind affinity chromatography and examines recent developments that have occurred in the use of this method for biomedical and pharmaceutical analysis. Techniques based on traditional affinity supports are discussed, but an emphasis is placed on methods in which affinity columns are used as part of HPLC systems or in combination with other analytical methods. General formats for affinity chromatography that are considered include step elution schemes, weak affinity chromatography, affinity extraction and affinity depletion. Specific separation techniques that are examined include lectin affinity chromatography, boronate affinity chromatography, immunoaffinity chromatography, and immobilized metal ion affinity chromatography. Approaches for the study of biological interactions by affinity chromatography are also presented, such as the measurement of equilibrium constants, rate constants, or competition and displacement effects. In addition, related developments in the use of immobilized enzyme reactors, molecularly imprinted polymers, dye ligands and aptamers are briefly considered. PMID:22305083

  8. Global functioning and disability in dissociative disorders.

    PubMed

    Mueller-Pfeiffer, Christoph; Rufibach, Kaspar; Perron, Noelle; Wyss, Daniela; Kuenzler, Cornelia; Prezewowsky, Cornelia; Pitman, Roger K; Rufer, Michael

    2012-12-30

    Dissociative disorders are frequent comorbid conditions of other mental disorders. Yet, there is controversy about their clinical relevance, and little systematic research has been done on how they influence global functioning. Outpatients and day care patients (N=160) of several psychiatric units in Switzerland were assessed with the Structured Clinical Interview for Diagnostic and Statistical Manual of Mental Disorders (DSM)-IV Axis I Disorders, Structured Clinical Interview for DSM-IV Dissociative Disorders, Global Assessment of Functioning Scale, and World Health Organization Disability Assessment Schedule-II. The association between subjects with a dissociative disorder (N=30) and functional impairment after accounting for non-dissociative axis I disorders was evaluated by linear regression models. We found a proportion of 18.8% dissociative disorders (dissociative amnesia=0%, dissociative fugue=0.6%, depersonalization disorder=4.4%, dissociative identity disorder=7.5%, dissociative disorder-not-otherwise-specified=6.3%) across treatment settings. Adjusted for other axis I disorders, subjects with a comorbid dissociative identity disorder or dissociative disorder-not-otherwise-specified had a median global assessment of functioning score that was 0.86 and 0.88 times, respectively, the score of subjects without a comorbid dissociative disorder. These findings support the hypothesis that complex dissociative disorders, i.e., dissociative identity disorder and dissociative disorder-not-otherwise-specified, contribute to functional impairment above and beyond the impact of co-existing non-dissociative axis I disorders, and that they qualify as "serious mental illness".

  9. Electron attachment and detachment and the electron affinity of cyclo-C4F8

    NASA Astrophysics Data System (ADS)

    Miller, Thomas M.; Friedman, Jeffrey F.; Viggiano, A. A.

    2004-04-01

    New measurements have been made of rate constants for electron attachment to c-C4F8 (octafluorocyclobutane) and thermal electron detachment from the parent anion, c-C4F8-, over the temperature range 298-400 K in 133 Pa of He gas in a flowing-afterglow Langmuir-probe apparatus. From these data the electron affinity for c-C4F8 was determined, EA(c-C4F8)=0.63±0.05 eV. The motivation was to resolve a discrepancy between our earlier EA estimate and a higher value (EA=1.05±0.10 eV) reported from a recent experiment of Hiraoka et al. [J. Chem. Phys. 116, 7574 (2002)]. The electron attachment rate constant is 9.3±3.0×10-9 cm3 s-1 at 298 K. The electron detachment rate constant is negligible at room temperature but climbs to 1945±680 s-1 at 400 K. G3(MP2) calculations were carried out for the neutral (D2d, 1A1) and anion (D4h, 2A2u) and yielded EA(c-C4F8-)=0.595 eV. Bond energies were also calculated for loss of F from c-C4F8 and loss of F or F- from c-C4F8-. From these, dissociative electron attachment is found to be endothermic by at least 1.55 eV.

  10. Electron attachment and detachment and the electron affinity of cyclo-C4F8.

    PubMed

    Miller, Thomas M; Friedman, Jeffrey F; Viggiano, A A

    2004-04-15

    New measurements have been made of rate constants for electron attachment to c-C(4)F(8) (octafluorocyclobutane) and thermal electron detachment from the parent anion, c-C(4)F(8) (-), over the temperature range 298-400 K in 133 Pa of He gas in a flowing-afterglow Langmuir-probe apparatus. From these data the electron affinity for c-C(4)F(8) was determined, EA(c-C(4)F(8))=0.63+/-0.05 eV. The motivation was to resolve a discrepancy between our earlier EA estimate and a higher value (EA=1.05+/-0.10 eV) reported from a recent experiment of Hiraoka et al. [J. Chem. Phys. 116, 7574 (2002)]. The electron attachment rate constant is 9.3+/-3.0x10(-9) cm(3) s(-1) at 298 K. The electron detachment rate constant is negligible at room temperature but climbs to 1945+/-680 s(-1) at 400 K. G3(MP2) calculations were carried out for the neutral (D(2d), (1)A(1)) and anion (D(4h), (2)A(2u)) and yielded EA(c-C(4)F(8) (-))=0.595 eV. Bond energies were also calculated for loss of F from c-C(4)F(8) and loss of F or F(-) from c-C(4)F(8) (-). From these, dissociative electron attachment is found to be endothermic by at least 1.55 eV.

  11. Kernel Affine Projection Algorithms

    NASA Astrophysics Data System (ADS)

    Liu, Weifeng; Príncipe, José C.

    2008-12-01

    The combination of the famed kernel trick and affine projection algorithms (APAs) yields powerful nonlinear extensions, named collectively here, KAPA. This paper is a follow-up study of the recently introduced kernel least-mean-square algorithm (KLMS). KAPA inherits the simplicity and online nature of KLMS while reducing its gradient noise, boosting performance. More interestingly, it provides a unifying model for several neural network techniques, including kernel least-mean-square algorithms, kernel adaline, sliding-window kernel recursive-least squares (KRLS), and regularization networks. Therefore, many insights can be gained into the basic relations among them and the tradeoff between computation complexity and performance. Several simulations illustrate its wide applicability.

  12. Does dissociation further our understanding of PTSD?

    PubMed

    Bryant, Richard A

    2007-01-01

    Peritraumatic dissociation, and other dissociative reactions, refer to alterations in awareness in the context of a traumatic experience. This review provides an overview of the current conceptualization of dissociation, critiques methodological approaches to studying dissociation, and reviews the evidence for the purported relationship between dissociative reactions and posttraumatic stress disorder. The evidence challenges the notion that a linear relationship exists between dissociation and psychiatric morbidity. Future research should abandon the global construct of dissociation, and study the specific responses that involve altered awareness under experimental conditions.

  13. Electrochemical and theoretical analysis of the reactivity of shikonin derivatives: dissociative electron transfer in esterified compounds.

    PubMed

    Armendáriz-Vidales, Georgina; Frontana, Carlos

    2014-09-07

    An electrochemical and theoretical analysis of a series of shikonin derivatives in aprotic media is presented. Results showed that the first electrochemical reduction signal is a reversible monoelectronic transfer, generating a stable semiquinone intermediate; the corresponding E(I)⁰ values were correlated with calculated values of electroaccepting power (ω(+)) and adiabatic electron affinities (A(Ad)), obtained with BH and HLYP/6-311++G(2d,2p) and considering the solvent effect, revealing the influence of intramolecular hydrogen bonding and the substituting group at position C-2 in the experimental reduction potential. For the second reduction step, esterified compounds isobutyryl and isovalerylshikonin presented a coupled chemical reaction following dianion formation. Analysis of the variation of the dimensionless cathodic peak potential values (ξ(p)) as a function of the scan rate (v) functions and complementary experiments in benzonitrile suggested that this process follows a dissociative electron transfer, in which the rate of heterogeneous electron transfer is slow (~0.2 cm s(-1)), and the rate constant of the chemical process is at least 10(5) larger.

  14. Involuntariness in hypnotic responding and dissociative symptoms.

    PubMed

    Dell, Paul F

    2010-01-01

    Clark Hull's (1933) research on dissociation was based on a 'straw man' formulation of dissociation; he claimed that dissociation requires noninterference. Hull completely ignored the then-current paradigm of dissociation--dissociation as automatism--and claimed that he had refuted the validity of the phenomenon of dissociation. Hull's view of dissociation held sway in the hypnosis field for 60 years. This essay seeks to retrieve the Janetian paradigm of dissociation as automatism. Automatisms are unexpected, uninitiated, involuntary behaviors that just 'happen.' The author argues that human sensitivity to the experience of involuntariness (a) is quite important, (b) was selected by evolution, and (c) is central to both hypnotic responses and dissociative symptoms. This editorial urges the hypnosis field and the dissociation field to jointly undertake a renewed investigation of the experience of involuntariness and to follow recent neuroimaging studies which indicate that the parietal cortex underlies the experience of involuntariness.

  15. Kinetic studies of drug-protein interactions by using peak profiling and high-performance affinity chromatography: examination of multi-site interactions of drugs with human serum albumin columns.

    PubMed

    Tong, Zenghan; Schiel, John E; Papastavros, Efthimia; Ohnmacht, Corey M; Smith, Quentin R; Hage, David S

    2011-04-15

    Carbamazepine and imipramine are drugs that have significant binding to human serum albumin (HSA), the most abundant serum protein in blood and a common transport protein for many drugs in the body. Information on the kinetics of these drug interactions with HSA would be valuable in understanding the pharmacokinetic behavior of these drugs and could provide data that might lead to the creation of improved assays for these analytes in biological samples. In this report, an approach based on peak profiling was used with high-performance affinity chromatography to measure the dissociation rate constants for carbamazepine and imipramine with HSA. This approach compared the elution profiles for each drug and a non-retained species on an HSA column and control column over a board range of flow rates. Various approaches for the corrections of non-specific binding between these drugs and the support were considered and compared in this process. Dissociation rate constants of 1.7 (±0.2) s(-1) and 0.67 (±0.04) s(-1) at pH 7.4 and 37°C were estimated by this approach for HSA in its interactions with carbamazepine and imipramine, respectively. These results gave good agreement with rate constants that have determined by other methods or for similar solute interactions with HSA. The approach described in this report for kinetic studies is not limited to these particular drugs or HSA but can also be extended to other drugs and proteins.

  16. Adjoint affine fusion and tadpoles

    NASA Astrophysics Data System (ADS)

    Urichuk, Andrew; Walton, Mark A.

    2016-06-01

    We study affine fusion with the adjoint representation. For simple Lie algebras, elementary and universal formulas determine the decomposition of a tensor product of an integrable highest-weight representation with the adjoint representation. Using the (refined) affine depth rule, we prove that equally striking results apply to adjoint affine fusion. For diagonal fusion, a coefficient equals the number of nonzero Dynkin labels of the relevant affine highest weight, minus 1. A nice lattice-polytope interpretation follows and allows the straightforward calculation of the genus-1 1-point adjoint Verlinde dimension, the adjoint affine fusion tadpole. Explicit formulas, (piecewise) polynomial in the level, are written for the adjoint tadpoles of all classical Lie algebras. We show that off-diagonal adjoint affine fusion is obtained from the corresponding tensor product by simply dropping non-dominant representations.

  17. Vibrational nonequilibrium effects on diatomic dissociation rates

    NASA Technical Reports Server (NTRS)

    Hansen, C. F.

    1993-01-01

    The collision-induced dissociation rate of diatomic molecules from a ladder of rotational and anharmonic vibrational states is developed, and the correction for vibrational nonequilibrium is considered. The result is similar to an analytic correction derived by Hammerling et al. (1959) for harmonic oscillators. An empirical correction algorithm suggested by Park (1987, 1990) gives similar results when vibrational temperature is comparable to kinetic temperature but underestimates the dissociation rate when vibrational temperature is small compared with the kinetic temperature. This algorithm uses an effective temperature in the experimentally determined Arrhenius expression for the rate coefficient, which is a weighted average of the vibrational and kinetic temperature, whereas theory indicates that kinetic temperature should appear only in the exponential term of the Arrhenius expression. Nevertheless, an effective temperature can always be found that will numerically duplicate the proper rate coefficient at any given condition, but a constant weighting factor cannot be expected to provide this. However, the algorithm can he adjusted to give reasonable results over a range of conditions if the geometric weighting factor is taken to be a simple linear function of the ratio of vibrational to kinetic temperature in the gas.

  18. Dissociative disorders among alcohol-dependent inpatients.

    PubMed

    Evren, Cuneyt; Sar, Vedat; Karadag, Figen; Tamar Gurol, Defne; Karagoz, Mustafa

    2007-08-30

    The aim of this study was to determine the prevalence of dissociative disorders among inpatients with alcohol dependency. The Dissociative Experiences Scale was used to screen 111 alcohol-dependent patients consecutively admitted to the inpatient unit of a dependency treatment center. Subgroups of 29 patients who scored 30.0 or above and 25 patients who scored below 10.0 were then evaluated with the Dissociative Disorders Interview Schedule and the Structured Interview for DSM-IV Dissociative Disorders. The interviewers were blind to the Dissociative Experiences Scale scores. Of the 54 patients evaluated, 10 (9.0% of the original 111) patients had a dissociative disorder. A considerable number of the remaining patients reported a high level of dissociative experiences. Among the dissociative disorder group, nine patients had dissociative disorder not otherwise specified and one patient had depersonalization disorder. Female gender, younger age, history of suicide attempt, childhood emotional and sexual abuse, and neglect were more frequent in the dissociative disorder group than among non-dissociative patients. The dissociative disorder group also had somatization disorder, borderline personality disorder, and lifetime major depression more frequently. For 9 of the 10 dissociative patients, dissociative symptoms started before the onset of alcohol use. Although the probability of having a comorbid dissociative disorder was not higher among alcohol-dependent inpatients than among the general psychiatric inpatients, the dissociative subgroup had distinct features. Many patients without a dissociative disorder diagnosis (predominantly men) provided hints of subtle dissociative psychopathology. Implications of comorbid dissociative disorders and dissociative experiences on prevention and treatment of alcohol dependency and the importance of gender-specific characteristics in this relationship require further study.

  19. Cosmological constant, fine structure constant and beyond

    NASA Astrophysics Data System (ADS)

    Wei, Hao; Zou, Xiao-Bo; Li, Hong-Yu; Xue, Dong-Ze

    2017-01-01

    In the present work, we consider the cosmological constant model Λ ∝ α ^{-6}, which is well motivated from three independent approaches. As is well known, the hint of varying fine structure constant α was found in 1998. If Λ ∝ α ^{-6} is right, it means that the cosmological constant Λ should also be varying. Here, we try to develop a suitable framework to model this varying cosmological constant Λ ∝ α ^{-6}, in which we view it from an interacting vacuum energy perspective. Then we consider the observational constraints on these models by using the 293 Δ α /α data from the absorption systems in the spectra of distant quasars. We find that the model parameters can be tightly constrained to the very narrow ranges of O(10^{-5}) typically. On the other hand, we can also view the varying cosmological constant model Λ ∝ α ^{-6} from another perspective, namely it can be equivalent to a model containing "dark energy" and "warm dark matter", but there is no interaction between them. We find that this is also fully consistent with the observational constraints on warm dark matter.

  20. Visuomotor Dissociation in Cerebral Scaling of Size.

    PubMed

    Potgieser, Adriaan R E; de Jong, Bauke M

    2016-01-01

    Estimating size and distance is crucial in effective visuomotor control. The concept of an internal coordinate system implies that visual and motor size parameters are scaled onto a common template. To dissociate perceptual and motor components in such scaling, we performed an fMRI experiment in which 16 right-handed subjects copied geometric figures while the result of drawing remained out of sight. Either the size of the example figure varied while maintaining a constant size of drawing (visual incongruity) or the size of the examples remained constant while subjects were instructed to make changes in size (motor incongruity). These incongruent were compared to congruent conditions. Statistical Parametric Mapping (SPM8) revealed brain activations related to size incongruity in the dorsolateral prefrontal and inferior parietal cortex, pre-SMA / anterior cingulate and anterior insula, dominant in the right hemisphere. This pattern represented simultaneous use of a 'resized' virtual template and actual picture information requiring spatial working memory, early-stage attention shifting and inhibitory control. Activations were strongest in motor incongruity while right pre-dorsal premotor activation specifically occurred in this condition. Visual incongruity additionally relied on a ventral visual pathway. Left ventral premotor activation occurred in all variably sized drawing while constant visuomotor size, compared to congruent size variation, uniquely activated the lateral occipital cortex additional to superior parietal regions. These results highlight size as a fundamental parameter in both general hand movement and movement guided by objects perceived in the context of surrounding 3D space.

  1. Protein Complex Affinity Capture from Cryomilled Mammalian Cells.

    PubMed

    LaCava, John; Jiang, Hua; Rout, Michael P

    2016-12-09

    Affinity capture is an effective technique for isolating endogenous protein complexes for further study. When used in conjunction with an antibody, this technique is also frequently referred to as immunoprecipitation. Affinity capture can be applied in a bench-scale and in a high-throughput context. When coupled with protein mass spectrometry, affinity capture has proven to be a workhorse of interactome analysis. Although there are potentially many ways to execute the numerous steps involved, the following protocols implement our favored methods. Two features are distinctive: the use of cryomilled cell powder to produce cell extracts, and antibody-coupled paramagnetic beads as the affinity medium. In many cases, we have obtained superior results to those obtained with more conventional affinity capture practices. Cryomilling avoids numerous problems associated with other forms of cell breakage. It provides efficient breakage of the material, while avoiding denaturation issues associated with heating or foaming. It retains the native protein concentration up to the point of extraction, mitigating macromolecular dissociation. It reduces the time extracted proteins spend in solution, limiting deleterious enzymatic activities, and it may reduce the non-specific adsorption of proteins by the affinity medium. Micron-scale magnetic affinity media have become more commonplace over the last several years, increasingly replacing the traditional agarose- and Sepharose-based media. Primary benefits of magnetic media include typically lower non-specific protein adsorption; no size exclusion limit because protein complex binding occurs on the bead surface rather than within pores; and ease of manipulation and handling using magnets.

  2. Protein Complex Affinity Capture from Cryomilled Mammalian Cells

    PubMed Central

    LaCava, John; Jiang, Hua; Rout, Michael P.

    2016-01-01

    Affinity capture is an effective technique for isolating endogenous protein complexes for further study. When used in conjunction with an antibody, this technique is also frequently referred to as immunoprecipitation. Affinity capture can be applied in a bench-scale and in a high-throughput context. When coupled with protein mass spectrometry, affinity capture has proven to be a workhorse of interactome analysis. Although there are potentially many ways to execute the numerous steps involved, the following protocols implement our favored methods. Two features are distinctive: the use of cryomilled cell powder to produce cell extracts, and antibody-coupled paramagnetic beads as the affinity medium. In many cases, we have obtained superior results to those obtained with more conventional affinity capture practices. Cryomilling avoids numerous problems associated with other forms of cell breakage. It provides efficient breakage of the material, while avoiding denaturation issues associated with heating or foaming. It retains the native protein concentration up to the point of extraction, mitigating macromolecular dissociation. It reduces the time extracted proteins spend in solution, limiting deleterious enzymatic activities, and it may reduce the non-specific adsorption of proteins by the affinity medium. Micron-scale magnetic affinity media have become more commonplace over the last several years, increasingly replacing the traditional agarose- and Sepharose-based media. Primary benefits of magnetic media include typically lower non-specific protein adsorption; no size exclusion limit because protein complex binding occurs on the bead surface rather than within pores; and ease of manipulation and handling using magnets. PMID:28060343

  3. Dissociative electron attachment to C2F5 radicals.

    PubMed

    Haughey, Sean A; Field, Thomas A; Langer, Judith; Shuman, Nicholas S; Miller, Thomas M; Friedman, Jeffrey F; Viggiano, A A

    2012-08-07

    Dissociative electron attachment to the reactive C(2)F(5) molecular radical has been investigated with two complimentary experimental methods; a single collision beam experiment and a new flowing afterglow Langmuir probe technique. The beam results show that F(-) is formed close to zero electron energy in dissociative electron attachment to C(2)F(5). The afterglow measurements also show that F(-) is formed in collisions between electrons and C(2)F(5) molecules with rate constants of 3.7 × 10(-9) cm(3) s(-1) to 4.7 × 10(-9) cm(3) s(-1) at temperatures of 300-600 K. The rate constant increases slowly with increasing temperature, but the rise observed is smaller than the experimental uncertainty of 35%.

  4. Dissociative electron attachment to C2F5 radicals

    NASA Astrophysics Data System (ADS)

    Haughey, Sean A.; Field, Thomas A.; Langer, Judith; Shuman, Nicholas S.; Miller, Thomas M.; Friedman, Jeffrey F.; Viggiano, A. A.

    2012-08-01

    Dissociative electron attachment to the reactive C2F5 molecular radical has been investigated with two complimentary experimental methods; a single collision beam experiment and a new flowing afterglow Langmuir probe technique. The beam results show that F- is formed close to zero electron energy in dissociative electron attachment to C2F5. The afterglow measurements also show that F- is formed in collisions between electrons and C2F5 molecules with rate constants of 3.7 × 10-9 cm3 s-1 to 4.7 × 10-9 cm3 s-1 at temperatures of 300-600 K. The rate constant increases slowly with increasing temperature, but the rise observed is smaller than the experimental uncertainty of 35%.

  5. Acetylated histone H3 increases nucleosome dissociation

    NASA Astrophysics Data System (ADS)

    Simon, Marek; Manohar, Mridula; Ottesen, Jennifer; Poirier, Michael

    2009-03-01

    Chromatin's basic unit structure is the nucleosome, i.e. genomic DNA wrapped around a particular class of proteins -- histones -- which due to their physical hindrance, block vital biological processes, such as DNA repair, DNA replication, and RNA transcription. Histone post-translational modifications, which are known to exist in vivo, are hypothesized to regulate these biological processes by directly altering DNA-histone interactions and thus nucleosome structure and stability. Using magnetic tweezers technique we studied the acetylation of histone H3 in the dyad region, i.e. at K115 and K122, on reconstituted arrays of nucleosomes under constant external force. Based on the measured increase in the probability of dissociation of modified nucleosomes, we infer that this double modification could facilitate histone chaperone mediated nucleosome disassembly in vivo.

  6. Spectroscopic constants and potential energy curves of SnF

    NASA Astrophysics Data System (ADS)

    Dai, Dingguo; Balasubramanian, K.

    1994-07-01

    Spectroscopic constants and potential energy curves of 20 electronic states of SnF arising from valence dissociation limits are computed using the complete active space MCSCF (CASSCF) followed by first- and second-order configuration interaction (FOCI, SOCI) methods, which included up to one million configurations. Our computed spectroscopic constants of five electronic states fit well with the experimentally observed X, A, a, and C states. The dipole moments and other yet to be observed properties of several electronic states are reported.

  7. In vitro selection, characterization, and biosensing application of high-affinity cylindrospermopsin-targeting aptamers.

    PubMed

    Elshafey, Reda; Siaj, Mohamed; Zourob, Mohammed

    2014-09-16

    Contamination of freshwater with cyanotoxin cylindrospermopsin (CYN) represents a significant global concern for public health. The sensitive detection of CYN is necessary to effectively manage and control the treatment of water resources. Here we report a novel, highly sensitive label-free aptasensor for CYN analysis, using aptamers as specific receptors. We have selected the DNA aptamers from a diverse random library using the in vitro screening SELEX approach. The aptamers exhibited high affinity for CYN with Kd of nanomolar range. One aptamer exhibited conformational change upon CYN recognition (CD analysis) and was used to fabricate the label-free impedimetric aptasensor for CYN. A self-assembled monolayer from a disulfide-derivatized aptamer was formed on a gold electrode to fabricate the aptasensor. Upon CYN capturing to the aptasensor surface, a marked drop in the electron transfer resistance was obtained, which was used as the principle of detection of CYN. This resulted from the aptamer's conformational change induced by CYN recognition. The present aptasensor could detect CYN with the limit of detection as low as 100 pM and a wide linear range of 0.1 to 80 nM. When mounted on the gold surface, the aptamer exhibited a lower dissociation constant for CYN than that observed in the fluorescence assay, implying that the anchoring of the aptamer on the Au surface improved its affinity to CYN. Moreover, the aptasensor showed high specificity toward other coexistent cyanobacterial toxins of microcystin-LR and Anatoxin-a. Further biosensor designs will be generated using those aptamers for simple and sensitive CYN monitoring.

  8. Reinforcement of frontal affinity chromatography for effective analysis of lectin-oligosaccharide interactions.

    PubMed

    Hirabayashi, J; Arata, Y; Kasai, K

    2000-08-25

    Frontal affinity chromatography is a method for quantitative analysis of biomolecular interactions. We reinforced it by incorporating various merits of a contemporary liquid chromatography system. As a model study, the interaction between an immobilized Caenorhabditis elegans galectin (LEC-6) and fluorescently labeled oligosaccharides (pyridylaminated sugars) was analyzed. LEC-6 was coupled to N-hydroxysuccinimide-activated Sepharose 4 Fast Flow (100 microm diameter), and packed into a miniature column (e.g., 10 x 4.0 mm, 0.126 ml). Twelve pyridylaminated oligosaccharides were applied to the column through a 2-ml sample loop, and their elution patterns were monitored by fluorescence. The volume of the elution front (V) determined graphically for each sample was compared with that obtained in the presence of an excess amount of hapten saccharide, lactose (V0); and the dissociation constant, Kd, was calculated according to the literature [K. Kasai, Y. Oda, M. Nishikawa, S. Ishii, J. Chromatogr. 376 (1986) 33]. This system also proved to be useful for an inverse confirmation; that is, application of galectins to an immobilized glycan column (in the present case, asialofetuin was immobilized on Sepharose 4 Fast Flow), and the elution profiles were monitored by fluorescence based on tryptophan. The relative affinity of various galectins for asialofetuin could be easily compared in terms of the extent of retardation. The newly constructed system proved to be extremely versatile. It enabled rapid (analysis time 12 min/cycle) and sensitive (20 nM for pyridylaminated derivatives, and 1 microg/ml for protein) analyses of lectin-carbohydrate interactions. It should become a powerful tool for elucidation of biomolecular interactions, in particular for functional analysis of a large number of proteins that should be the essential issues of post-genome projects.

  9. Spectroscopic constants and potential energy curves of PF

    NASA Astrophysics Data System (ADS)

    Latifzadeh, Lida; Balasubramanian, K.

    1995-09-01

    Spectroscopic constants of low-lying bound electronic states and potential energy curves of 19 electronic states of PF arising from the valence dissociation limits are computed using the complete active space self-consistent field (CASSCF) followed by first- and second-order configuration interaction (FOCI, SOCI) methods, which included up to 0.9 million configurations.

  10. Dissociative symptomatology in cancer patients

    PubMed Central

    Civilotti, Cristina; Castelli, Lorys; Binaschi, Luca; Cussino, Martina; Tesio, Valentina; Di Fini, Giulia; Veglia, Fabio; Torta, Riccardo

    2015-01-01

    Introduction: The utilization of the post-traumatic stress disorder (PTSD) diagnostic spectrum is currently being debated to categorize psychological adjustment in cancer patients. The aims of this study were to: (1) evaluate the presence of cancer-related traumatic dissociative symptomatology in a sample of cancer patients; (2) examine the correlation of cancer-related dissociation and sociodemographic and medical variables, anxiety, depression, and post-traumatic stress symptomatology; (3) investigate the predictors of cancer-related dissociation. Methods: Ninety-two mixed cancer patients (mean age: 58.94, ds = 10.13) recruited from two hospitals in northern Italy were administered a questionnaire on sociodemographic and medical characteristics, the Karnofsky Scale to measure the level of patient activity and medical care requirements, the Hospital Anxiety and Depression Scale (HADS) to evaluate the presence of anxiety and depression, the Impact of Event Scale Revised (IES-R) to assess the severity of intrusion, avoidance, and hypervigilance, and the Peritraumatic Dissociative Experiences Questionnaire (PDEQ) to quantify the traumatic dissociative symptomatology. Results: 31.5% of participants report a PDEQ score above the cutoff. The results indicated that dissociative symptomatology was positively correlated with HADS scores (HADS-Anxiety: r = 0.476, p < 0.001; HADS-Depression: r = 0.364, p < 0.001) and with IES-R scores (IES-R-Intrusion: r = 0.698, p < 0.001; IES-R-Avoidance: r = 0.619, p < 0.001; IES-R- Hypervigilance: r = 0.681, p < 0.001). A stepwise regression analysis was performed in order to find the predictors of cancer-related traumatic dissociative symptomatology. The results converged on a three predictor model revealing that IES-R-Intrusion, IES-R-Avoidance, and IES-R-Hyperarousal accounted for 53.9% of the explained variance. Conclusion: These findings allow us to hypothesize a specific psychological reaction which may be ascribed to the traumatic

  11. Identification and characterization of a novel high affinity metal-binding site in the hammerhead ribozyme.

    PubMed Central

    Hansen, M R; Simorre, J P; Hanson, P; Mokler, V; Bellon, L; Beigelman, L; Pardi, A

    1999-01-01

    A novel metal-binding site has been identified in the hammerhead ribozyme by 31P NMR. The metal-binding site is associated with the A13 phosphate in the catalytic core of the hammerhead ribozyme and is distinct from any previously identified metal-binding sites. 31P NMR spectroscopy was used to measure the metal-binding affinity for this site and leads to an apparent dissociation constant of 250-570 microM at 25 degrees C for binding of a single Mg2+ ion. The NMR data also show evidence of a structural change at this site upon metal binding and these results are compared with previous data on metal-induced structural changes in the core of the hammerhead ribozyme. These NMR data were combined with the X-ray structure of the hammerhead ribozyme (Pley HW, Flaherty KM, McKay DB. 1994. Nature 372:68-74) to model RNA ligands involved in binding the metal at this A13 site. In this model, the A13 metal-binding site is structurally similar to the previously identified A(g) metal-binding site and illustrates the symmetrical nature of the tandem G x A base pairs in domain 2 of the hammerhead ribozyme. These results demonstrate that 31P NMR represents an important method for both identification and characterization of metal-binding sites in nucleic acids. PMID:10445883

  12. Kinetic exclusion assay of monoclonal antibody affinity to the membrane protein Roundabout 1 displayed on baculovirus.

    PubMed

    Kusano-Arai, Osamu; Fukuda, Rie; Kamiya, Wakana; Iwanari, Hiroko; Hamakubo, Takao

    2016-07-01

    The reliable assessment of monoclonal antibody (mAb) affinity against membrane proteins in vivo is a major issue in the development of cancer therapeutics. We describe here a simple and highly sensitive method for the evaluation of mAbs against membrane proteins by means of a kinetic exclusion assay (KinExA) in combination with our previously developed membrane protein display system using budded baculovirus (BV). In our BV display system, the membrane proteins are displayed on the viral surface in their native form. The BVs on which the liver cancer antigen Roundabout 1 (Robo1) was displayed were adsorbed onto magnetic beads without fixative (BV beads). The dissociation constant (Kd, ∼10(-11) M) that was measured on the Robo1 expressed BV beads correlated well with the value from a whole cell assay (the coefficient of determination, R(2) = 0.998) but not with the value for the soluble extracellular domains of Robo1 (R(2) = 0.834). These results suggest that the BV-KinExA method described here provides a suitably accurate Kd evaluation of mAbs against proteins on the cell surface.

  13. Seasonal changes in cortisol sensitivity and glucocorticoid receptor affinity and number in leukocytes of coho salmon

    USGS Publications Warehouse

    Maule, Alec G.; Schreck, Carl B.; Sharpe, Cameron

    1993-01-01

    To determine if there were organ-specific changes in immune responses or immune-endocrine interaction, we monitored in vitro immune response, cortisol sensitivity and number and affinity of glucocorticoid receptors (GR) in leukocytes from freshwater-adapted juvenile coho salmon (Oncorhynchus kisutch) during the physiological changes that prepare them to enter the marine environment. During this period, absolute immune response declined, but splenic leukocytes generated more antibody-producing cells than did cells from anterior kidney. Splenic leukocytes were initially more sensitive to the suppressive effects of cortisol and had fewer GR than leukocytes from the anterior kidney. Leukocytes from the anterior kidney were initially insensitive to cortisol but developed sensitivity at about the same time as the dissociation constant and number of GR increased. In vitro incubation of anterior kidney leukocytes in cortisol altered GR variables when experiments were conducted during March through September but not during November through February. In some years, changes in GR or immune responses were correlated with plasma cortisol titers, but in other years there was no correlation. Thus, the exact relation between cortisol, GR and immune response in anadromous salmonids is unclear and other factors are involved.

  14. Rapid screening of textile dyes employed as affinity ligands to purify enzymes from yeast.

    PubMed

    Raya-Tonetti, G; Perotti, N

    1999-04-01

    A rapid method for screening potential dye ligands for use in affinity chromatography is described. Textile dyes were non-covalently coupled to a cross-linked polysaccharide Sepharose(R) matrix. Yeast alcohol dehydrogenase (ADH) was used as the model protein for evaluating the screening system. A homogenate from baker's yeast was used as the crude source of enzyme. Batchwise adsorption and elution were used to evaluate the individual dyes. The influence of pH and ionic strength in the binding and elution steps was evaluated. Batch isotherms were used to evaluate parameter characteristics. Experimental data obtained were fitted to Langmuir isotherms to determine the maximum binding capacity and the dissociation constant for each dye evaluated in this study. A dynamic binding capacity of 107.6 units of ADH/g of resin was determined for Procion Turquoise MXG dye by frontal analysis. Specific elution with NAD+ and non-specific elution with 50 mM Tris/HCl buffer, pH 8.5, were tested when Procion Turquoise MXG was used, giving purification factors of 53.5 and 4.4 respectively. This screening technique is inexpensive and can be performed in a few hours. It was possible to predict the performance of different reactive dyes in this way, and the influence of pH and salt on the binding behaviour was demonstrated.

  15. Quantitative Affinity Determination by Fluorescence Anisotropy Measurements of Individual Nanoliter Droplets

    PubMed Central

    2017-01-01

    Fluorescence anisotropy measurements of reagents compartmentalized into individual nanoliter droplets are shown to yield high-resolution binding curves from which precise dissociation constants (Kd) for protein–peptide interactions can be inferred. With the current platform, four titrations can be obtained per minute (based on ∼100 data points each), with stoichiometries spanning more than 2 orders of magnitude and requiring only tens of microliters of reagents. In addition to affinity measurements with purified components, Kd values for unpurified proteins in crude cell lysates can be obtained without prior knowledge of the concentration of the expressed protein, so that protein purification can be avoided. Finally, we show how a competition assay can be set up to perform focused library screens, so that compound labeling is not required anymore. These data demonstrate the utility of droplet compartments for the quantitative characterization of biomolecular interactions and establish fluorescence anisotropy imaging as a quantitative technique in a miniaturized droplet format, which is shown to be as reliable as its macroscopic test tube equivalent. PMID:28192993

  16. CHARACTERIZATION OF DRUG INTERACTIONS WITH SERUM PROTEINS BY USING HIGH-PERFORMANCE AFFINITY CHROMATOGRAPHY

    PubMed Central

    Hage, David S.; Anguizola, Jeanethe; Barnaby, Omar; Jackson, Abby; Yoo, Michelle J.; Papastavros, Efthimia; Pfaunmiller, Erika; Sobansky, Matt; Tong, Zenghan

    2011-01-01

    The binding of drugs with serum proteins can affect the activity, distribution, rate of excretion, and toxicity of pharmaceutical agents in the body. One tool that can be used to quickly analyze and characterize these interactions is high-performance affinity chromatography (HPAC). This review shows how HPAC can be used to study drug-protein binding and describes the various applications of this approach when examining drug interactions with serum proteins. Methods for determining binding constants, characterizing binding sites, examining drug-drug interactions, and studying drug-protein dissociation rates will be discussed. Applications that illustrate the use of HPAC with serum binding agents such as human serum albumin, α1-acid glycoprotein, and lipoproteins will be presented. Recent developments will also be examined, such as new methods for immobilizing serum proteins in HPAC columns, the utilization of HPAC as a tool in personalized medicine, and HPAC methods for the high-throughput screening and characterization of drug-protein binding. PMID:21395530

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

    SciTech Connect

    Raza, H.; Chung, W.L.; Mukhtar, H. )

    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.

  18. Determining force dependence of two-dimensional receptor-ligand binding affinity by centrifugation.

    PubMed Central

    Piper, J W; Swerlick, R A; Zhu, C

    1998-01-01

    Analyses of receptor-ligand interactions are important to the understanding of cellular adhesion. Traditional methods of measuring the three-dimensional (3D) dissociation constant (Kd) require at least one of the molecular species in solution and hence cannot be directly applied to the case of cell adhesion. We describe a novel method of measuring 2D binding characteristics of receptors and ligands that are attached to surfaces and whose bonds are subjected to forces. The method utilizes a common centrifugation assay to quantify adhesion. A model for the experiment has been formulated, solved exactly, and tested carefully. The model is stochastically based and couples the bond force to the binding affinity. The method was applied to examine tumor cell adherence to recombinant E-selectin. Satisfactory agreement was found between predictions and data. The estimated zero-force 2D Kd for E-selectin/carbohydrate ligand binding was approximately 5 x 10(3) microm(-2), and the bond interaction range was subangstrom. Our results also suggest that the number of bonds mediating adhesion was small (<5). PMID:9449350

  19. Local release from affinity-based polymers increases urethral concentration of the stem cell chemokine CCL7 in rats.

    PubMed

    Rivera-Delgado, Edgardo; Sadeghi, Zhina; Wang, Nick X; Kenyon, Jonathan; Satyanarayan, Sapna; Kavran, Michael; Flask, Chris; Hijaz, Adonis Z; von Recum, Horst A

    2016-04-21

    The protein chemokine (C-C motif) ligand 7 (CCL7) is significantly over-expressed in urethral and vaginal tissues immediately following vaginal distention in a rat model of stress urinary incontinence. Further evidence, in this scenario and other clinical scenarios, indicates CCL7 stimulates stem cell homing for regenerative repair. This CCL7 gradient is likely absent or compromised in the natural repair process of women who continue to suffer from SUI into advanced age. We evaluated the feasibility of locally providing this missing CCL7 gradient by means of an affinity-based implantable polymer. To engineer these polymers we screened the affinity of different proteoglycans, to use them as CCL7-binding hosts. We found heparin to be the strongest binding host for CCL7 with a 0.323 nM dissociation constant. Our experimental approach indicates conjugation of heparin to a polymer backbone (using either bovine serum albumin or poly (ethylene glycol) as the base polymer) can be used as a delivery system capable of providing sustained concentrations of CCL7 in a therapeutically useful range up to a month in vitro. With this approach we are able to detect, after polymer implantation, significant increase in CCL7 in the urethral tissue directly surrounding the polymer implants with only trace amounts of human CCL7 present in the blood of the animals. Whole animal serial sectioning shows evidence of retention of locally injected human mesenchymal stem cells (hMSCs) only in animals with sustained CCL7 delivery, 2 weeks after affinity-polymers were implanted.

  20. Dissociative Electron Attachment to chloroacetylene

    NASA Astrophysics Data System (ADS)

    Ngassam, V.; Orel, A. E.

    2007-06-01

    The production of two fragments with σ symmetry from electron-impact dissociation of C2H2, which has only a low lying &*circ; resonance at equilibrium geometry, has been explained by the existence of interactions with &*circ; resonances at bent geometries. We are investigating the presence of such multidimensional effects in the dissociative attachment of chloroacetylene (C2HCl). We have performed electron scattering calculations using the Complex Kohn variational method to determine the resonance energies and widths of the chloroacetylene resonances as a function of both the Cl--C2H bond distance as well as the variation with C-C stretch and bend. We will discuss our results and our prediction of the dissociation dynamics in comparison to the findings for for C2H2. This work was supported by the U.S. DOE Office of Basic Energy Sciences, Division of Chemical Sciences and the National Science Foundation, PHY-05-55401.

  1. A homogeneous assay for relative affinity of binding proteins using a green fluorescent protein tag and membrane disk.

    PubMed

    Aoki, Takashi; Kazama, Hitoshi; Satoh, Marie; Mizuki, Kazuhiro; Watabe, Hiroyuki

    2005-09-01

    When the association between a ligand immobilized on a membrane disk and a fluorescence-labeled analyte was monitored with a fluorescent microplate reader, the time-dependent increase in fluorescence intensity of the reaction mixture was observed. A novel assay system for the specific interaction based on this phenomenon was designated the homogeneous assay for fluorescence concentrated on membrane (HAFCOM). In this study, streptococcal protein G (SpG) and glycogen-binding subunit R5 of protein phosphatase 1 (PPP1R5) tagged by green fluorescent protein (GFP) were used as the fluorescence-labeled analytes, and the affinity change caused by various amino acid substitutions was measured with HAFCOM. From the site-directed mutagenesis of SpG and PPP1R5, it was clarified that (i) the association rate constant of the Lys454Pro/Glu456Gln mutant of SpG to goat immunoglobulin G was almost equivalent to that of the wild-type but its dissociation rate constant was about 2.7 times that of the wild-type and (ii) the amino acid substitutions of Phe180 in PPP1R5 reduced glycogen-binding by 30-50%. Since HAFCOM using the GFP-tagged analyte requires no special chemicals and instruments, this system can easily and economically assay the specific interaction between target protein and ligand.

  2. Affine conformal vectors in space-time

    NASA Astrophysics Data System (ADS)

    Coley, A. A.; Tupper, B. O. J.

    1992-05-01

    All space-times admitting a proper affine conformal vector (ACV) are found. By using a theorem of Hall and da Costa, it is shown that such space-times either (i) admit a covariantly constant vector (timelike, spacelike, or null) and the ACV is the sum of a proper affine vector and a conformal Killing vector or (ii) the space-time is 2+2 decomposable, in which case it is shown that no ACV can exist (unless the space-time decomposes further). Furthermore, it is proved that all space-times admitting an ACV and a null covariantly constant vector (which are necessarily generalized pp-wave space-times) must have Ricci tensor of Segré type {2,(1,1)}. It follows that, among space-times admitting proper ACV, the Einstein static universe is the only perfect fluid space-time, there are no non-null Einstein-Maxwell space-times, and only the pp-wave space-times are representative of null Einstein-Maxwell solutions. Otherwise, the space-times can represent anisotropic fluids and viscous heat-conducting fluids, but only with restricted equations of state in each case.

  3. Molecular Dynamics Simulation of Ligand Dissociation from Liver Fatty Acid Binding Protein

    PubMed Central

    Long, Dong; Mu, Yuguang; Yang, Daiwen

    2009-01-01

    The mechanisms of how ligands enter and leave the binding cavity of fatty acid binding proteins (FABPs) have been a puzzling question over decades. Liver fatty acid binding protein (LFABP) is a unique family member which accommodates two molecules of fatty acids in its cavity and exhibits the capability of interacting with a variety of ligands with different chemical structures and properties. Investigating the ligand dissociation processes of LFABP is thus a quite interesting topic, which however is rather difficult for both experimental approaches and ordinary simulation strategies. In the current study, random expulsion molecular dynamics simulation, which accelerates ligand motions for rapid dissociation, was used to explore the potential egress routes of ligands from LFABP. The results showed that the previously hypothesized “portal region” could be readily used for the dissociation of ligands at both the low affinity site and the high affinity site. Besides, one alternative portal was shown to be highly favorable for ligand egress from the high affinity site and be related to the unique structural feature of LFABP. This result lends strong support to the hypothesis from the previous NMR exchange studies, which in turn indicates an important role for this alternative portal. Another less favored potential portal located near the N-terminal end was also identified. Identification of the dissociation pathways will allow further mechanistic understanding of fatty acid uptake and release by computational and/or experimental techniques. PMID:19564911

  4. Affinity enhancement of an in vivo matured therapeutic antibody using structure-based computational design.

    PubMed

    Clark, Louis A; Boriack-Sjodin, P Ann; Eldredge, John; Fitch, Christopher; Friedman, Bethany; Hanf, Karl J M; Jarpe, Matthew; Liparoto, Stefano F; Li, You; Lugovskoy, Alexey; Miller, Stephan; Rushe, Mia; Sherman, Woody; Simon, Kenneth; Van Vlijmen, Herman

    2006-05-01

    Improving the affinity of a high-affinity protein-protein interaction is a challenging problem that has practical applications in the development of therapeutic biomolecules. We used a combination of structure-based computational methods to optimize the binding affinity of an antibody fragment to the I-domain of the integrin VLA1. Despite the already high affinity of the antibody (Kd approximately 7 nM) and the moderate resolution (2.8 A) of the starting crystal structure, the affinity was increased by an order of magnitude primarily through a decrease in the dissociation rate. We determined the crystal structure of a high-affinity quadruple mutant complex at 2.2 A. The structure shows that the design makes the predicted contacts. Structural evidence and mutagenesis experiments that probe a hydrogen bond network illustrate the importance of satisfying hydrogen bonding requirements while seeking higher-affinity mutations. The large and diverse set of interface mutations allowed refinement of the mutant binding affinity prediction protocol and improvement of the single-mutant success rate. Our results indicate that structure-based computational design can be successfully applied to further improve the binding of high-affinity antibodies.

  5. Electron Affinity Calculations for Thioethers

    NASA Technical Reports Server (NTRS)

    Sulton, Deley L.; Boothe, Michael; Ball, David W.; Morales, Wilfredo

    1997-01-01

    Previous work indicated that polyphenyl thioethers possessed chemical properties, related to their electron affinities, which could allow them to function as vapor phase lubricants (VPL). Indeed, preliminary tribological tests revealed that the thioethers could function as vapor phase lubricants but not over a wide temperature and hertzian pressure range. Increasing the electron affinity of the thioethers may improve their VPL properties over this range. Adding a substituent group to the thioether will alter its electron affinity in many cases. Molecular orbital calculations were undertaken to determine the effect of five different substituent groups on the electron affinity of polyphenyl thioethers. It was found that the NO2, F, and I groups increased the thioethers electron affinity by the greatest amount. Future work will involve the addition of these groups to the thioethers followed by tribological testing to assess their VPL properties.

  6. Diffraction dissociation at the LHC

    SciTech Connect

    Jenkovszky, Laszlo; Orava, Risto; Salii, Andrii

    2013-04-15

    We report on recent calculations of low missing mass single (SD) and double (DD) diffractive dissociation at LHC energies. The calculations are based on a dual-Regge model, dominated by a single Pomeron exchange. The diffractively excited states lie on the nucleon trajectory N*, appended by the isolated Roper resonance. Detailed predictions for the squared momentum transfer and missing mass dependence of the differential and integrated single-and double diffraction dissociation in the kinematical range of present and future LHC measurements are given.

  7. [Dissociative identity disorder or schizophrenia?].

    PubMed

    Tschöke, S; Steinert, T

    2010-01-01

    We present a case of dissociative identity disorder in which Schneiderian first rank symptoms were present besides of various states of consciousness. Thus the diagnosis of schizophrenia had to be considered. Formally, the symptoms met ICD-10 criteria for schizophrenia. However, taking into account the lack of formal thought disorder and of negative symptoms as well as a typical history of severe and prolonged traumatisation, we did not diagnose a co-morbid schizophrenic disorder. There is good evidence for the existence of psychotic symptoms among patients with dissociative disorders. However, in clinical practice this differential diagnosis is rarely considered.

  8. Rate constants of atomic hydrogen formation in H3O+(H2O) n + e → H + (H2O) n gas-phase processes

    NASA Astrophysics Data System (ADS)

    Stepanov, N. F.; Novakovskaya, Yu. V.

    2009-09-01

    Using the Maxwellian electron velocity distribution and the Breit-Wigner approximation of the reaction cross section, the kinetic parameters of the hydrogen atom formation upon the electron capture by positively charged hydronium-water clusters are estimated. Calculations of the cross sections and rate constants are based on the data of quantum chemical studies of H3O+(H2O) n and H3O(H2O) n clusters, particularly on the detailed analysis of the spacing of high-lying states of the radicals and the character of the unpaired electron density distribution, as well as on the general trend in the electron affinity change of the cations depending on the number of water molecules. The lifetimes of the radicals before the dissociation are taken from the classical nonempirical molecular dynamics runs. The results are compared to available experimental data.

  9. Trajectory study of dissociation reactions. The single-ensemble method. II

    NASA Astrophysics Data System (ADS)

    Kutz, H. Douglas; Burns, George

    1981-04-01

    The single uniform ensemble method was previously employed in 3D classical trajectory calculations [H. D. Kutz and G. Burns, J. Chem. Phys. 72, 3652 (1980)]. Presently it is applied to the Br2+Ar system to study nonequilbrium effects in diatom dissociation over a wide temperature range. It was found that, for a given large set of trajectories, observables, such as reaction cross sections or rate constants, are indepedent within four significant figures of the initial distribution function. This indicates a high degree of reliability of the single uniform ensemble method, once the choice of a set of trajectories is made. In order to study dissociation from the low lying energy states, the uniform velocity selection method in trajectory calculations was used. It was found that dissociation from these states contributes but little to the overall dissociation reaction. The latter finding is consistent with the attractive nature of the potential energy surface used, and constitutes an argument against those current theories of diatom dissociation reaction which explains experimental data by postulating a high probability of dissociation from low lying energy states of diatoms. It was found that the contribution from the low lying states to dissociation can be estimated with good accuracy using information theory expressions. Temperature dependence of nonequilibrium effects was investigated between 1 500 and 6 000 °K. In this range the nonequilibrium correction factor varies between 0.2 and 0.5. Angular momentum dependence of such observables as reaction rate constant and reaction cross section was investigated.

  10. On the Khinchin Constant

    NASA Technical Reports Server (NTRS)

    Bailey, David H.; Borwein, Jonathan M.; Crandall, Richard E.; Craw, James M. (Technical Monitor)

    1995-01-01

    We prove known identities for the Khinchin constant and develop new identities for the more general Hoelder mean limits of continued fractions. Any of these constants can be developed as a rapidly converging series involving values of the Riemann zeta function and rational coefficients. Such identities allow for efficient numerical evaluation of the relevant constants. We present free-parameter, optimizable versions of the identities, and report numerical results.

  11. Kinetic Studies of Biological Interactions By Affinity Chromatography

    PubMed Central

    Schiel, John E.; Hage, David S.

    2009-01-01

    The rates at which biological interactions occur can provide important information on the mechanism and behavior of such processes in living systems. This review will discuss how affinity chromatography can be used as a tool to examine the kinetics of biological interactions. This approach, referred to here as biointeraction chromatography, uses a column with an immobilized binding agent to examine the association or dissociation of this agent with other compounds. The use of HPLC-based affinity columns in kinetic studies has received particular attention in recent years. Advantages of using HPLC with affinity chromatography for this purpose include the ability to reuse the same ligand within a column for a large number of experiments, and the good precision and accuracy of this approach. A number of techniques are available for kinetic studies through the use of affinity columns and biointeraction chromatography. These approaches include plate height measurements, peak profiling, peak fitting, split-peak measurements, and peak decay analysis. The general principles for each of these methods are discussed in this review and some recent applications of these techniques are presented. The advantages and potential limitations of each approach are also considered. PMID:19391173

  12. [Gender differences in dissociative disorders].

    PubMed

    Spitzer, C; Freyberger, H J

    2008-01-01

    The relationship between mental illness, on the one hand, and sex and gender, on the other hand, has received interest since the beginning of medicine in antique times. A prototypical example of a seemingly woman-specific disease is hysteria. The term itself, which is derived from the Greek word for womb, denotes a psychosexual dimension comprising the current attitude towards sexuality and the dominating gender relationship. In addition, the colourful history of hysteria indicates that illness is not exclusively determined by biological factors, but also significantly by socio-cultural influences, for example in the treatment of hysterical women. Even nowadays, there is a wide-spread belief that dissociative symptoms and disorders, which have succeeded hysteria in current classification systems, are predominantly seen in women. However, empirical studies in the general population and in different clinical samples using sound instruments have indicated that dissociative symptoms do not differ between the genders. The seemingly dominance of dissociative disorders in women may also depend on the socio-cultural context, because men with dissociative disorders usually do not enter the general health system, but rather the legal system, i.e. they can be found in jail or forensic institutions.

  13. Heterogeneity of the chondroitin sulfate portion of phosphacan/6B4 proteoglycan regulates its binding affinity for pleiotrophin/heparin binding growth-associated molecule.

    PubMed

    Maeda, Nobuaki; He, Jue; Yajima, Yuki; Mikami, Tadahisa; Sugahara, Kazuyuki; Yabe, Tomio

    2003-09-12

    PTP zeta is a receptor-type protein-tyrosine phosphatase that is synthesized as a chondroitin sulfate proteoglycan and uses pleiotrophin as a ligand. The chondroitin sulfate portion of this receptor is essential for high affinity binding to pleiotrophin. Here, we purified phosphacan, which corresponds to the extracellular domain of PTP zeta, from postnatal day 7 (P7) and P12 rat cerebral cortex (PG-P7 and PG-P12, respectively) and from P20 rat whole brain (PG-P20). The chondroitin sulfate of these preparations displayed immunologically and compositionally different structures. In particular, only PG-P20 reacted with the monoclonal antibody MO-225, which recognizes chondroitin sulfate containing the GlcA(2S)beta 1-3GalNAc(6S) disaccharide unit (D unit). Analysis of the chondroitinase digestion products revealed that GlcA beta 1-3GalNAc(4S) disaccharide unit (A unit) was the major component in these preparations and that PG-P20 contained 1.3% D unit, which was not detected in PG-P7 and PG-P12. Interaction analysis using a surface plasmon resonance biosensor indicated that PG-P20 had approximately 5-fold stronger affinity for pleiotrophin (dissociation constant (KD) = 0.14 nM) than PG-P7 and PG-P12, although all these preparations showed similar low affinity binding to pleiotrophin after chondroitinase ABC digestion (KD = 1.4 approximately 1.6 nM). We also found that shark cartilage chondroitin sulfate D containing approximately 20% D unit bound to pleiotrophin with moderate affinity (KD = 2.7 nM), whereas whale cartilage chondroitin sulfate A showed no binding to this growth factor. These results suggest that variation of chondroitin sulfate plays important roles in the regulation of signal transduction in the brain.

  14. 3- and 4-O-sulfoconjugated and methylated dopamine: highly reduced binding affinity to dopamine D2 receptors in rat striatal membranes.

    PubMed

    Werle, E; Lenz, T; Strobel, G; Weicker, H

    1988-07-01

    The binding properties of 3- and 4-O-sulfo-conjugated dopamine (DA-3-O-S, DA-4-O-S) as well as 3-O-methylated dopamine (MT) to rat striatal dopamine D2 receptors were investigated. 3H-spiperone was used as a radioligand in the binding studies. In saturation binding experiments (+)butaclamol, which has been reported to bind to dopaminergic D2 and serotoninergic 5HT2 receptors, was used in conjunction with ketanserin and sulpiride, which preferentially label 5HT2 and D2 receptors, respectively, in order to discriminate between 3H-spiperone binding to D2 and to 5HT2 receptors. Under our particular membrane preparation and assay conditions, 3H-spiperone binds to D2 and 5HT2 receptors with a maximal binding capacity (Bmax) of 340 fmol/mg protein in proportions of about 75%:25% with similar dissociation constants KD (35 pmol/l; 43 pmol/l). This result was verified by the biphasic competition curve of ketanserin, which revealed about 20% high (KD = 24 nmol/l) and 80% low (KD = 420 nmol/l) affinity binding sites corresponding to 5HT2 and D2 receptors, respectively. Therefore, all further competition experiments at a tracer concentration of 50 pmol/l were performed in the presence of 0.1 mumol/l ketanserin to mask the 5HT2 receptors. DA competition curves were best fitted assuming two binding sites, with high (KH = 0.12 mumol/l) and low (KL = 18 mumol/l) affinity, present in a ratio of 3:1. The high affinity binding sites were interconvertible by 100 mumol/l guanyl-5-yl imidodiphosphate [Gpp(NH)p], resulting in a homogenous affinity state of DA receptors (KD = 2.8 mumol/l).2+ off

  15. Study of heat transfer characteristics during dissociation of gas hydrates in porous media

    SciTech Connect

    Kamath, V.A.

    1984-01-01

    An experimental technique was developed to measure the rate of formation and dissociation of hydrates in porous media. In the first phase of the work, hydrates of propane and methane were studied. Propane hydrate cores were formed by contacting liquid propane with compacted porous ice cores at 274 K for 24 to 100 hours, whereas the formation of methane hydrates was achieved by contacting ice cores with gaseous methane at about 7000 kPa and 274 K, for 24 to 200 hours. These hydrate cores were dissociated by circulating warm water over the top of the core, under controlled temperatures and pressures. The major findings of these experiments are as follows: 1) the phenomena of dissociation of hydrates to liquid water and gas is similar to nucleate boiling of liquids; 2) the rate of dissociation of hydrates at constant ..delta..T, is directly proportional to the area of hydrates exposed to the warm fluid or the composition of hydrates in the core; and 3) the rate of heat transfer and dissociation increase with increase in pressure and the rate of circulation of the warm fluid. Unified correlations for heat transfer and dissociation rates were successfully obtained for both methane and propane hydrate dissociation. These correlations will be useful to predict the rate of dissociation and gas production in hydrate reservoirs. In the second phase of his work, in order to simulate the conditions of hydrate dissociation in the earth, methane hydrates were formed and dissociated in unconsolidated cores of sand. The results of these experiments have demonstrated that the heat transfer resistance of the media (rock) plays an important role in dissociation of hydrates in earth.

  16. Super-dissociative recombination of H2+?

    NASA Astrophysics Data System (ADS)

    Mitchell, J. B. A.; Yousif, F. B.; van der Donk, P. J. T.; Morgan, T. J.; Chibisov, M. I.

    1995-11-01

    The dissociative recombination of vibrationally excited H2+ ions to form products in high Rydberg states has been measured. Surprisingly large cross-sections are found for this channel. This seems to be an example of super-dissociative recombination.

  17. Fundamental Physical Constants

    National Institute of Standards and Technology Data Gateway

    SRD 121 CODATA Fundamental Physical Constants (Web, free access)   This site, developed in the Physics Laboratory at NIST, addresses three topics: fundamental physical constants, the International System of Units (SI), which is the modern metric system, and expressing the uncertainty of measurement results.

  18. Dissociative Mothers' Subjective Experience of Parenting.

    ERIC Educational Resources Information Center

    Benjamin, Lynn R.; And Others

    1996-01-01

    A study of 54 mothers with a dissociative disorder, 20 mothers with other mental problems, and 20 normal mothers investigated what effect, if any, dissociation had on parenting. When tested on the Subjective Experiences of Parenting Scale, mothers with dissociation presented significantly more negative parenting behavior and attitudes. (CR)

  19. Contractions of affine spherical varieties

    SciTech Connect

    Arzhantsev, I V

    1999-08-31

    The language of filtrations and contractions is used to describe the class of G-varieties obtainable as the total spaces of the construction of contraction applied to affine spherical varieties, which is well-known in invariant theory. These varieties are local models for arbitrary affine G-varieties of complexity 1 with a one-dimensional categorical quotient. As examples, reductive algebraic semigroups and three-dimensional SL{sub 2}-varieties are considered.

  20. Affine generalization of the Komar complex of general relativity

    NASA Astrophysics Data System (ADS)

    Mielke, Eckehard W.

    2001-02-01

    On the basis of the ``on shell'' Noether identities of the metric-affine gauge approach of gravity, an affine superpotential is derived which comprises the energy- and angular-momentum content of exact solutions. In the special case of general relativity (GR) or its teleparallel equivalent, the Komar or Freud complex, respectively, are recovered. Applying this to the spontaneously broken anti-de Sitter gauge model of McDowell and Mansouri with an induced Euler term automatically yields the correct mass and spin of the Kerr-AdS solution of GR with a (induced) cosmological constant without the factor two discrepancy of the Komar formula.

  1. In Vitro Opioid Receptor Affinity and in Vivo Behavioral Studies of Nelumbo nucifera Flower

    PubMed Central

    Kumarihamy, Mallika; León, Francisco; Pettaway, Sara; Wilson, Lisa; Lambert, Janet A.; Wang, Mei; Hill, Christopher; McCurdy, Christopher R.; ElSohly, Mahmoud A.; Cutler, Stephen J.; Muhammad, Ilias

    2015-01-01

    Ethnopharmacological relevance Nelumbo nucifera Geartn., known as sacred lotus, has been used traditionally in South East Asia as a traditional medicine for various CNS disorders including stress, fever, depression, insomnia, and cognitive conditions. Aim of the study To investigate the in vitro cannabinoid and opioid receptor binding affinities, and in vivo behavioral actions of Nelumbo flower extracts and to isolate the potential compounds to treat CNS associated disorders. Materials and methods The white and pink flowers of N. nucifera were extracted with 95% EtOH, followed by acid-base partitioning using CHCl3 to give acidic and basic partitions. These partitions were subjected to Centrifugal Preparative TLC (CPTLC) to yield benzyltetrahydroisoquinoline (BTIQ) alkaloids and long chain fatty acids, identified by physical and spectroscopic methods. In addition, EtOH extracts and partitions were analyzed for chemical markers by UHPLC/MS and GC/MS. In vitro neuropharmacological effects were evaluated by cannabinoid (CB1 and CB2) and opioid [delta (δ), kappa (κ), and mu (μ)] competitive radioligand binding and GTPγS functional assays. The in vivo behavioral effect was studied through the use of the mouse tetrad assay at 10, 30, 75 and 100 mg/kg/ip doses that revealed the effect on locomotion, catalepsy, body temperature, and nociception of acidic and basic CHCl3 partitions, fractions, and compounds. Results Three aporphines, nuciferine (1), N-nor-nuciferine (2), asimilobine (3), and five BTIQs, armepavine (4), O-methylcoclaurine (5), N-methylcoclaurine (6), coclaurine (7), neferine (10), and a mixture of linoleic and palmitic acids (LA and PA), were identified and evaluated for cannabinoid and opioid receptor displacement activities. Compounds 5–7 showed binding affinities for the κ opioid receptor with equilibrium dissociation constant (Ki) values of 3.5±0.3, 0.9±0.1, 2.2±0.2 µM, respectively. Compound 10 displayed affinities for δ-and μ- opioid

  2. "Recognizing Numerical Constants"

    NASA Technical Reports Server (NTRS)

    Bailey, David H.; Craw, James M. (Technical Monitor)

    1995-01-01

    The advent of inexpensive, high performance computer and new efficient algorithms have made possible the automatic recognition of numerically computed constants. In other words, techniques now exist for determining, within certain limits, whether a computed real or complex number can be written as a simple expression involving the classical constants of mathematics. In this presentation, some of the recently discovered techniques for constant recognition, notably integer relation detection algorithms, will be presented. As an application of these methods, the author's recent work in recognizing "Euler sums" will be described in some detail.

  3. Peritraumatic versus persistent dissociation in acute stress disorder.

    PubMed

    Panasetis, Paula; Bryant, Richard A

    2003-12-01

    The DSM-IV definition of acute stress disorder (ASD) regards dissociation that occurs during a trauma (peritraumatic dissociation) comparably to persistent dissociation. This study investigated the relative contributions of peritraumatic dissociation and persistent dissociation to acute posttraumatic stress reactions. Civilian trauma (N = 53) survivors with either acute stress disorder (ASD), subclinical ASD, or no ASD were administered modified versions of the Peritraumatic Dissociative Experiences Questionnaire that indexed both dissociation during the trauma and dissociation at the time of assessment. Persistent dissociation was more strongly associated with ASD severity and intrusive symptoms than peritraumatic dissociation. These results are consistent with the proposition that persistent, rather than peritraumatic, dissociation is associated with posttraumatic psychopathology.

  4. Coupling of disulfide bond and distal histidine dissociation in human ferrous cytoglobin regulates ligand binding.

    PubMed

    Beckerson, Penny; Reeder, Brandon J; Wilson, Michael T

    2015-02-13

    Earlier kinetics studies on cytoglobin did not assign functional properties to specific structural forms. Here, we used defined monomeric and dimeric forms and cysteine mutants to show that an intramolecular disulfide bond (C38-C83) alters the dissociation rate constant of the intrinsic histidine (H81) (∼1000 fold), thus controlling binding of extrinsic ligands. Through time-resolved spectra we have unequivocally assigned CO binding to hexa- and penta-coordinate forms and have made direct measurement of histidine rebinding following photolysis. We present a model that describes how the cysteine redox state of the monomer controls histidine dissociation rate constants and hence extrinsic ligand binding.

  5. Radiative capture versus Coulomb dissociation.

    SciTech Connect

    Esbensen, H.; Physics

    2006-01-01

    Measurements of the Coulomb dissociation of {sup 8}B have been used to infer the rate of the inverse radiative proton capture on {sup 7}Be. The analysis is usually based on the assumptions that the two processes are related by detailed balance and described by E1 transitions. However, there are corrections to this relation. The Coulomb form factors for the two processes, for example, are not identical. There are also E2 transitions and higher-order effects in the Coulomb dissociation, and the nuclear induced breakup cannot always be ignored. While adding first-order E2 transitions enhances the decay energy spectrum, the other mechanisms cause a suppression at low relative energies. The net result may accidentally be close to the conventional first-order E1 calculation, but there are differences which cannot be ignored if accuracies of 10% or better are needed.

  6. Affinity Interaction between Hexamer Peptide Ligand HWRGWV and Immunoglobulin G Studied by Quartz Crystal Microbalance and Surface Plasmon Resonance

    NASA Astrophysics Data System (ADS)

    Shen, Fei

    sensors for the immobilization of peptide ligands with low nonspecific binding. The silica surface was first modified by the formation of self-assembling monolayer (SAM) of 3-amino-propyl triethoxy silane as an anchor layer. Short chains of poly(ethylene glycol) (PEG) with Fmoc-protected amino groups at one end and carboxyl groups at the other end were then coupled through the carboxyl terminal to the amino groups on the silane. The short PEG chains served as spacer arms to reduce nonspecific binding to the substrate. The gold surface was modified by a two-component SAM using mixtures of HS(CH 2)11(CH2CH2O)6NH2 and HS(CH2)11(CH2CH2O)3OH. The advantage of using a modified silica surface is its relatively higher stability than the SAM on gold during the peptide functionalization step, however the SPR sensors do not work on silica surfaces. In addition, the modification process of the gold surface is relatively simple compared with that of the silica surface. The peptide immobilization process was optimized with silica surfaces and the best conditions were applied for the immobilization on gold surfaces. The results of surface modifications and peptide immobilizations were characterized by various surface analysis techniques including, ellipsometry, contact angle goniometer, chemical force microscopy (CFM), x-ray photoelectron spectroscopy (XPS) and time of flight secondary ion mass spectroscopy (ToF-SIMS). QCM and SPR results indicated that this peptide ligand HWRGWV immobilized on modified silica or gold surfaces has high affinity and specificity to hIgG binding even in a complex medium such as cMEM. Both thermodynamic and kinetic parameters of affinity interaction were obtained by the analysis of QCM and SPR data. Compared with QCM, SPR is more suitable for quantitative analysis of the protein binding, which is essential for the investigation of thermodynamics and kinetics parameters. The maximum binding capacity (4.15 mg m-2 ) and the dissociation constant (1.83 mu

  7. Kinetics of CH4 and CO2 hydrate dissociation and gas bubble evolution via MD simulation.

    PubMed

    Uddin, M; Coombe, D

    2014-03-20

    Molecular dynamics simulations of gas hydrate dissociation comparing the behavior of CH4 and CO2 hydrates are presented. These simulations were based on a structurally correct theoretical gas hydrate crystal, coexisting with water. The MD system was first initialized and stabilized via a thorough energy minimization, constant volume-temperature ensemble and constant volume-energy ensemble simulations before proceeding to constant pressure-temperature simulations for targeted dissociation pressure and temperature responses. Gas bubble evolution mechanisms are demonstrated as well as key investigative properties such as system volume, density, energy, mean square displacements of the guest molecules, radial distribution functions, H2O order parameter, and statistics of hydrogen bonds. These simulations have established the essential similarities between CH4 and CO2 hydrate dissociation. The limiting behaviors at lower temperature (no dissociation) and higher temperature (complete melting and formation of a gas bubble) have been illustrated for both hydrates. Due to the shift in the known hydrate stability curves between guest molecules caused by the choice of water model as noted by other authors, the intermediate behavior (e.g., 260 K) showed distinct differences however. Also, because of the more hydrogen-bonding capability of CO2 in water, as reflected in its molecular parameters, higher solubility of dissociated CO2 in water was observed with a consequence of a smaller size of gas bubble formation. Additionally, a novel method for analyzing hydrate dissociation based on H-bond breakage has been proposed and used to quantify the dissociation behaviors of both CH4 and CO2 hydrates. Activation energies Ea values from our MD studies were obtained and evaluated against several other published laboratory and MD values. Intrinsic rate constants were estimated and upscaled. A kinetic reaction model consistent with macroscale fitted kinetic models has been proposed to

  8. The cosmological constant problem

    SciTech Connect

    Dolgov, A.D.

    1989-05-01

    A review of the cosmological term problem is presented. Baby universe model and the compensating field model are discussed. The importance of more accurate data on the Hubble constant and the Universe age is stressed. 18 refs.

  9. Science Is Constantly Cool.

    ERIC Educational Resources Information Center

    Eichinger, John

    1996-01-01

    Presents an activity in which students attempt to keep water at a constant temperature. Helps students in grades three to six hone their skills in prediction, observation, measurement, data collection, graphing, data analysis, and communication. (JRH)

  10. Thermal Dissociation of Halogen Azides

    DTIC Science & Technology

    1994-09-01

    inefficient due to the low gain provided by this emitter and the short duration of the shock tube experiment . Much higher gain coefficients were...with the SF6 sensitizers (in the C02 laser triggered experiment ) and thermal dissociation of the tetraazldodiborane donor ;nto non-productive...Excitation of FN3/S2CI 2/SF6 gas mixtures by the pulsed C02 laser yielded NS(B-X) chemiluminescence 5 . Cold trap experiments showed that small levels

  11. The single Cys2-His2 zinc finger domain of the GAGA protein flanked by basic residues is sufficient for high-affinity specific DNA binding.

    PubMed Central

    Pedone, P V; Ghirlando, R; Clore, G M; Gronenborn, A M; Felsenfeld, G; Omichinski, J G

    1996-01-01

    Specific DNA binding to the core consensus site GAGAGAG has been shown with an 82-residue peptide (residues 310-391) taken from the Drosophila transcription factor GAGA. Using a series of deletion mutants, it was demonstrated that the minimal domain required for specific binding (residues 310-372) includes a single zinc finger of the Cys2-His2 family and a stretch of basic amino acids located on the N-terminal end of the zinc finger. In gel retardation assays, the specific binding seen with either the peptide or the whole protein is zinc dependent and corresponds to a dissociation constant of approximately 5 x 10(-9) M for the purified peptide. It has previously been thought that a single zinc finger of the Cys2-His2 family is incapable of specific, high-affinity binding to DNA. The combination of an N-terminal basic region with a single Cys2-His2 zinc finger in the GAGA protein can thus be viewed as a novel DNA binding domain. This raises the possibility that other proteins carrying only one Cys2-His2 finger are also capable of high-affinity specific binding to DNA. Images Fig. 2 Fig. 3 Fig. 4 PMID:8610125

  12. A biomimetic Protein G affinity adsorbent: an Ugi ligand for immunoglobulins and Fab fragments based on the third IgG-binding domain of Protein G.

    PubMed

    El Khoury, Graziella; Lowe, Christopher R

    2013-04-01

    This work reports the development of a synthetic affinity adsorbent for immunoglobulins based on the Fab-binding domain of Streptococcal Protein G (SpG-domain III). The ligand (A2C7I1) was synthesized by the four-component Ugi reaction to generate a substituted peptoidal scaffold mimicking key amino acid residues of SpG. Computer-aided analysis suggests a putative binding site on the CH 1 domain of the Fab molecule. In silico studies, supported by affinity chromatography in comparison with immobilized SpG, as well as analytical characterization by liquid chromatography/electrospray ionization-mass spectrometry and (1) H nuclear magnetic resonance of the ligand synthesized in solution, indicated the authenticity and suitability of the designed ligand for the purification of immunoglobulins. The immobilized ligand displayed an apparent static binding capacity of ~17 mg IgG ml(-1) and a dissociation constant of 5.34 × 10(-5)  M. Preparative chromatography demonstrated the ability of the immobilized ligand to purify IgG and Fab fragments from crude mammalian and yeast cell cultures, under near physiological ionic strength and pH, to yield proteins of 99% and 93% purity, respectively.

  13. Tityus gamma toxin, a high affinity effector of the Na+ channel in muscle, with a selectivity for channels in the surface membrane.

    PubMed

    Barhanin, J; Ildefonse, M; Rougier, O; Sampaio, S V; Giglio, J R; Lazdunski, M

    1984-01-01

    Toxin gamma from the venom of Tityus serrulatus scorpion produces a partial block of the surface Na+ channel in frog muscle. This block occurs with no change in the voltage-dependence or in the kinetics of the remaining surface Na+ current. The partial blockade of Na+ channel activity occurs with no change in tubular Na+ currents nor in twitch tension. The maximum effect of the toxin is attained at concentrations as low as 3 X 10(-10) M. Hyperpolarization to potentials more negative than the resting potential (E = -90 mV) reduces or abolishes the effect of the toxin. Radioiodinated toxin gamma binds to frog muscle membranes with a very high affinity corresponding to a dissociation constant of about 1 X 10(-11) M. Data obtained with both rabbit and frog muscle indicate that toxin gamma is specific for Na+ channels in surface membranes. Toxin gamma does not seem to bind to Na+ channels in T-tubule membranes. The biochemical data are in good agreement with electrophysiological studies and data on contraction. There is one Tityus gamma toxin binding site per tetrodotoxin binding site in surface membranes. Competition experiments have confirmed that Tityus gamma toxin binds to a new toxin receptor site on the Na+ channel structure. This site is the same that the toxin II from Centruroides suffusus binding site, but this toxin has 100 times less affinity for the Na+ channel than Tityus gamma toxin.

  14. Dopamine inhibition of anterior pituitary adenylate cyclase is mediated through the high-affinity state of the D/sub 2/ receptor

    SciTech Connect

    Borgundvaag, B.; George, S.R.

    1985-07-29

    The diterpinoid forskolin stimulated adenylate cyclase activity (measured by conversion of (/sup 3/H)-ATP to (/sup 3/H)-cAMP) in anterior pituitary from male and female rats. Inhibition of stimulated adenylate cyclase activity by potent dopaminergic agonists was demonstrable only in female anterior pituitary. The inhibition of adenylate cyclase activity displayed a typically dopaminergic rank order of agonist potencies and could be completely reversed by a specific dopamine receptor antagonist. The IC/sub 50/ values of dopamine agonist inhibition of adenylate cyclase activity correlated with equal molarity with the dissociation constant of the high-affinity dopamine agonist-detected receptor binding site and with the IC/sub 50/ values for inhibition of prolactin secretion. These findings support the hypothesis that it is the high-affinity form of the D/sub 2/ dopamine receptor in anterior pituitary which is responsible for mediating the dopaminergic function of attenuating adenylate cyclase activity. 12 references, 4 figures, 1 table.

  15. Gonadotropin stimulation of cyclic adenosine monophosphate and testosterone production without detectable high-affinity binding sites in purified Leydig cells from rat testis

    SciTech Connect

    Browne, E.S.; Bhalla, V.K. )

    1991-02-01

    Rat testicular interstitial cells were separated by three different gradient-density procedures and, with each, two biochemically and morphologically distinct cell fractions were isolated. The lighter density cells in fraction-I bound iodine 125-labeled human chorionic gonadotropin (hCG) with high-affinity (apparent equilibrium dissociation constant, Kd, approximately 10{sup {minus} 10} M) without producing either cyclic adenosine monophosphate or testosterone in response to hormone action. The heavier-density cells displayed morphologic features typical of Leydig cells and produced cyclic adenosine monophosphate and testosterone in the presence of hCG without detectable {sup 125}I-labeled hCG high-affinity binding. These cell fractions were further characterized by studies using deglycosylated hCG, a known antagonist to hCG action. Cell concentration-dependent studies with purified Leydig cells revealed that maximal testosterone production was achieved when lower cell concentrations (0.5 x 10(6) cells/250 microliters) were used for in vitro hCG stimulation assays. Under these conditions, the {sup 125}I-labeled hCG binding was barely detectable (2.24 fmol; 2,698 sites/cell). Furthermore, these studies revealed that the hCG-specific binding in Leydig cells is overestimated by the classic method for nonspecific binding correction using excess unlabeled hormone. An alternate method is presented.

  16. Allostery in the lac operon: population selection or induced dissociation?

    PubMed

    Sharp, Kim A

    2011-11-01

    Allostery, the modulation of function of a protein at one site by the binding of a ligand at a different site, is a property of many proteins. Two kinetically distinct models have been proposed: i) The induced fit model in which the ligand binds to the protein and then induces the conformational change. ii) The population selection model, in which the protein spontaneously undergoes a conformational change, which is then 'captured' by the ligand. Using measured kinetic constants for the lac repressor the contribution of population selection vs. induced dissociation is quantified by simulating the kinetics of allostery. At very low inducer concentration, both mechanisms contribute significantly. Total induction, though, is small under these conditions. At increasing levels of induction the induced dissociation mechanism soon dominates, first due to binding of one inducer, and then from two inducers binding.

  17. [Differential diagnosis between dissociative disorders and schizophrenia].

    PubMed

    Shibayama, Masatoshi

    2011-01-01

    The differential diagnosis of dissociative disorders includes many psychiatric disorders, such as schizophrenia, bipolar disorders (especially bipolar II disorder), depressive disorder (especially atypical depression), epilepsy, Asperger syndrome, and borderline personality disorder. The theme of this paper is the differential diagnosis between dissociative disorders and schizophrenia. Schneiderian first-rank symptoms in schizophrenia are common in dissociative disorders, especially in dissociative identity disorder (DID). Many DID patients have been misdiagnosed as schizophrenics and treated with neuroleptics. We compared and examined Schneiderian symptoms of schizophrenia and those of dissociative disorders from a structural viewpoint. In dissociative disorders, delusional perception and somatic passivity are not seen. "Lateness" and "Precedence of the Other" originated from the concept of "Pattern Reversal" (H. Yasunaga)" is characteristic of schizophrenia. It is important to check these basic structure of schizophrenia in subjective experiences in differential diagnosis between dissociative disorders and schizophrenia.

  18. Limits for antibody affinity maturation and repertoire diversification in hypervaccinated humans.

    PubMed

    Poulsen, Tine Rugh; Jensen, Allan; Haurum, John S; Andersen, Peter S

    2011-10-15

    The immune system is known to generate a diverse panel of high-affinity Abs by adaptively improving the recognition of pathogens during ongoing immune responses. In this study, we report the biological limits for Ag-driven affinity maturation and repertoire diversification by analyzing Ab repertoires in two adult volunteers after each of three consecutive booster vaccinations with tetanus toxoid. Maturation of on-rates and off-rates occurred independently, indicating a kinetically controlled affinity maturation process. The third vaccination induced no significant changes in the distribution of somatic mutations and binding rate constants implying that the limits for affinity maturation and repertoire diversification had been reached. These fully matured Ab repertoires remained similar in size, genetically diverse, and dynamic. Somatic mutations and kinetic rate constants showed normal and log-normal distribution profiles, respectively. Mean values can therefore be considered as biological constants defining the observed boundaries. At physiological temperature, affinity maturation peaked at k(on) = 1.6 × 10(4) M(-1) s(-1) and k(off) = 1.7 × 10(-4) s(-1) leading to a maximum mean affinity of K(D) = 1.0 × 10(-9) M. At ambient temperature, the average affinity increased to K(D) = 3.4 × 10(-10) M mainly due to slower off-rates. This experimentally determined set of constants can be used as a benchmark for analysis of the maturation level of human Abs and Ab responses.

  19. Charge States of y Ions in the Collision-Induced Dissociation of Doubly Charged Tryptic Peptide Ions

    NASA Astrophysics Data System (ADS)

    Neta, Pedatsur; Stein, Stephen E.

    2011-05-01

    Bonds that break in collision-induced dissociation (CID) are often weakened by a nearby proton, which can, in principle, be carried away by either of the product fragments. Since peptide backbone dissociation is commonly charge-directed, relative intensities of charge states of product y- and b-ions depend on the final location of that proton. This study examines y-ion charge distributions for dissociation of doubly charged peptide ions, using a large reference library of peptide ion fragmentation generated from ion-trap CID of peptide ions from tryptic digests. Trends in relative intensities of y2+ and y1+ ions are examined as a function of bond cleavage position, peptide length (n), residues on either side of the bond and effects of residues remote from the bond. It is found that yn-2/b2 dissociation is the most sensitive to adjacent amino acids, that y2+/y1+ steadily increase with increasing peptide length, that the N-terminal amino acid can have a major influence in all dissociations, and in some cases other residues remote from the bond cleavage exert significant effects. Good correlation is found between the values of y2+/y1+ for the peptide and the proton affinities of the amino acids present at the dissociating peptide bond. A few deviations from this correlation are rationalized by specific effects of the amino acid residues. These correlations can be used to estimate trends in y2+/y1+ ratios for peptide ions from amino acid proton affinities.

  20. [Clinical Handling of Patients with Dissociative Disorders].

    PubMed

    Okano, Kenichiro

    2015-01-01

    This paper discusses the way informed psychiatrists are expected to handle dissociative patients in clinical situations, with a specific focus on dissociative identity disorders and dissociative fugue. On the initial interview with dissociative patients, information on their history of trauma and any nascent dissociative symptoms in their childhood should be carefully obtained. Their level of stress in their current life should also be assessed in order to understand their symptomatology, as well as to predict their future clinical course. A psychoeducational approach is crucial; it might be helpful to give information on dissociative disorder to these patients as well as their family members in order to promote their adherence to treatment. Regarding the symptomatology of dissociative disorders, detailed symptoms and the general clinical course are presented. It was stressed that dissociative identity disorder and dissociative fugue, the most high-profile dissociative disorders, are essentially different in their etiology and clinical presentation. Dissociative disorders are often confused with and misdiagnosed as psychotic disorders, such as schizophrenia. Other conditions considered in terms of the differential diagnosis include borderline personality disorder as well as temporal lobe epilepsy. Lastly, the therapeutic approach to dissociative identity disorder is discussed. Each dissociative identity should be understood as potentially representing some traumatically stressful event in the past. The therapist should be careful not to excessively promote the creation or elaboration of any dissociative identities. Three stages are proposed in the individual psychotherapeutic process. In the initial stage, a secure environment and stabilization of symptoms should be sought. The second stage consists of aiding the "host" personality to make use of other more adaptive coping skills in their life. The third stage involves coaching as well as continuous awareness of

  1. Dynamic friction of self-affine surfaces

    NASA Astrophysics Data System (ADS)

    Schmittbuhl, Jean; Vilotte, Jean-Pierre; Roux, Stéphane

    1994-02-01

    We investigate the velocity dependence of the friction between two rigid blocks limited by a self-affine surface such as the one generated by a crack. The upper solid is subjected either to gravity or to an external elastic stiffness, and is driven horizontally at constant velocity, V, while the lower solid is fixed. For low velocities, the apparent friction coefficient is constant. For high velocities, the apparent friction is shown to display a velocity weakening. The weakening can be related to the variation of the mean contact time due to the occurrence of jumps during the motions. The cross-over between these two regimes corresponds to a characteristic velocity which depends on the geometry of the surfaces and on the mean normal force. In the case of simple gravity loading, the velocity dependence of the apparent friction at high velocities is proportional to 1/V^2 where V is the imposed tangential velocity. In the case of external elastic stiffness, two velocity weakening regimes can be identified, the first is identical to the gravity case with a 1/V^2 dependence, the second appears at higher velocities and is characterized by a 1/V variation. The characteristic velocity of this second cross-over depends on the roughness and the elastic stiffness. The statistical distribution of ballistic flight distances is analysed, and is shown to reveal in all cases the self-affinity of the contacting surfaces. Nous analysons la dépendence en vitesse du frottement entre deux solides limités par une surface rugueuse auto-affine comme celle d'une surface de fracture. Le solide supérieur est soumis soit à la gravité, soit à une raideur élastique externe, et est entraîné à vitesse horizontale constante V sur le solide inférieur fixe. A faible vitesse, le coefficient de friction apparent, est constant. A forte vitesse, le coefficient de friction apparent devient inversement proportionnel à la vitesse. Cette dépendance peut être reliée à la variation du temps

  2. Reexamination of the I 2 spectrum near the B( 3Π0u+) state dissociation limit

    NASA Astrophysics Data System (ADS)

    Tromp, John W.; Le Roy, Robert J.; Gerstenkorn, Simon; Luc, Paul

    1983-07-01

    The disagreement of Danyluk and King's ( Chem. Phys.25, 343 (1977)) rotational constants for levels lying near the dissociation limit of B-state I 2 with the mechanical behavior predicted by near-dissociation theory is investigated. The discrepancies are shown to be much too large to be explained by either the neglect of centrifugal distortion effects in the original analysis or by rotational or spin-rotation coupling to a nearby repulsive 1 u state. These differences are therefore attributed to experimental error, a conclusion which is confirmed by more recent experimental results. A reanalysis of the best available data for levels near the dissociation limit of B-state I 2 then yields improved values for the B-state dissociation limit D = 20 043.16 (±0.02) cm -1 of the vibrational index at dissociation vD = 87.32 (±0.04) and of the long-range potential constant C 5 = 2.88 (±0.03) × 10 5cm -1Å5. This in turn implies a slightly improved ground-state dissociation energy of D0 = 12 440.18 (±0.02) cm -1.

  3. Binding of a Calcium Antagonist, [3H]Nitrendipine, to High Affinity Sites in Bovine Aortic Smooth Muscle and Canine Cardiac Membranes

    PubMed Central

    Williams, Lewis T.; Tremble, Patrice

    1982-01-01

    [3H]Nitrendipine, a potent calcium channel antagonist [3-ethyl-5-methyl-1-1,4-dihydro-2,6 - dimethyl - 4 - (3 - nitrophenyl) - 3,5 - pyridine carboxylate], was used to label high affinity binding sites on membranes prepared from bovine aortic smooth muscle. The binding of [3H]nitrendipine is rapid (t1/2 < 5 min) and reversible at 37°C. The binding sites have a high affinity for [3H]nitrendipine with an equilibrium dissociation constant of 2.1 nM. The density of sites is 40-60 fmol/mg of membrane protein. Analogues of nitrendipine compete for the binding sites with affinities consistent with their known biological effects as calcium antagonists. Nisoldipine, [isobutyl methyl 1,4-dihydro-2,6-dimethyl-4-(2-nitrophenyl)-3,5-pyridine carboxylate], a calcium antagonist more potent than nifedipine [2,6-dimethyl-3,5-dicarbomethoxy-4-(2-nitrophenyl)-1,4-dihydropyridine] in relaxing vascular smooth muscle, has an affinity three-fold higher than that of nifedipine in competing for the binding sites. A biologically inactive derivative of nifedipine does not compete for [3H]nitrendipine binding. Verapamil (α-isopropyl-α[(N-methyl - N-homoveratryl) -α-aminopropyl]-3,4-dimethyoxyphenyl acetonitrile), a structurally different calcium antagonist, only partially (25%) inhibits binding at high concentrations (1 μM). Prazosin, an alpha adrenergic antagonist does not compete for [3H]nitrendipine binding sites. The binding of [3H]nitrendipine is not affected by 1.5 mM calcium. Canine cardiac membranes also have high affinity [3H]nitrendipine binding sites, (KD = 6 nM) but bovine erythrocytes do not. The relative affinities of nisoldipine and nifedipine for the cardiac membrane binding sites reflect the relative activities of these compounds as calcium channel antagonists. These results suggest that the [3H]nitrendipine binding sites are the sites through which dihydropyridines act as calcium channel antagonists. PMID:6282938

  4. Dissociative experiences on ice--peritraumatic and trait dissociation during the cold pressor test.

    PubMed

    Giesbrecht, Timo; Smeets, Tom; Merckelbach, Harald

    2008-01-15

    The present study investigated the relationship between trait dissociation and peritraumatic dissociation during acute painful stimulation. In a sample of 70 undergraduate students, peritraumatic dissociation was induced by means of a cold pressor test, which basically consists of participants holding an arm in ice water for as long as possible. Results indicate that heightened trait dissociation scores were related to shorter durations that participants could sustain the task. However, trait dissociation was not associated with increases in acute dissociative symptoms (i.e., peritraumatic dissociation). These findings are in sharp contrast to the defensive function ascribed to both types of dissociation and also the commonly held assumption that peritraumatic and trait dissociation are intimately linked.

  5. The influence of porosity and structural parameters on different kinds of gas hydrate dissociation

    NASA Astrophysics Data System (ADS)

    Misyura, S. Y.

    2016-07-01

    Methane hydrate dissociation at negative temperatures was studied experimentally for different artificial and natural samples, differing by macro- and micro-structural parameters. Four characteristic dissociation types are discussed in the paper. The internal kinetics of artificial granule gas hydrates and clathrate hydrates in coal is dependent on the porosity, defectiveness and gas filtration rate. The density of pores distribution in the crust of formed ice decreases by the several orders of magnitude and this change significantly the rate of decay. Existing models for describing dissociation at negative temperatures do not take into account the structural parameters of samples. The dissociation is regulated by internal physical processes that must be considered in the simulation. Non-isothermal dissociation with constant external heat flux was simulated numerically. The dissociation is simulated with consideration of heat and mass transfer, kinetics of phase transformation and gas filtering through a porous medium of granules for the negative temperatures. It is shown that the gas hydrate dissociation in the presence of mainly microporous structures is fundamentally different from the disintegration of gas hydrates containing meso and macropores.

  6. The influence of porosity and structural parameters on different kinds of gas hydrate dissociation.

    PubMed

    Misyura, S Y

    2016-07-22

    Methane hydrate dissociation at negative temperatures was studied experimentally for different artificial and natural samples, differing by macro- and micro-structural parameters. Four characteristic dissociation types are discussed in the paper. The internal kinetics of artificial granule gas hydrates and clathrate hydrates in coal is dependent on the porosity, defectiveness and gas filtration rate. The density of pores distribution in the crust of formed ice decreases by the several orders of magnitude and this change significantly the rate of decay. Existing models for describing dissociation at negative temperatures do not take into account the structural parameters of samples. The dissociation is regulated by internal physical processes that must be considered in the simulation. Non-isothermal dissociation with constant external heat flux was simulated numerically. The dissociation is simulated with consideration of heat and mass transfer, kinetics of phase transformation and gas filtering through a porous medium of granules for the negative temperatures. It is shown that the gas hydrate dissociation in the presence of mainly microporous structures is fundamentally different from the disintegration of gas hydrates containing meso and macropores.

  7. The influence of porosity and structural parameters on different kinds of gas hydrate dissociation

    PubMed Central

    Misyura, S. Y.

    2016-01-01

    Methane hydrate dissociation at negative temperatures was studied experimentally for different artificial and natural samples, differing by macro- and micro-structural parameters. Four characteristic dissociation types are discussed in the paper. The internal kinetics of artificial granule gas hydrates and clathrate hydrates in coal is dependent on the porosity, defectiveness and gas filtration rate. The density of pores distribution in the crust of formed ice decreases by the several orders of magnitude and this change significantly the rate of decay. Existing models for describing dissociation at negative temperatures do not take into account the structural parameters of samples. The dissociation is regulated by internal physical processes that must be considered in the simulation. Non-isothermal dissociation with constant external heat flux was simulated numerically. The dissociation is simulated with consideration of heat and mass transfer, kinetics of phase transformation and gas filtering through a porous medium of granules for the negative temperatures. It is shown that the gas hydrate dissociation in the presence of mainly microporous structures is fundamentally different from the disintegration of gas hydrates containing meso and macropores. PMID:27445113

  8. Dynamics of nitrogen dissociation from direct molecular simulation

    NASA Astrophysics Data System (ADS)

    Valentini, Paolo; Schwartzentruber, Thomas E.; Bender, Jason D.; Candler, Graham V.

    2016-08-01

    We present a molecular-level investigation of nitrogen dissociation at high temperature. The computational technique, called direct molecular simulation (DMS), solely relies on an ab initio potential energy surface and both N2+N2 and N +N2 processes are simulated as they concurrently take place in an evolving nonequilibrium gas system. Quasiclassical trajectory calculations (QCT) reveal that dissociation rate coefficients calculated at thermal equilibrium, i.e., assuming Boltzmann energy distributions, are approximately equal (within less than 15%) for both N2+N2 and N +N2 collisions for the range of temperatures considered. The DMS (nonequilibrium) results indicate, however, that the presence of atomic nitrogen significantly affects the dissociation rate of molecular nitrogen, but indirectly. In fact, the presence of atomic nitrogen causes an important reduction of the vibrational relaxation time of N2, by almost one order of magnitude. This, in turn, speeds up the replenishment of high-v states that are otherwise significantly depleted if only N2+N2 collisions are considered. Because of the strong favoring of dissociation from high-v states, this results in dissociation rates that are 2-3 times higher when significant atomic nitrogen is present compared to systems composed of mainly diatomic nitrogen, such as during the initial onset of dissociation. Specifically, we find that exchange events occur frequently during N +N2 collisions and that such exchange collisions constitute an effective mechanism of scrambling the internal energy states, resulting in multiquantum jumps in vibrational energy levels that effectively promote energy transfers. The resulting vibrational relaxation time constant we calculate for N +N2 collisions is significantly lower than the widely used Millikan-White model. Significant discrepancies are found between predictions of the Park two-temperature model (using the Millikan-White vibrational relaxation model) and the DMS results for

  9. Barrierless association of CF2 and dissociation of C2F4 by variational transition-state theory and system-specific quantum Rice-Ramsperger-Kassel theory.

    PubMed

    Bao, Junwei Lucas; Zhang, Xin; Truhlar, Donald G

    2016-11-29

    Bond dissociation is a fundamental chemical reaction, and the first principles modeling of the kinetics of dissociation reactions with a monotonically increasing potential energy along the dissociation coordinate presents a challenge not only for modern electronic structure methods but also for kinetics theory. In this work, we use multifaceted variable-reaction-coordinate variational transition-state theory (VRC-VTST) to compute the high-pressure limit dissociation rate constant of tetrafluoroethylene (C2F4), in which the potential energies are computed by direct dynamics with the M08-HX exchange correlation functional. To treat the pressure dependence of the unimolecular rate constants, we use the recently developed system-specific quantum Rice-Ramsperger-Kassel theory. The calculations are carried out by direct dynamics using an exchange correlation functional validated against calculations that go beyond coupled-cluster theory with single, double, and triple excitations. Our computed dissociation rate constants agree well with the recent experimental measurements.

  10. Dielectric Constant of Suspensions

    NASA Astrophysics Data System (ADS)

    Mendelson, Kenneth S.; Ackmann, James J.

    1997-03-01

    We have used a finite element method to calculate the dielectric constant of a cubic array of spheres. Extensive calculations support preliminary conclusions reported previously (K. Mendelson and J. Ackmann, Bull. Am. Phys. Soc. 41), 657 (1996).. At frequencies below 100 kHz the real part of the dielectric constant (ɛ') shows oscillations as a function of the volume fraction of suspension. These oscillations disappear at low conductivities of the suspending fluid. Measurements of the dielectric constant (J. Ackmann, et al., Ann. Biomed. Eng. 24), 58 (1996). (H. Fricke and H. Curtis, J. Phys. Chem. 41), 729 (1937). are not sufficiently sensitive to show oscillations but appear to be consistent with the theoretical results.

  11. Elastic constants of calcite

    USGS Publications Warehouse

    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.

  12. Comprehensive comparison of collision induced dissociation and electron transfer dissociation.

    PubMed

    Molina, Henrik; Matthiesen, Rune; Kandasamy, Kumaran; Pandey, Akhilesh

    2008-07-01

    Electron transfer dissociation (ETD) is a recently introduced mass spectrometric technique which has proven to be an excellent tool for the elucidation of labile post-translational modifications such as phosphorylation and O-GlcNAcylation of serine and threonine residues. However, unlike collision induced dissociation (CID), which has been studied for decades, the intricacies of ETD-based fragmentation have not yet been firmly established or systematically addressed. In this analysis, we have systematically compared the CID and ETD fragmentation patterns for the large majority of the peptides that do not contain such labile modifications. Using a standard 48 protein mix, we were able to measure false-positive rates for the experiments and also assess a large number of peptides for a detailed comparison of CID and ETD fragmentation pattern. Analysis of approximately 19,000 peptides derived from both standard proteins and complex protein samples revealed that (i) CID identified 50% more peptides than ETD; (ii) ETD resulted in approximately 20% increase in amino acid sequence coverage over CID; and (iii) combining CID and ETD fragmentation increased the sequence coverage for an average tryptic peptide to 92%. Interestingly, our analysis revealed that nearly 60% of all ETD-identified peptides carried two positive charges, which is in sharp contrast to what has been generally accepted. We also present a novel strategy for automatic validation of peptide assignments based on identification of a peptide by consecutive CID and ETD fragmentation in an alternating mode.

  13. Dissociation of Retinal Ganglion Cells Without Enzymes

    PubMed Central

    Hayashida, Yuki; Partida, Gloria J.; Ishida, Andrew T.

    2011-01-01

    We describe here methods for dissociating retinal ganglion cells from adult goldfish and rat without proteolytic enzymes, and show responses of ganglion cells isolated this way to step-wise voltage changes and fluctuating current injections. Taking advantage of the laminar organization of vertebrate retinas, photoreceptors and other cells were lifted away from the distal side of freshly isolated goldfish retinas, after contact with pieces of membrane filter. Likewise, cells were sliced away from the distal side of freshly isolated rat retinas, after these adhered to a membrane filter. The remaining portions of retina were incubated in an enzyme-free, low Ca2+ solution, and triturated. After aliquots of the resulting cell suspension were plated, ganglion cells could be identified by dye retrogradely transported via the optic nerve. These cells showed no obvious morphological degeneration for several days of culture. Perforated-patch whole-cell recordings showed that the goldfish ganglion cells spike tonically in response to depolarizing constant current injections, that these spikes are temporally precise in response to fluctuating current injections, and that the largest voltage-gated Na+ currents of these cells were larger than those of ganglion cells isolated with a neutral protease. PMID:15196824

  14. Chemical binding affinity estimation using MSB

    NASA Astrophysics Data System (ADS)

    Weaver, John B.; Rauwerdink, Adam M.

    2011-03-01

    Binding affinity can be estimated in several ways in the laboratory but there is no viable way to estimate binding affinity in vivo without assumptions on the number of binding sites. Magnetic spectroscopy of nanoparticle Brownian motion, MSB, measures the rotational Brownian motion. The MSB signal is affected by nanoparticle binding affinity so it provides a mechanism to measure the chemical binding affinity. We present a possible mechanism to quantify the binding affinity and test that mechanism using viscous solutions.

  15. Coenzyme-like ligands for affinity isolation of cholesterol oxidase.

    PubMed

    Xin, Yu; Lu, Liushen; Wang, Qing; Zhang, Ling; Tong, Yanjun; Wang, Wu

    2016-05-15

    Two coenzyme-like chemical ligands were designed and synthesized for affinity isolation of cholesterol oxidase (COD). To simulate the structure of natural coenzyme of COD (flavin adenine dinucleotide (FAD)), on Sepharose beads, 5-aminouracil, cyanuric chloride and 1, 4-butanediamine were composed and then modified. The COD gene from Brevibacterium sp. (DQ345780) was expressed in Escherichia coli BL21 (DE3), and then the sorbents were applied to adsorption analysis with the pure enzyme. Subsequently, the captured enzyme was applied to SDS-PAGE and activity analysis. As calculated, the theoretical maximum adsorption (Qmax) of the two affinity sorbents (RL-1 and RL-2) were ∼83.5 and 46.3mg/g wet gel; and the desorption constant Kd of the two sorbents were ∼6.02×10(-4) and 1.19×10(-4)μM. The proteins after cell lysis were applied to affinity isolation, and then after one step of affinity binding on the two sorbents, the protein recoveries of RL-1 and RL-2 were 9.2% and 9.7%; the bioactivity recoveries were 92.7% and 91.3%, respectively. SDS-PAGE analysis revealed that the purities of COD isolated with the two affinity sorbents were approximately 95%.

  16. Switchable binding affinity of mannose tethered to collagen peptide by temperature-dependent triple-helix formation.

    PubMed

    Okada, Tomoko; Isobe, Chika; Wada, Takeaki; Ezaki, Sumiyo; Minoura, Norihiko

    2013-06-19

    Novel glycopeptides were created with a view to regulate the bindings of carbohydrates to lectins as a means of controlling biological function. We synthesized glycopeptides containing mannose (Man) tethered to a collagen peptide moiety (MPOG10: -(Pro-Hyp-Gly)10- or MGPP10: -(Gly-Pro-Pro)10-). Circular dichroism spectra showed formation of a triple helical structure for MPOG10, and the melting temperature indicates that MPOG10 forms a more stable triple helical structure than MGPP10 in phosphate buffered saline (PBS). At 25 °C, fluorescence polarization (FP) values of MPOG10 and MGPP10 increased following the addition of concanavalin A (ConA), and the addition of α-methyl-mannose (MeMan) to a mixed solution of each glycopeptide with ConA resulted in a decrease in FP values. These results confirm that the previous increase in FP values observed was caused by ConA binding to Man on MPOG10 or MGPP10. The binding affinity of MPOG10 was higher than that of MGPP10, and the dissociation constant of MPOG10 to ConA was 1.9 × 10(-5) (mol/L). The observed binding of MPOG10 to ConA at 25 °C was reduced at higher temperature (50 °C). Therefore, the enhanced binding affinity of MPOG10 to ConA could be accounted for by formation of a clustered Man moiety triggered by the formation of a more stable triple helical structure of MPOG10 compared with MGPP10.

  17. Affinity labeling of the plasma membrane 3,3',5-triiodo-L-thyronine receptor in GH3 cells.

    PubMed Central

    Horiuchi, R; Johnson, M L; Willingham, M C; Pastan, I; Cheng, S

    1982-01-01

    The binding of 3,3',5-triiodo-L-thyronine (T3) to GH3 rat pituitary tumor cells was studied at 15 degrees C and was shown to be saturable, reversible, and stereospecific. Least-squares analysis of the binding data showed two classes of binding sites with dissociation constants of 1.8 +/- 0.2 nM and 260 +/- 30 nM and binding capacities of (5.2 +/- 0.2) X 10(4) and (1.6 +/- 0.2) X 10(6) sites per cell, respectively. Affinity labeling of intact cells was carried out by incubation of cells with 0.3 nM N-bromoacetyl-[125I]T3 at 15 degrees C for 1 hr. Analysis of the cellular extracts by sodium dodecyl sulfate gel electrophoresis showed three labeled protein bands with apparent molecular masses of 55, 47, and 33 kilodaltons (kDal) in a ratio of 86:7:7. The labeling of only the 55-kDal protein band was selectively reduced to 50% by 20 microM unlabeled T3. Highly purified plasma membranes of GH3 cells were prepared and shown to be free of nuclei. Affinity labeling of the purified plasma membranes gave the same labeling pattern as with intact cells. Peptide mapping by Staphylococcus aureus V8 digestion of the 55-kDal protein from cells or plasma membranes gave the identical peptide fragments. Thus the 55-kDal protein labeled from intact cells is the same protein as that from purified plasma membranes. These results together with our earlier findings [Horiuchi, R., Cheng, S.-y., Willingham, M. & Pastan, I. (1982) J. Biol. Chem. 257, 3139-3144] suggest that the 55-kDal protein may be involved in mediating the uptake of T3 in GH3 cells. Images PMID:6291049

  18. Measurement of free glucocorticoids: quantifying corticosteroid-binding globulin binding affinity and its variation within and among mammalian species

    PubMed Central

    Delehanty, Brendan; Hossain, Sabrina; Jen, Chao Ching; Crawshaw, Graham J.; Boonstra, Rudy

    2015-01-01

    Plasma glucocorticoids (GCs) are commonly used as measures of stress in wildlife. A great deal of evidence indicates that only free GC (GC not bound by the specific binding protein, corticosteroid-binding globulin, CBG) leaves the circulation and exerts biological effects on GC-sensitive tissues. Free hormone concentrations are difficult to measure directly, so researchers estimate free GC using two measures: the binding affinity and the binding capacity in plasma. We provide an inexpensive saturation binding method for calculating the binding affinity (equilibrium dissociation constant, Kd) of CBG that can be run without specialized laboratory equipment. Given that other plasma proteins, such as albumin, also bind GCs, the method compensates for this non-specific binding. Separation of bound GC from free GC was achieved with dextran-coated charcoal. The method provides repeatable estimates (12% coefficient of variation in the red squirrel, Tamiasciurus hudsonicus), and there is little evidence of inter-individual variation in Kd (range 2.0–7.3 nM for 16 Richardson's ground squirrels, Urocitellus richardsonii). The Kd values of 28 mammalian species we assessed were mostly clustered around a median of 4 nM, but five species had values between 13 and 61 nM. This pattern may be distinct from birds, for which published values are more tightly distributed (1.5–5.1 nM). The charcoal separation method provides a reliable and robust method for measuring the Kd in a wide range of species. It uses basic laboratory equipment to provide rapid results at very low cost. Given the importance of CBG in regulating the biological activity of GCs, this method is a useful tool for physiological ecologists. PMID:27293705

  19. Religious Dissociation and Economic Appraisal in Brazil.

    PubMed

    François Dengah, H J

    2016-04-01

    Research on the association between religion and health often neglects to provide an explicit theoretical mechanism of influence between faith and well-being. This research posits that dissociative behaviors, such as glossolalia, may provide a biological pathway that influences both physiological and psychological health. This paper argues that religious dissociation acts as a moderator between economic stressors and psychobiological appraisal. Brazil, with its economic inequality and preponderance of religious dissociative rituals, provides an ideal context to examine religious dissociation as a moderator of stress. Utilizing data from a cross section of Brazilian faiths, this paper examines: (1) Whether individuals with low socioeconomic status preferentially participate and experience religious dissociative states and (2) whether dissociative states are correlated with greater psychological appraisal of status.

  20. Sodium Cation Affinities of Commonly Used MALDI Matrices Determined by Guided Ion Beam Tandem Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Chinthaka, S. D. M.; Rodgers, M. T.

    2012-04-01

    The sodium cation affinities of six commonly used MALDI matrices are determined here using guided ion beam tandem mass spectrometry techniques. The collision-induced dissociation behavior of six sodium cationized MALDI matrices, Na+(MALDI), with Xe is studied as a function of kinetic energy. The MALDI matrices examined here include: nicotinic acid, quinoline, 3-aminoquinoline, 4-nitroaniline, picolinic acid, and 3-hydroxypicolinic acid. In all cases, the primary dissociation pathway corresponds to endothermic loss of the intact MALDI matrix. The cross section thresholds are interpreted to yield zero and 298 K Na+-MALDI bond dissociation energies (BDEs), or sodium cation affinities, after accounting for the effects of multiple ion-neutral collisions, the kinetic and internal energy distributions of the reactants, and dissociation lifetimes. Density functional theory calculations at the B3LYP/6-311+G(2d,2p)//B3LYP/6-31G* and MP2(full)/6-311+G(2d,2p)//B3LYP/6-31G* levels of theory are used to characterized the structures and energetics for these systems. The calculated BDEs exhibit very good agreement with the measured values for most systems. The experimental and theoretical Na+-MALDI BDEs determined here are compared with those previously measured by cation transfer equilibrium methods.

  1. Cloning and expression of two distinct high-affinity receptors cross-reacting with acidic and basic fibroblast growth factors.

    PubMed Central

    Dionne, C A; Crumley, G; Bellot, F; Kaplow, J M; Searfoss, G; Ruta, M; Burgess, W H; Jaye, M; Schlessinger, J

    1990-01-01

    The fibroblast growth factor (FGF) family consists of at least seven closely related polypeptide mitogens which exert their activities by binding and activation of specific cell surface receptors. Unanswered questions have been whether there are multiple FGF receptors and what factors determine binding specificity and biological response. We report the complete cDNA cloning of two human genes previously designated flg and bek. These genes encode two similar but distinct cell surface receptors comprised of an extracellular domain with three immunoglobulin-like regions, a single transmembrane domain, and a cytoplasmic portion containing a tyrosine kinase domain with a typical kinase insert. The expression of these two cDNAs in transfected NIH 3T3 cells led to the biosynthesis of proteins of 150 kd and 135 kd for flg and bek, respectively. Direct binding experiments with radiolabeled acidic FGF (aFGF) or basic FGF (bFGF), inhibition of binding with native growth factors, and Scatchard analysis of the binding data indicated that bek and flg bind either aFGF or bFGF with dissociation constants of (2-15) x 10(-11) M. The high affinity binding of two distinct growth factors to each of two different receptors represents a unique double redundancy without precedence among polypeptide growth factor-receptor interactions. Images Fig. 1. Fig. 2. Fig. 3. Fig. 4. PMID:1697263

  2. Antibody affinities and relative titers in polyclonal populations: surface plasmon resonance analysis of anti-DNA antibodies.

    PubMed

    Sem, D S; McNeeley, P A; Linnik, M D

    1999-12-01

    This paper presents the equations and methodology for the measurement and interpretation of apparent dissociation constants for polyclonal populations of antibodies, where antigen is kept trace relative to antibody concentration. Surface plasmon resonance is used to determine K(d)s for the binding of anti-DNA antibodies to trace amounts of DNA antigen on a chip. Since the approach taken relies on equilibrium measurements, kinetic mass transport artifacts are avoided. The apparent K(d) is a weighted average of all the K(d)s for the clonally related subpopulations within the polyclonal pool, where each weighting factor is the relative titer (fractional presence) of the subpopulation. Titration curves appear as if there is one monoclonal population with that titer-weighted-average K(d). Implications of changes in the antibody affinity distribution within the population are discussed. The equations described herein provide a better physical understanding of the apparent K(d) that is obtained when a heterogeneous population of receptors is titrated against a trace ligand.

  3. Human Monoclonal Antibody Fragments Binding to Insulin-like Growth Factors 1 and 2 with Picomolar Affinity

    PubMed Central

    Zhao, Qi; Feng, Yang; Zhu, Zhongyu; Dimitrov, Dimiter S.

    2011-01-01

    The type 1 insulin-like growth factor receptor (IGF1R) and its ligands (IGF1 and IGF2) have been implicated in a variety of physiological processes and in diseases such as cancer. In addition to IGF1R, IGF2 also activates the insulin receptor (IR) isoform A and therefore antibodies against IGF2 can inhibit cell proliferation mediated by the signaling through both IGF1R and IR triggered by IGF2. We identified a new human monoclonal antibody (mAb), m708.2, which bound to IGF1 and IGF2 but not to insulin. m708.2 potently inhibited signal transduction mediated by the interaction of IGF1 or IGF2 with the IGF1R and IGF2 with the IR. It also inhibited the growth of the breast cancer cell line MCF-7. An affinity-matured derivative of m708.2, m708.5, bound to IGF1 with equilibrium dissociation constant, KD = 200 pM and to IGF2 with KD = 60 pM. m708.5 inhibited signal transduction mediated by IGF1 and IGF2 and cancer cell growth more potently than m708.2. These results suggest that m708.5 could have potential as a candidate therapeutic for cancers driven by the IGF1,2 interactions with IGF1R and IR. PMID:21750218

  4. Affine Contractions on the Plane

    ERIC Educational Resources Information Center

    Celik, D.; Ozdemir, Y.; Ureyen, M.

    2007-01-01

    Contractions play a considerable role in the theory of fractals. However, it is not easy to find contractions which are not similitudes. In this study, it is shown by counter examples that an affine transformation of the plane carrying a given triangle onto another triangle may not be a contraction even if it contracts edges, heights or medians.…

  5. Affinity-aware checkpoint restart

    SciTech Connect

    Saini, Ajay; Rezaei, Arash; Mueller, Frank; Hargrove, Paul; Roman, Eric

    2014-12-08

    Current checkpointing techniques employed to overcome faults for HPC applications result in inferior application performance after restart from a checkpoint for a number of applications. This is due to a lack of page and core affinity awareness of the checkpoint/restart (C/R) mechanism, i.e., application tasks originally pinned to cores may be restarted on different cores, and in case of non-uniform memory architectures (NUMA), quite common today, memory pages associated with tasks on a NUMA node may be associated with a different NUMA node after restart. Here, this work contributes a novel design technique for C/R mechanisms to preserve task-to-core maps and NUMA node specific page affinities across restarts. Experimental results with BLCR, a C/R mechanism, enhanced with affinity awareness demonstrate significant performance benefits of 37%-73% for the NAS Parallel Benchmark codes and 6-12% for NAMD with negligible overheads instead of up to nearly four times longer an execution times without affinity-aware restarts on 16 cores.

  6. Affinity-aware checkpoint restart

    DOE PAGES

    Saini, Ajay; Rezaei, Arash; Mueller, Frank; ...

    2014-12-08

    Current checkpointing techniques employed to overcome faults for HPC applications result in inferior application performance after restart from a checkpoint for a number of applications. This is due to a lack of page and core affinity awareness of the checkpoint/restart (C/R) mechanism, i.e., application tasks originally pinned to cores may be restarted on different cores, and in case of non-uniform memory architectures (NUMA), quite common today, memory pages associated with tasks on a NUMA node may be associated with a different NUMA node after restart. Here, this work contributes a novel design technique for C/R mechanisms to preserve task-to-core mapsmore » and NUMA node specific page affinities across restarts. Experimental results with BLCR, a C/R mechanism, enhanced with affinity awareness demonstrate significant performance benefits of 37%-73% for the NAS Parallel Benchmark codes and 6-12% for NAMD with negligible overheads instead of up to nearly four times longer an execution times without affinity-aware restarts on 16 cores.« less

  7. ELECTRON AFFINITIES OF INORGANIC RADICALS.

    DTIC Science & Technology

    energy in the latter compound is 110 kcals/mole, distinctly higher than in ammonia. Cyanogen (CN)2 and hydrocyanic acid (HCN) yield values for the...ions very readily, and the electron affinity is 49 kcals/mole. A comparison with the results from thiocyanic acid (HNCS) indicates that the H-N bond

  8. Dissociation rate of bromine diatomics in an argon heat bath

    NASA Technical Reports Server (NTRS)

    Razner, R.; Hopkins, D.

    1973-01-01

    The evolution of a collection of 300 K bromine diatomics embedded in a heat bath of argon atoms at 1800 K was studied by computer, and a dissociation-rate constant for the reaction Br2 + BR + Ar yields Br + Ar was determined. Previously published probability distributions for energy and angular momentum transfers in classical three-dimensional Br2-Ar collisions were used in conjunction with a newly developed Monte Carlo scheme for this purpose. Results are compared with experimental shock-tube data and the predictions of several other theoretical models. A departure from equilibrium is obtained which is significantly greater than that predicted by any of these other theories.

  9. Dissociative recombination in planetary ionospheres

    NASA Technical Reports Server (NTRS)

    Fox, J. L.

    1993-01-01

    Ionization in planetary atmospheres can be produced by solar photoionization, photoelectron impact ionization, and, in auroral regions, by impact of precipitating particles. This ionization is lost mainly in dissociative recombination (DR) of molecular ions. Although atomic ions cannot undergo DR, they can be transformed locally through ion-molecule reactions into molecular ions, or they may be transported vertically or horizontally to regions of the atmosphere where such transformations are possible. Because DR reactions tend to be very exothermic, they can be an important source of kinetically or internally excited fragments. In interplanetary thermospheres, the neutral densities decrease exponentially with altitude. Below the homopause (or turbopause), the atmosphere is assumed to be throughly mixed by convection and/or turbulence. Above the homopause, diffusion is the major transport mechanism, and each species is distributed according to its mass, with the logarithmic derivative of the density with repect to altitude given approximately by -1/H, where H = kT/mg is the scale height. In this expression, T is the neutral temperature, g is the local acceleratiion of gravity, and m is the mass of the species. Thus lighter species become relatively more abundant, and heavier species less abundant, as the altitude increases. This variation of the neutral composition can lead to changes in the ion composition; furthermore, as the neutral densities decrease, dissociative recombination becomes more important relative to ion-neutral reactions as a loss mechanism for molecular ions.

  10. Dissociative identity disorder: Medicolegal challenges.

    PubMed

    Farrell, Helen M

    2011-01-01

    Persons with dissociative identity disorder (DID) often present in the criminal justice system rather than the mental health system and perplex experts in both professions. DID is a controversial diagnosis with important medicolegal implications. Defendants have claimed that they committed serious crimes, including rape or murder, while they were in a dissociated state. Asserting that their alter personality committed the bad act, defendants have pleaded not guilty by reason of insanity (NGRI). In such instances, forensic experts are asked to assess the defendant for DID and provide testimony in court. Debate continues over whether DID truly exists, whether expert testimony should be allowed into evidence, and whether it should exculpate defendants for their criminal acts. This article reviews historical and theoretical perspectives on DID, presents cases that illustrate the legal implications and controversies of raising an insanity defense based on multiple personalities, and examines the role of forensic experts asked to comment on DID with the goal of assisting clinicians in the medicolegal assessment of DID in relation to crimes.

  11. Dissociation and functional neurologic disorders.

    PubMed

    Brown, R J

    2017-01-01

    Dissociation has been cited as a possible psychologic mechanism underpinning functional neurologic disorders (FND) since the 19th century. Since that time, changes in psychiatric classification have created confusion about what the term dissociation actually means. The available evidence suggests that it now refers to at least two qualitatively distinct types of phenomena: detachment (an altered state of consciousness characterized by a sense of separation from the self or world) and compartmentalization (a reversible loss of voluntary control over apparently intact processes and functions), as well as their underlying mechanisms. This chapter considers some of the problems with conflating these phenomena under a single heading as well as the relationship between detachment, compartmentalization, and FND. It is argued that FNDs are fundamentally compartmentalization disorders, but that detachment is often part of the clinical picture and may contribute to the development and maintenance of functional symptoms in many cases. By this view, understanding compartmentalization requires an appreciation of the mechanisms involved in controlling and accessing mental processes and contents. Two possible mechanisms in this regard are described and the evidence for these is considered, followed by a discussion of clinical and empiric implications.

  12. Dissociative disorders and suicidality in psychiatric outpatients.

    PubMed

    Foote, Brad; Smolin, Yvette; Neft, Deborah I; Lipschitz, Deborah

    2008-01-01

    Although it is common for patients with dissociative disorders to report a history of suicide attempts, there is very little data systematically comparing suicidality in patients with dissociative disorders versus patients without these disorders. The subjects in our study were 231 patients consecutively admitted to an inner-city, hospital-based outpatient psychiatric clinic. Eighty-two of these patients completed structured interviews for dissociative disorders, borderline personality disorder, and trauma history (dissociative disorders interview schedule) and for posttraumatic stress disorder and substance abuse (Structured Clinical Interview for DSM-IV). Patients receiving a dissociative disorder diagnosis were compared with nondissociative patients on measures of self-harm and suicidality. Presence of a dissociative disorder was strongly associated with all measures of self-harm and suicidality. When we focused on patients with a history of multiple suicide attempts, significant associations were found between several diagnoses (dissociative disorder; borderline personality disorder; posttraumatic stress disorder; alcohol abuse/dependence) and multiple suicide attempter status. When these diagnoses were entered in a logistic regression, a highly significant association remained for dissociative diagnosis and multiple suicide attempter status (odds ratio, 15.09; 95% confidence interval, 2.67-85.32; p = 0.002). Dissociative disorders are commonly overlooked in studies of suicidality, but in this population they were the strongest predictor of multiple suicide attempter status.

  13. Dissociative Recombination without a Curve Crossing

    NASA Technical Reports Server (NTRS)

    Guberman, Steven L.

    1994-01-01

    Ab initio calculations show that a curve crossing is not always needed for a high dissociative- recombination cross section. For HeH(+), in which no neutral states cross the ion potential curve, dissociative recombination is driven by the nuclear kinetic-energy operator on adiabatic potential curves. The kinetic-energy derivative operator allows for capture into repulsive curves that are outside of the classical turning points for the nuclear motion. The dominant dissociative route is the C (2)Sigma(+) state leading to H(n = 2) atoms. An analogous mechanism is proposed for the dissociative recombination of H3(+).

  14. Mirror writing and a dissociative identity disorder.

    PubMed

    Le, Catherine; Smith, Joyce; Cohen, Lewis

    2009-01-01

    Individuals with dissociative identity disorder (DID) have been known to show varied skills and talents as they change from one dissociative state to another. For example, case reports have described people who have changed their handedness or have spoken foreign languages during their dissociative states. During an interview with a patient with DID, a surprising talent emerged when she wrote a sentence for the Folstein Mini-Mental State Exam-mirror writing. It is not known whether her mirror writing had a deeper level of meaning; however, it does emphasize the idiosyncratic nature of dissociative identity disorder.

  15. Truncated Amyloid-β(11–40/42) from Alzheimer Disease Binds Cu2+ with a Femtomolar Affinity and Influences Fiber Assembly*

    PubMed Central

    Barritt, Joseph D.; Viles, John H.

    2015-01-01

    Alzheimer disease coincides with the formation of extracellular amyloid plaques composed of the amyloid-β (Aβ) peptide. Aβ is typically 40 residues long (Aβ(1–40)) but can have variable C and N termini. Naturally occurring N-terminally truncated Aβ(11–40/42) is found in the cerebrospinal fluid and has a similar abundance to Aβ(1–42), constituting one-fifth of the plaque load. Based on its specific N-terminal sequence we hypothesized that truncated Aβ(11–40/42) would have an elevated affinity for Cu2+. Various spectroscopic techniques, complemented with transmission electron microscopy, were used to determine the properties of the Cu2+-Aβ(11–40/42) interaction and how Cu2+ influences amyloid fiber formation. We show that Cu2+-Aβ(11–40) forms a tetragonal complex with a 34 ± 5 fm dissociation constant at pH 7.4. This affinity is 3 orders of magnitude tighter than Cu2+ binding to Aβ(1–40/42) and more than an order of magnitude tighter than that of serum albumin, the extracellular Cu2+ transport protein. Furthermore, Aβ(11–40/42) forms fibers twice as fast as Aβ(1–40) with a very different morphology, forming bundles of very short amyloid rods. Substoichiometric Cu2+ drastically perturbs Aβ(11–40/42) assembly, stabilizing much longer fibers. The very tight fm affinity of Cu2+ for Aβ(11–40/42) explains the high levels of Cu2+ observed in Alzheimer disease plaques. PMID:26408196

  16. The Interaction of Arp2/3 Complex with Actin: Nucleation, High Affinity Pointed End Capping, and Formation of Branching Networks of Filaments

    NASA Astrophysics Data System (ADS)

    Dyche Mullins, R.; Heuser, John A.; Pollard, Thomas D.

    1998-05-01

    The Arp2/3 complex is a stable assembly of seven protein subunits including two actin-related proteins (Arp2 and Arp3) and five novel proteins. Previous work showed that this complex binds to the sides of actin filaments and is concentrated at the leading edges of motile cells. Here, we show that Arp2/3 complex purified from Acanthamoeba caps the pointed ends of actin filaments with high affinity. Arp2/3 complex inhibits both monomer addition and dissociation at the pointed ends of actin filaments with apparent nanomolar affinity and increases the critical concentration for polymerization at the pointed end from 0.6 to 1.0 μ M. The high affinity of Arp2/3 complex for pointed ends and its abundance in amoebae suggest that in vivo all actin filament pointed ends are capped by Arp2/3 complex. Arp2/3 complex also nucleates formation of actin filaments that elongate only from their barbed ends. From kinetic analysis, the nucleation mechanism appears to involve stabilization of polymerization intermediates (probably actin dimers). In electron micrographs of quick-frozen, deep-etched samples, we see Arp2/3 bound to sides and pointed ends of actin filaments and examples of Arp2/3 complex attaching pointed ends of filaments to sides of other filaments. In these cases, the angle of attachment is a remarkably constant 70 ± 7 degrees. From these in vitro biochemical properties, we propose a model for how Arp2/3 complex controls the assembly of a branching network of actin filaments at the leading edge of motile cells.

  17. Modeling heating curve for gas hydrate dissociation in porous media.

    PubMed

    Dicharry, Christophe; Gayet, Pascal; Marion, Gérard; Graciaa, Alain; Nesterov, Anatoliy N

    2005-09-15

    A method for modeling the heating curve for gas hydrate dissociation in porous media at isochoric conditions (constant cell volume) is presented. This method consists of using an equation of state of the gas, the cumulative volume distribution (CVD) of the porous medium, and a van der Waals-Platteeuw-type thermodynamic model that includes a capillary term. The proposed method was tested to predict the heating curves for methane hydrate dissociation in a mesoporous silica glass for saturated conditions (liquid volume = pore volume) and for a fractional conversion of water to hydrate of 1 (100% of the available water was converted to hydrate). The shape factor (F) of the hydrate-water interface was found equal to 1, supporting a cylindrical shape for the hydrate particles during hydrate dissociation. Using F = 1, it has been possible to predict the heating curve for different ranges of pressure and temperature. The excellent agreement between the calculated and experimental heating curves supports the validity of our approach.

  18. Redshift in Hubble's constant.

    NASA Astrophysics Data System (ADS)

    Temple-Raston, M.

    1997-01-01

    A topological field theory with Bogomol'nyi solitons is examined. The Bogomol'nyi solitons have much in common with the instanton in Yang-Mills theory; consequently the author called them 'topological instantons'. When periodic boundary conditions are imposed, the field theory comments indirectly on the speed of light within the theory. In this particular model the speed of light is not a universal constant. This may or may not be relevant to the current debate in astronomy and cosmology over the large values of the Hubble constant obtained by the latest generation of ground- and space-based telescopes. An experiment is proposed to detect spatial variation in the speed of light.

  19. Wall of fundamental constants

    SciTech Connect

    Olive, Keith A.; Peloso, Marco; Uzan, Jean-Philippe

    2011-02-15

    We consider the signatures of a domain wall produced in the spontaneous symmetry breaking involving a dilatonlike scalar field coupled to electromagnetism. Domains on either side of the wall exhibit slight differences in their respective values of the fine-structure constant, {alpha}. If such a wall is present within our Hubble volume, absorption spectra at large redshifts may or may not provide a variation in {alpha} relative to the terrestrial value, depending on our relative position with respect to the wall. This wall could resolve the contradiction between claims of a variation of {alpha} based on Keck/Hires data and of the constancy of {alpha} based on Very Large Telescope data. We derive the properties of the wall and the parameters of the underlying microscopic model required to reproduce the possible spatial variation of {alpha}. We discuss the constraints on the existence of the low-energy domain wall and describe its observational implications concerning the variation of the fundamental constants.

  20. Percolation with Constant Freezing

    NASA Astrophysics Data System (ADS)

    Mottram, Edward

    2014-06-01

    We introduce and study a model of percolation with constant freezing ( PCF) where edges open at constant rate , and clusters freeze at rate independently of their size. Our main result is that the infinite volume process can be constructed on any amenable vertex transitive graph. This is in sharp contrast to models of percolation with freezing previously introduced, where the limit is known not to exist. Our interest is in the study of the percolative properties of the final configuration as a function of . We also obtain more precise results in the case of trees. Surprisingly the algebraic exponent for the cluster size depends on the degree, suggesting that there is no lower critical dimension for the model. Moreover, even for , it is shown that finite clusters have algebraic tail decay, which is a signature of self organised criticality. Partial results are obtained on , and many open questions are discussed.

  1. Dissociative symptoms and dissociative disorder comorbidity in patients with obsessive-compulsive disorder.

    PubMed

    Belli, Hasan; Ural, Cenk; Vardar, Melek Kanarya; Yesılyurt, Sema; Oncu, Fatıh

    2012-10-01

    The present study attempted to assess the dissociative symptoms and overall dissociative disorder comorbidity in patients with obsessive-compulsive disorder (OCD). In addition, we examined the relationship between the severity of obsessive-compulsive symptoms and dissociative symptoms. All patients admitted for the first time to the psychiatric outpatient unit were included in the study. Seventy-eight patients had been diagnosed as having OCD during the 2-year study period. Patients had to meet the Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition criteria for OCD. Most (76.9%; n = 60) of the patients were female, and 23.1% (n = 18) of the patients were male. Dissociation Questionnaire was used to measure dissociative symptoms. The Structured Clinical Interview for Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition Dissociative Disorders interviews and Yale-Brown Obsessive Compulsive Checklist and Severity Scale were used. Eleven (14%) of the patients with OCD had comorbid dissociative disorder. The most prevalent disorder in our study was dissociative depersonalization disorder. Dissociative amnesia and dissociative identity disorder were common as well. The mean Yale-Brown score was 23.37 ± 7.27 points. Dissociation Questionnaire scores were between 0.40 and 3.87 points, and the mean was 2.23 ± 0.76 points. There was a statistically significant positive correlation between Yale-Brown points and Dissociation Questionnaire points. We conclude that dissociative symptoms among patients with OCD should alert clinicians for the presence of a chronic and complex dissociative disorder. Clinicians may overlook an underlying dissociative process in patients who have severe symptoms of OCD. However, a lack of adequate response to cognitive-behavioral and drug therapy may be a consequence of dissociative process.

  2. Calcium ion gradients modulate the zinc affinity and antibacterial activity of human calprotectin.

    PubMed

    Brophy, Megan Brunjes; Hayden, Joshua A; Nolan, Elizabeth M

    2012-10-31

    Calprotectin (CP) is an antimicrobial protein produced and released by neutrophils that inhibits the growth of pathogenic microorganisms by sequestering essential metal nutrients in the extracellular space. In this work, spectroscopic and thermodynamic metal-binding studies are presented to delineate the zinc-binding properties of CP. Unique optical absorption and EPR spectroscopic signatures for the interfacial His(3)Asp and His(4) sites of human calprotectin are identified by using Co(II) as a spectroscopic probe. Zinc competition titrations employing chromophoric Zn(II) indicators provide a 2:1 Zn(II):CP stoichiometry, confirm that the His(3)Asp and His(4) sites of CP coordinate Zn(II), and reveal that the Zn(II) affinity of both sites is calcium-dependent. The calcium-insensitive Zn(II) competitor ZP4 affords dissociation constants of K(d1) = 133 ± 58 pM and K(d2) = 185 ± 219 nM for CP in the absence of Ca(II). These values decrease to K(d1) ≤ 10 pM and K(d2) ≤ 240 pM in the presence of excess Ca(II). The K(d1) and K(d2) values are assigned to the His(3)Asp and His(4) sites, respectively. In vitro antibacterial activity assays indicate that the metal-binding sites and Ca(II)-replete conditions are required for CP to inhibit the growth of both Gram-negative and -positive bacteria. Taken together, these data provide a working model whereby calprotectin responds to physiological Ca(II) gradients to become a potent Zn(II) chelator in the extracellular space.

  3. Muscarinic cholinergic receptor binding sites differentiated by their affinity for pirenzepine do not interconvert

    SciTech Connect

    Gil, D.W.; Wolfe, B.B.

    1986-05-01

    Although it has been suggested by many investigators that subtypes of muscarinic cholinergic receptors exist, physical studies of solubilized receptors have indicated that only a single molecular species may exist. To test the hypothesis that the putative muscarinic receptor subtypes in rat forebrain are interconvertible states of the same receptor, the selective antagonist pirenzepine (PZ) was used to protect muscarinic receptors from blockade by the irreversible muscarinic receptor antagonist propylbenzilylcholine mustard (PBCM). If interconversion of high (M1) and low (M2) affinity binding sites for PZ occurs, incubation of cerebral cortical membranes with PBCM in the presence of PZ should not alter the proportions of M1 and M2 binding sites that are unalkylated (i.e., protected). If, on the other hand, the binding sites are not interconvertible, PZ should be able to selectively protect M1 sites and alter the proportions of unalkylated M1 and M2 binding sites. In the absence of PZ, treatment of cerebral cortical membranes with 20 nM PBCM at 4 degrees C for 50 min resulted in a 69% reduction in the density of M1 binding sites and a 55% reduction in the density of M2 binding sites with no change in the equilibrium dissociation constants of the radioligands (/sup 3/H)quinuclidinyl benzilate or (/sup 3/H)PZ. The reasons for this somewhat selective effect of PBCM are not apparent. In radioligand binding experiments using cerebral cortical membranes, PZ inhibited the binding of (/sup 3/H)quinuclidinyl benzilate in a biphasic manner.

  4. Constant-pressure Blowers

    NASA Technical Reports Server (NTRS)

    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.

  5. Universe of constant

    NASA Astrophysics Data System (ADS)

    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

  6. The Hubble Constant.

    PubMed

    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.

  7. Iodination of (Tyr11)somatostatin yields a super high affinity ligand for somatostatin receptors in GH4C1 pituitary cells

    SciTech Connect

    Presky, D.H.; Schonbrunn, A.

    1988-11-01

    GH4C1 cells are a clonal strain of rat pituitary tumor cells which contain high affinity receptors for the inhibitory neuropeptide somatostatin (SRIF). In contrast to other peptides that bind to specific receptors on these cells, receptor-bound (125I-Tyr1)SRIF does not undergo rapid endocytosis. Rather, partial degradation to 125I-tyrosine occurs concomitantly with the dissociation of (125I-Tyr1)SRIF from cell surface receptors. In this study we characterize the binding, biological activity and receptor-mediated degradation of (125I-Tyr11)SRIF, a SRIF analog that is radiolabeled in the center of the molecule. The binding of trace concentrations of (125I-Tyr11)SRIF (less than 50 pM) required 6 hr to reach equilibrium at 37 degrees compared with the 60 min required for (125I-Tyr1)SRIF. Analysis of the kinetics of (125I- Tyr11)SRIF binding showed that the rate constant for association (kon = 1.7 x 10(8) M-8min-1) was similar to that for (125I-Tyr1)SRIF (0.8 x 10(8) M-1min-1). However, the two radioligands exhibited markedly different dissociation kinetics; the koff for (125I-Tyr11)SRIF was 0.002 min-1 compared with the value of 0.02 min-1 for (125I-Tyr1) SRIF. In agreement with its much slower rate of dissociation, (125I-Tyr11)SRIF bound to the SRIF receptor with higher affinity (Kd = 70 pM) than did (125I-Tyr1)SRIF (Kd = 350 pM). However, the apparent ED50 for (I-Tyr11)SRIF to inhibit cAMP accumulation (1.9 +/- 0.4 nM) was greater than the ED50 for SRIF (0.19 +/- 0.04 nM). The low potency of (I-Tyr11)SRIF probably resulted from the fact that subsaturating concentrations of this peptide did not achieve equilibrium binding during the 30-min incubation used to assay biological activity. As previously reported for (125I-Tyr1)SRIF, receptor-bound (125I-Tyr11)SRIF was not internalized and was released from the cells as a mixture of intact (125I-Tyr11)SRIF (30%) and the degradation product 125I-tyrosine (65%).

  8. The multidimensional inventory of dissociation (MID): A comprehensive measure of pathological dissociation.

    PubMed

    Dell, Paul F

    2006-01-01

    This article describes the development and validation of the Multidimensional Inventory of Dissociation (MID). The MID is a 218-item, self-administered, multiscale instrument that comprehensively assesses the phenomenological domain of pathological dissociation and diagnoses the dissociative disorders. The MID measures 14 major facets of pathological dissociation; it has 23 dissociation diagnostic scales that simultaneously operationalize (1) the subjective/ phenomenological domain of pathological dissociation and (2) the hypothesized dissociative symptoms of dissociative identity disorder (Dell, 2001a). The MID was designed for clinical research and for diagnostic assessment of patients who present with a mixture of dissociative, posttraumatic, and borderline symptoms. The MID demonstrated internal reliability, temporal stability, convergent validity, discriminant validity, and construct validity. The MID also exhibited incremental validity over the Dissociative Experiences Scale (DES) by predicting an additional 18% of the variance in weighted abuse scores on the Traumatic Experiences Questionnaire (TEQ). Confirmatory factor analysis (CFA) did not support a one-factor model of the MID's clinical scales (i.e., the 14 facets and the 23 diagnostic symptoms). In contrast, however, CFA of the MID's factor scales (Dell & Lawson, 2005) has strongly supported a one-factor model. It was concluded that both the MID's 168 dissociation items and the construct of pathological dissociation have a second-order, unifactorial structure.

  9. Molecular basis for the dissociation dynamics of protein A-immunoglobulin G1 complex.

    PubMed

    Liu, Fu-Feng; Huang, Bo; Dong, Xiao-Yan; Sun, Yan

    2013-01-01

    Staphylococcus aureus protein A (SpA) is the most popular affinity ligand for immunoglobulin G1 (IgG1). However, the molecular basis for the dissociation dynamics of SpA-IgG1 complex is unclear. Herein, coarse-grained (CG) molecular dynamics (MD) simulations with the Martini force field were used to study the dissociation dynamics of the complex. The CG-MD simulations were first verified by the agreement in the structural and interactional properties of SpA and human IgG1 (hIgG1) in the association process between the CG-MD and all-atom MD at different NaCl concentrations. Then, the CG-MD simulation studies focused on the molecular insight into the dissociation dynamics of SpA-hIgG1 complex at pH 3.0. It is found that there are four steps in the dissociation process of the complex. First, there is a slight conformational adjustment of helix II in SpA. This is followed by the phenomena that the electrostatic interactions provided by the three hot spots (Glu143, Arg146 and Lys154) of helix II of SpA break up, leading to the dissociation of helix II from the binding site of hIgG1. Subsequently, breakup of the hydrophobic interactions between helix I (Phe132, Tyr133 and His137) in SpA and hIgG1 occurs, resulting in the disengagement of helix I from its binding site of hIgG1. Finally, the non-specific interactions between SpA and hIgG1 decrease slowly till disappearance, leading to the complete dissociation of the SpA-hIgG1 complex. This work has revealed that CG-MD coupled with the Martini force field is an effective method for studying the dissociation dynamics of protein-protein complex.

  10. A Redetermination of the Dissociation Energy of MgO(+)

    NASA Technical Reports Server (NTRS)

    Bauschlicher, Charles W., Jr.; Langhoff, Stephen R.; Partridge, Harry

    1994-01-01

    In 1986, we reported a dissociation energy (D(sub 0) of 2.31 eV for the X(sup 2)Pi ground state of MgO(+). This value was determined by computing the dissociation energy to the Mg(2+) + O(-) limit and adjusting the value to the Mg(+) + O limit using the experimental Ionization Potential (IP) of Mg(+) and the Electron Affinity (EA) of O. The success of this method relies on the assumption that there is little covalent contribution to the bonding. The very small (0.04 eV) correlation contribution to the binding energy was taken as corroboration for the validity of this approach. Our earlier theoretical value was estimated to be accurate to at least 0.2 eV. It is in excellent agreement with the subsequent value of 2.30 +/- 0.13 eV determined by Freiser and co-workers from photodissociation experiments. It is also consistent with the upper (less than 3.1 eV) and lower (greater than 1.1 eV) bounds determined by Rowe obtained by studying the reactions of Mg(+) with 03 and NO2. However, it is inconsistent with an upper bound of 1.7 eV reported by Kappes and Staley based on their failure to observe MgO(+) in the reaction of Mg(+) with N2O. The picture became somewhat clouded, however, by the recent guided-ion beam mass spectrometric studies of Dalleska and Armentrout. Their initial analysis of the reaction data for Mg(+) + O2 lead to a bond dissociation energy of 2.92 +/- 0.25 eV, which is considerably larger than the value of 2.47 +/- 0.06 eV deduced from their studies of the Mg(+)+NO2 reaction.

  11. Dissociative attachment reactions of electrons with strong acid molecules

    SciTech Connect

    Adams, N.G.; Smith, D.; Viggiano, A.A.; Paulson, J.F.; Henchman, M.J.

    1986-06-15

    Using the flowing afterglow/Langmuir probe (FALP) technique, we have determined (at variously 300 and 570 K) the dissociative attachment coefficients ..beta.. for the reactions of electrons with the common acids HNO/sub 3/ (producing NO/sup -//sub 2/) and H/sub 2/SO/sub 4/ (HSO/sup -//sub 4/), the superacids FSO/sub 3/H (FSO/sup -//sub 3/), CF/sub 3/SO/sub 3/H (CF/sub 3/SO/sup -//sub 3/), ClSO/sub 3/H (ClSO/sup -//sub 3/,Cl/sup -/), the acid anhydride (CF/sub 3/SO/sub 2/)/sub 2/O (CF/sub 3/SO/sup -//sub 3/), and the halogen halides HBr (Br/sup -/) and HI (I/sup -/). The anions formed in the reactions are those given in the parentheses. The reactions with HF and HCl were investigated, but did not occur at a measurable rate since they are very endothermic. Dissociative attachment is rapid for the common acids, the superacids, and the anhydride, the measured ..beta.. being appreciable fractions of the theoretical maximum ..beta.. for such reactions, ..beta../sub max/. The HI reaction is very fast ( ..beta..approx...beta../sub max/) but the HBr reaction occurs much more slowly because it is significantly endothermic. The data indicate that the extreme acidity of the (Bronsted-type) superacids has its equivalence in the very efficient gas-phase dissociative attachment which these species undergo when reacting with free electrons. The anions of the superacids generated in these reactions, notably FSO/sup -//sub 3/ and CF/sub 3/SO/sup -//sub 3/, are very stable (unreactive) implying exceptionally large electron affinities for the FSO/sub 3/ and CF/sub 3/SO/sub 3/ radicals.

  12. Collision-induced thermochemistry of reactions of dissociation of glycyl-homopeptides-An experimental and theoretical analysis.

    PubMed

    Ivanova, Bojidarka; Spiteller, Michael

    2017-02-01

    The research draws on experimental and theoretical data about energetics and kinetics of mass spectrometric (MS) reactions of glycyl homopenta- (G5) and glycyl homohexapeptides (G6). It shows the great applicability of the methods of quantum chemistry to predict MS profile of peptides using energetics of collision induced dissociation (CID) fragment species. Mass spectrometry is among irreplaceable methods, providing unambiguous qualitative, quantitative and structural information about analytes, applicable to many scientific areas like environmental chemistry; food chemistry; medicinal chemistry; and more. Our study could be considered of substantial interdisciplinary significance, where MS proteomics is widely used. The experimental design involves electrospray ionization (ESI) and CID MS/MS. Theoretical design is based on ab initio and density functional theory (DFT) methods. Experimental MS and theoretical free Gibbs energies as well as rate constants of fragment reactions are compared. The thermodynamic encompasses gas-phase and polar continuum analysis, including polar protic and aprotic solvents within temperature T = 10-500 K; dielectric constant ε = 0-78, pH, and ionic strengths μ = 0.001-1.0 mol dm(-1) . There are computed and discussed 39 protonated forms of peptides at amide N- and -(NHC)=O centers; corresponding fragment ions studying their thermodynamic stability depending on experimental conditions. A correlation analysis between molecular conformations of parent ions and fragment species; their proton accepting ability and internal energy distribution is carried out. Data about ionization potentials (IPs) and electron affinities (EAs) are discussed, as well.

  13. Theoretical study of the unimolecular dissociation of HCO

    NASA Astrophysics Data System (ADS)

    Whittier, Gregory Scott

    This thesis offers a detailed theoretical study of the unimolecular dissociation of formyl radical, HCO, which is an important intermediate in combustion chemistry. A quantum mechanical treatment of the dissociation of isolated HCO is presented along with a mixed quantum/classical study of the excitation and deexcitation of HCO in collisions of HCO with the bath gas Ar. The results are then used to model the kinetics of the collision-induced dissociation of HCO by Ar. Resonance states of HCO are calculated for total angular momentum J = 0, 1, and 3 using the artificial boundary inhomogeneity (ABI) method of Jang and Light [J. Chem. Phys. 102, 3262 (1995)]. Resonance energies and widths are determined by analyzing the Smith lifetime matrix. A resonance search algorithm and a method for resolving overlapping resonances are described. The accurate prediction of J = 3 resonances from J = 0 and 1 data is tested with good results for excited stretch resonance and less accurate results for bending resonances, demonstrating the degree of separability of vibration from overall rotation for these quasi-bound states. A quantum/classical time-dependent self-consistent field (Q/C TDSCF) approach is used to simulate the dynamics of collisions of Ar with HCO. State-to-state cross sections and thermal rate constants for vibrational transitions are presented. Using this model together with assumptions about the rotational energy transfer and a master equation treatment of the kinetics, the low-pressure thermal rate of collision-induced dissociation was calculated over the 300-4000 K temperature range. Comparison with experiment shows good agreement at high temperatures and poor agreement at low temperatures. The high temperature results were sufficient to obtain an Arrhenius expression for the rate that agrees with all experimental results of which we are aware.

  14. Vibrational photodetachment spectroscopy near the electron affinity of S2

    NASA Astrophysics Data System (ADS)

    Barrick, J. B.; Yukich, J. N.

    2016-02-01

    We have conducted laser photodetachment spectroscopy near the detachment threshold of the electron affinity of S2 in a 1.8-T field. The ions are prepared by dissociative electron attachment to carbonyl sulfide. The experiment is conducted in a Penning ion trap and with a narrow-band, tunable, Ti:sapphire laser. A hybrid model for photodetachment in an ion trap is fit to the data using the appropriate Franck-Condon factors. The observations reveal detachment from and to the first few vibrational levels of the anion and the neutral molecule, respectively. Evaporative cooling of the anion ensemble condenses the thermal distribution to the lowest initial vibrational states. The subsequent detachment spectroscopy yields results consistent with a vibrationally cooled anion population.

  15. The Dissociative Recombination of OH(+)

    NASA Technical Reports Server (NTRS)

    Guberman, Steven L.

    1995-01-01

    Theoretical quantum chemical calculations of the cross sections and rates for the dissociative recombination of the upsilon = 0 level of the ground state of OH(+) show that recombination occurs primarily along the 2 (2)Pi diabatic route. The products are 0((1)D) and a hot H atom with 6.1 eV kinetic energy. The coupling to the resonances is very small and the indirect recombination mechanism plays only a minor role. The recommended value for the rate coefficient is (6.3 +/- 0.7) x 10(exp -9)x (T(e)/1300)(exp -0.48) cu.cm/s for 10 less than T(e) less than 1000 K.

  16. Dissociating conflict adaptation from feature integration: a multiple regression approach.

    PubMed

    Notebaert, Wim; Verguts, Tom

    2007-10-01

    Congruency effects are typically smaller after incongruent than after congruent trials. One explanation is in terms of higher levels of cognitive control after detection of conflict (conflict adaptation; e.g., M. M. Botvinick, T. S. Braver, D. M. Barch, C. S. Carter, & J. D. Cohen, 2001). An alternative explanation for these results is based on feature repetition and/or integration effects (e.g., B. Hommel, R. W. Proctor, & K.-P. Vu, 2004; U. Mayr, E. Awh, & P. Laurey, 2003). Previous attempts to dissociate feature integration from conflict adaptation focused on a particular subset of the data in which feature transitions were held constant (J. G. Kerns et al., 2004) or in which congruency transitions were held constant (C. Akcay & E. Hazeltine, in press), but this has a number of disadvantages. In this article, the authors present a multiple regression solution for this problem and discuss its possibilities and pitfalls.

  17. Frequency-constant Q, unity and disorder

    SciTech Connect

    Hargreaves, N.D.

    1995-12-31

    In exploration geophysics we obtain information about the earth by observing its response to different types of applied force. The response can cover the full range of possible Q values (where Q, the quality factor, is a measure of energy dissipation), from close to infinity in the case of deep crustal seismic to close to 0 in the case of many electromagnetic methods. When Q is frequency-constant, however, the various types of response have a common scaling behavior and can be described as being self-affine. The wave-equation then takes on a generalised form, changing from the standard wave-equation at Q = {infinity} to the diffusion equation at Q = 0, via lossy, diffusive, propagation at intermediate Q values. Solutions of this wave-diffusion equation at any particular Q value can be converted to an equivalent set of results for any other Q value. In particular it is possible to convert from diffusive to wave propagation by a mapping from Q < {infinity} to Q = {infinity}. In the context of seismic sounding this is equivalent to applying inverse Q-filtering; in a more general context the mapping integrates different geophysical observations by referencing them to the common result at Q = {infinity}. The self-affinity of the observations for frequency-constant Q is an expression of scale invariance in the fundamental physical properties of the medium of propagation, this being the case whether the mechanism of diffusive propagation is scattering of intrinsic attenuation. Scale invariance, or fractal scaling, is a general property of disordered systems; the assumption of frequency-constant Q not only implies a unity between different geophysical observations, but also suggests that it is the disordered nature of the earth`s sub-surface that is the unifying factor.

  18. Dissociation behavior of methane--ethane mixed gas hydrate coexisting structures I and II.

    PubMed

    Kida, Masato; Jin, Yusuke; Takahashi, Nobuo; Nagao, Jiro; Narita, Hideo

    2010-09-09

    Dissociation behavior of methane-ethane mixed gas hydrate coexisting structures I and II at constant temperatures less than 223 K was studied with use of powder X-ray diffraction and solid-state (13)C NMR techniques. The diffraction patterns at temperatures less than 203 K showed both structures I and II simultaneously convert to Ih during the dissociation, but the diffraction pattern at temperatures greater than 208 K showed different dissociation behavior between structures I and II. Although the diffraction peaks from structure II decreased during measurement at constant temperatures greater than 208 K, those from structure I increased at the initial step of dissociation and then disappeared. This anomalous behavior of the methane-ethane mixed gas hydrate coexisting structures I and II was examined by using the (13)C NMR technique. The (13)C NMR spectra revealed that the anomalous behavior results from the formation of ethane-rich structure I. The structure I hydrate formation was associated with the dissociation rate of the initial methane-ethane mixed gas hydrate.

  19. [Dissociative disorders: neurologists and psychiatrists working together].

    PubMed

    Hubschmid, Monica; Aybek, Selma; Vingerhoets, François; Berney, Alexandre

    2008-02-13

    Dissociative disorders often have an atypical neurological presentation requiring a complex diagnostic process at the interface between neurology and psychiatry. A strong interdisciplinary collaboration is needed for diagnosis restitution and patient treatment. Current knowledge is still scarce but recent studies in clinical neuroscience enrich this field. This article presents the main aspects of dissociative disorders and suggests a treatment framework.

  20. Dissociations, Developmental Psychology, and Pedagogical Design

    ERIC Educational Resources Information Center

    Lillard, Angeline

    2006-01-01

    Although dissociations in children's responses are sometimes about "getting it right" for an experimenter, they might also often reflect differences between conscious and subconscious processing that are not geared to correct performance. Research with adults also reveals many cases of dissociation, and adults can more easily be subjected to…

  1. Verbal-Behavioral Dissociations in Development

    ERIC Educational Resources Information Center

    Woolley, Jacqueline D.

    2006-01-01

    Verbal and behavioral measures of children's knowledge are frequently dissociated. These situations represent a largely untapped but important resource for furthering an understanding of human cognition. In this paper, verbal-behavioral dissociations in children are discussed and analyzed, drawing from a wide range of domains. The article explores…

  2. Dissociative depression among women in the community.

    PubMed

    Sar, Vedat; Akyüz, Gamze; Oztürk, Erdinç; Alioğlu, Firdevs

    2013-01-01

    This study screened the prevalence and correlates of dissociative disorders among depressive women in the general population. The Dissociative Disorders Interview Schedule and the posttraumatic stress disorder (PTSD) and borderline personality disorder sections of the Structured Clinical Interview for DSM-IV were administered to 628 women in 500 homes. The prevalence of current major depressive episode was 10.0%. Of the women, 26 (40.6%) had the lifetime diagnosis of a DSM-IV, dissociative disorder, yielding a prevalence of 4.1% for dissociative depression. This group was younger (mean age = 30.7 years) than the nondissociative depression women (mean age = 39.6 years). There was no difference between the 2 groups on comorbid somatization disorder, PTSD, or borderline personality disorder. Besides suicide attempts, the dissociative group was characterized by secondary features of dissociative identity disorder; Schneiderian symptoms; borderline personality disorder criteria; and extrasensory perceptions, including possession experiences. They reported suicidality, thoughts of guilt and worthlessness, diminished concentration and indecisiveness, and appetite and weight changes more frequently than the nondissociative group. Early cessation of school education and childhood sexual abuse were frequently reported by the dissociative depression group. With its distinct features, the concept of dissociative depression may facilitate understanding of treatment resistance in, development of better psychotherapy strategies for, and new thinking on the neurobiology and pharmacotherapy of depressive disorders.

  3. [Dissociative disorder and self-injury].

    PubMed

    Noma, Shun'ichi

    2011-01-01

    Both the number of patients with dissociative disorder and that of those with self-injury have been increasing since the end of the twentieth century, suggesting that dissociation and self-injury might be closely related. When dissociative disorder coexists with self-injury, it implies self-punishment and a wish to be understood by others. Although many cases of self-injury observed since 2000 lacked traumatic experiences and were not accompanied by pathological dissociative symptoms, the patients did have dissociative tendencies. According to the results of our study examining self-injury in patients with eating disorders, we observed that self-injury, dissociative tendency and insulation from others are related to each other. This suggests that affects, sensations and representations are dissociated, losing their normal response order, and that the pervasive idea that "pain=secure" is formed in a patient from childhood based on influence from their parents. Self-injury appears to be an activation of this pervasive idea that is triggered by a stressful situation, when the dissociative psychological segmentation of effects and their representations are present in the background.

  4. A Hierarchical Process-Dissociation Model

    ERIC Educational Resources Information Center

    Rouder, Jeffrey N.; Lu, Jun; Morey, Richard D.; Sun, Dongchu; Speckman, Paul L.

    2008-01-01

    In fitting the process-dissociation model (L. L. Jacoby, 1991) to observed data, researchers aggregate outcomes across participant, items, or both. T. Curran and D. L. Hintzman (1995) demonstrated how biases from aggregation may lead to artifactual support for the model. The authors develop a hierarchical process-dissociation model that does not…

  5. Dissociation in schizophrenia and borderline personality disorder

    PubMed Central

    Pec, Ondrej; Bob, Petr; Raboch, Jiri

    2014-01-01

    Background Dissociation likely plays a key role in schizophrenia and borderline personality disorder (BPD), although empirical studies that compare specific manifestations of these symptoms in schizophrenia and BPD are rare. In this context, the purpose of this study was to compare the occurrence of dissociative and other psychopathological symptoms in these disorders, and to assess the possible influence of antipsychotic medication on the dissociative symptoms. Methods We assessed 31 patients with schizophrenia and 36 patients with BPD. Dissociative symptoms were measured by the Dissociative Experiences Scale (DES), symptoms related to stress and traumatic experiences were assessed using the Trauma Symptom Checklist-40 (TSC-40), and other psychopathological symptoms were measured with the Health of the Nation Outcome Scales (HoNOS). We also assessed actual daily doses of antipsychotic medication in chlorpromazine equivalents in all participants. Results The results show that symptoms of traumatic stress measured by the TSC-40 had significantly higher scores in the BPD group. The data also show that dissociative symptoms (DES) were significantly correlated with symptoms of traumatic stress (TSC-40) and with symptoms assessed by the HoNOS. Remarkably significant correlations were found between levels of antipsychotic medication and the DES and between antipsychotic medication and the depersonalization/derealization component of the DES in BPD patients. Conclusion The results support an important role of dissociative processes in schizophrenia and BPD and suggest a significant relationship between manifestations of dissociative symptoms in BPD and antipsychotic medication. PMID:24672239

  6. Dissociative recombination of highly symmetric polyatomic ions.

    PubMed

    Douguet, Nicolas; Orel, Ann E; Greene, Chris H; Kokoouline, Viatcheslav

    2012-01-13

    A general first-principles theory of dissociative recombination is developed for highly symmetric molecular ions and applied to H(3)O(+) and CH(3)(+), which play an important role in astrophysical, combustion, and laboratory plasma environments. The theoretical cross sections obtained for the dissociative recombination of the two ions are in good agreement with existing experimental data from storage ring experiments.

  7. Dissociation and the Development of Psychopathology.

    ERIC Educational Resources Information Center

    Putnam, Frank W.; Trickett, Penelope K.

    This paper reviews the research on dissociation and the development of psychopathology in children and adolescents. Definitions and dimensions of dissociation are addressed, noting its range from normative daydreaming to the extremes found in individuals with multiple personality disorder. Memory dysfunctions, disturbances of identity, passive…

  8. Dissociation heat transfer characteristics of methane hydrates

    SciTech Connect

    Kamath, V.A.; Holder, G.D.

    1987-02-01

    Knowledge of the interfacial heat transfer phenomenon during the dissociation of gas hydrates is essential in modeling the hydrate dissociation process. Such knowledge has applications in natural gas processing, storage, or transportation; in the drilling and recovery of oil and gas in the presence of gas hydrates; in the desalination of sea water; and in the production of natural gas from hydrate reservoirs. The process of hydrate dissociation is a unique phenomenon in which gas and water are simultaneously produced at the dissociated hydrate surface and play an important role in the mechanism of heat transfer to hydrates. An earlier study of propane hydrate dissociation showed that hydrate dissociation is a heat-transfer-limited process and somewhat similar to the nucleate boiling of liquids. In the present study, heat transfer limitations for methane hydrate dissociation were studied for two reasons. First, a comparison of the results of this study with propane hydrate was desired. Second, the effect of hydrate structure and gas molecule type on the rate of heat transfer during hydrate dissociation was sought.

  9. The Hubble Constant

    NASA Astrophysics Data System (ADS)

    Freedman, Wendy; Madore, Barry; Mager, Violet; Persson, Eric; Rigby, Jane; Sturch, Laura

    2008-12-01

    We present a plan to measure a value of the Hubble constant having a final systematic uncertainty of only 3% by taking advantage of Spitzer's unique mid-infrared capabilities. This involves using IRAC to undertake a fundamental recalibration of the Cepheid distance scale and progressively moving it out to pure Hubble flow by an application of a revised mid-IR Tully-Fisher relation. The calibration and application, in one coherent and self-consistent program, will go continuously from distances of parsecs to several hundred megaparsecs. It will provide a first-ever mid-IR calibration of Cepheids in the Milky Way, LMC and Key Project spiral galaxies and a first-ever measurement and calibration of the TF relation at mid-infrared wavelengths, and finally a calibration of Type Ia SNe. Most importantly this program will be undertaken with a single instrument, on a single telescope, working exclusively at mid-infrared wavelengths that are far removed from the obscuring effects of dust extinction. Using Spitzer in this focused way will effectively eliminate all of the major systematics in the Cepheid and TF distance scales that have been the limiting factors in all previous applications, including the HST Key Project. By executing this program, based exclusively on Spitzer data, we will deliver a value of the Hubble constant, having a statistical precision better than 11%, with all currently known systematics quantified and constrained to a level of less than 3%. A value of Ho determined to this level of systematic accuracy is required for up-coming cosmology experiments, including Planck. A more accurate value of the Hubble constant will directly result in other contingently measured cosmological parameters (e.g., Omega_m, Omega_L, & w) having their covariant uncertainties reduced significantly now. Any further improvements using this route will have to await JWST, for which this study is designed to provide a lasting and solid foundation, and ultimately a value of Ho

  10. The Dynamics Behind the Affinity: Controlling Heme-Gas Affinity via Geminate Recombination and Heme Propionate Conformation in the NO Carrier Cytochrome c'.

    PubMed

    Andrew, Colin R; Petrova, Olga N; Lamarre, Isabelle; Lambry, Jean-Christophe; Rappaport, Fabrice; Negrerie, Michel

    2016-11-18

    Nitric oxide (NO) sensors are heme proteins which may also bind CO and O2. Control of heme-gas affinity and their discrimination are achieved by the structural properties and reactivity of the heme and its distal and proximal environments, leading to several energy barriers. In the bacterial NO sensor cytochrome c' from Alcaligenes xylosoxidans (AXCP), the single Leu16Ala distal mutation boosts the affinity for gas ligands by a remarkable 10(6)-10(8)-fold, transforming AXCP from one of the lowest affinity gas binding proteins to one of the highest. Here, we report the dynamics of diatomics after photodissociation from wild type and L16A-AXCP over 12 orders of magnitude in time. For the L16A variant, the picosecond geminate rebinding of both CO and NO appears with an unprecedented 100% yield, and no exit of these ligands from protein to solvent could be observed. Molecular dynamic simulations saliently demonstrate that dissociated CO stays within 4 Å from Fe(2+), in contrast to wild-type AXCP. The L16A mutation confers a heme propionate conformation and docking site which traps the diatomics, maximizing the probability of recombination and directly explaining the ultrahigh affinities for CO, NO, and O2. Overall, our results point to a novel mechanism for modulating heme-gas affinities in proteins.

  11. Change is a Constant.

    PubMed

    Lubowitz, James H; Provencher, Matthew T; Brand, Jefferson C; Rossi, Michael J; Poehling, Gary G

    2015-06-01

    In 2015, Henry P. Hackett, Managing Editor, Arthroscopy, retires, and Edward A. Goss, Executive Director, Arthroscopy Association of North America (AANA), retires. Association is a positive constant, in a time of change. With change comes a need for continuing education, research, and sharing of ideas. While the quality of education at AANA and ISAKOS is superior and most relevant, the unique reason to travel and meet is the opportunity to interact with innovative colleagues. Personal interaction best stimulates new ideas to improve patient care, research, and teaching. Through our network, we best create innovation.

  12. Cosmology with varying constants.

    PubMed

    Martins, Carlos J A P

    2002-12-15

    The idea of possible time or space variations of the 'fundamental' constants of nature, although not new, is only now beginning to be actively considered by large numbers of researchers in the particle physics, cosmology and astrophysics communities. This revival is mostly due to the claims of possible detection of such variations, in various different contexts and by several groups. I present the current theoretical motivations and expectations for such variations, review the current observational status and discuss the impact of a possible confirmation of these results in our views of cosmology and physics as a whole.

  13. A classical trajectory study of the dissociation and isomerization of C2H5.

    PubMed

    Wagner, Albert F; Rivera-Rivera, Luis A; Bachellerie, Damien; Perry, Jamin W; Thompson, Donald L

    2013-11-21

    Motivated by photodissociation experiments in which non-RRKM nanosecond lifetimes of the ethyl radical were reported, we have performed a classical trajectory study of the dissociation and isomerization of C2H5 over the energy range 100-150 kcal/mol. We used a customized version of the AIREBO semiempirical potential (Stuart, S. J.; et al. J. Chem. Phys. 2000, 112, 6472-6486) to more accurately describe the gas-phase decomposition of C2H5. This study constitutes one of the first gas-phase applications of this potential form. At each energy, 10,000 trajectories were run and all underwent dissociation in less than 100 ps. The calculated dissociation rate constants are consistent with RRKM models; no evidence was found for nanosecond lifetimes. An analytic kinetics model of isomerization/dissociation competition was developed that incorporated incomplete mode mixing through a postulated divided phase space. The fits of the model to the trajectory data are good and represent the trajectory results in detail through repeated isomerizations at all energies. The model correctly displays single exponential decay at lower energies, but at higher energies, multiexponential decay due to incomplete mode mixing becomes more apparent. At both ends of the energy range, we carried out similar trajectory studies on CD2CH3 to examine isotopic scrambling. The results largely support the assumption that a H or a D atom is equally likely to dissociate from the mixed-isotope methyl end of the molecule. The calculated fraction of products that have the D atom dissociation is ∼20%, twice the experimental value available at one energy within our range. The calculated degree of isotopic scrambling is non-monotonic with respect to energy due to a non-monotonic ratio of the isomerization to dissociation rate constants.

  14. High fidelity modeling of thermal relaxation and dissociation of oxygen

    NASA Astrophysics Data System (ADS)

    Andrienko, Daniil A.; Boyd, Iain D.

    2015-11-01

    A master equation study of vibrational relaxation and dissociation of oxygen is conducted using state-specific O2-O transition rates, generated by extensive trajectory simulations. Both O2-O and O2-O2 collisions are concurrently simulated in the evolving nonequilibrium gas system under constant heat bath conditions. The forced harmonic oscillator model is incorporated to simulate the state-to-state relaxation of oxygen in O2-O2 collisions. The system of master equations is solved to simulate heating and cooling flows. The present study demonstrates the importance of atom-diatom collisions due to the extremely efficient energy randomization in the intermediate O3 complex. It is shown that the presence of atomic oxygen has a significant impact on vibrational relaxation time at temperatures observed in hypersonic flow. The population of highly-excited O2 vibrational states is affected by the amount of atomic oxygen when modeling the relaxation under constant heat bath conditions. A model of coupled state-to-state vibrational relaxation and dissociation of oxygen is also discussed.

  15. High fidelity modeling of thermal relaxation and dissociation of oxygen

    SciTech Connect

    Andrienko, Daniil A. Boyd, Iain D.

    2015-11-15

    A master equation study of vibrational relaxation and dissociation of oxygen is conducted using state-specific O{sub 2}–O transition rates, generated by extensive trajectory simulations. Both O{sub 2}–O and O{sub 2}–O{sub 2} collisions are concurrently simulated in the evolving nonequilibrium gas system under constant heat bath conditions. The forced harmonic oscillator model is incorporated to simulate the state-to-state relaxation of oxygen in O{sub 2}–O{sub 2} collisions. The system of master equations is solved to simulate heating and cooling flows. The present study demonstrates the importance of atom-diatom collisions due to the extremely efficient energy randomization in the intermediate O{sub 3} complex. It is shown that the presence of atomic oxygen has a significant impact on vibrational relaxation time at temperatures observed in hypersonic flow. The population of highly-excited O{sub 2} vibrational states is affected by the amount of atomic oxygen when modeling the relaxation under constant heat bath conditions. A model of coupled state-to-state vibrational relaxation and dissociation of oxygen is also discussed.

  16. The Shutdown Dissociation Scale (Shut-D)

    PubMed Central

    Schalinski, Inga; Schauer, Maggie; Elbert, Thomas

    2015-01-01

    The evolutionary model of the defense cascade by Schauer and Elbert (2010) provides a theoretical frame for a short interview to assess problems underlying and leading to the dissociative subtype of posttraumatic stress disorder. Based on known characteristics of the defense stages “fright,” “flag,” and “faint,” we designed a structured interview to assess the vulnerability for the respective types of dissociation. Most of the scales that assess dissociative phenomena are designed as self-report questionnaires. Their items are usually selected based on more heuristic considerations rather than a theoretical model and thus include anything from minor dissociative experiences to major pathological dissociation. The shutdown dissociation scale (Shut-D) was applied in several studies in patients with a history of multiple traumatic events and different disorders that have been shown previously to be prone to symptoms of dissociation. The goal of the present investigation was to obtain psychometric characteristics of the Shut-D (including factor structure, internal consistency, retest reliability, predictive, convergent and criterion-related concurrent validity). A total population of 225 patients and 68 healthy controls were accessed. Shut-D appears to have sufficient internal reliability, excellent retest reliability, high convergent validity, and satisfactory predictive validity, while the summed score of the scale reliably separates patients with exposure to trauma (in different diagnostic groups) from healthy controls. The Shut-D is a brief structured interview for assessing the vulnerability to dissociate as a consequence of exposure to traumatic stressors. The scale demonstrates high-quality psychometric properties and may be useful for researchers and clinicians in assessing shutdown dissociation as well as in predicting the risk of dissociative responding. PMID:25976478

  17. Dynamics of Na(+)(Benzene) + Benzene Association and Ensuing Na(+)(Benzene)2* Dissociation.

    PubMed

    Paul, Amit K; Kolakkandy, Sujitha; Hase, William L

    2015-07-16

    Chemical dynamics simulations were used to study Bz + Na(+)(Bz) → Na(+)(Bz)2* association and the ensuing dissociation of the Na(+)(Bz)2* cluster (Bz = benzene). An interesting and unexpected reaction found from the simulations is direct displacement, for which the colliding Bz molecule displaces the Bz molecule attached to Na(+), forming Na(+)(Bz). The rate constant for Bz + Na(+)(Bz) association was calculated at 750 and 1000 K, and found to decrease with increase in temperature. By contrast, the direct displacement rate constant increases with temperature. The cross section and rate constant for direct displacement are approximately an order of magnitude lower than those for association. The Na(+)(Bz)2* cluster, formed by association, dissociates with a biexponential probability, with the rate constant for the short-time component approximately an order of magnitude larger than that for the longer time component. The latter rate constant agrees with that of Rice-Ramsperger-Kassel-Marcus (RRKM) theory, consistent with rapid intramolecular vibrational energy redistribution (IVR) and intrinsic RRKM dynamics for the Na(+)(Bz)2* cluster. A coupled phase space model was used to analyze the biexponential dissociation probability.

  18. Dissociative electron attachment to water

    NASA Astrophysics Data System (ADS)

    Haxton, Daniel James

    Dissociative electron attachment to water, H2O+e-→ H+OH-3.27eV H2+O-3.56eV H-+OHX P2 4.35eVH+H+O-8.04e VH-+OH* S2 8.38eVH-+H+O 8.75eV 0.1 is a physical process that has been studied since 1930[1]. It may be viewed as an inherently is non-Born-Oppenheimer process, for the initial state belongs to the electronic continuum, and the final state is electronically bound. As such, it presents a particular challenge for theory. We present a first-principles theoretical treatment of this process, in which we calculate the cross sections for production of the three major atom - diatom products observed by experiment, H2 + O-, H- + OH (X 2pi), and H- + OH (2Sigma). These states comprise the bulk of the experimentally determined cross section. In the present work we employ a Born-Oppenheimer expansion of the wavefunction for nuclear and electronic motion, and treat both quantum mechanically. The adiabatic treatment of the electronic motion incorporates multiconfiguration, correlated wavefunctions and includes an explicit treatment of the electronic continuum. For the nuclear dynamics we incorporate the full dimensionality of nuclear motion, including the three internal degrees of freedom and the effect of rotation. This is the first ab initio treatment of dissociative electron attachment which incorporates more than one degree of freedom in the nuclear dynamics. The first step in our Born-Oppenheimer treatment is to calculate adiabatic potential energy surfaces for the three metastable electronic states which are involved, the 2B1, 2A1, and 2B 2 electronic Feshbach resonances. Global potential energy surfaces are defined which incorporate the results of fixed-nuclei, ab initio calculations: quantum-mechanical electron scattering calculations using the complex Kohn method are performed and augmented by the results of large-scale configuration-interaction calculations performed in a restricted Hilbert space. The global surfaces are defined by a very accurate fit combining a 35

  19. Dissociation of dicarboxylate and disulfonate dianions.

    PubMed

    Ard, Shaun; Mirsaleh-Kohan, Nasrin; Steill, Jeffrey D; Oomens, Jos; Nielsen, Steen Brøndsted; Compton, R N

    2010-03-07

    Collision-induced dissociation (CID), along with infrared multiple photon dissociation/detachment (IRMPD) techniques, is utilized to study a series of doubly substituted aromatic dianions containing sulfonate and carboxylate functionalities (1,2- and 1,3-benzenedisulfonate, 1,5-naphthalenedisulfonate, 2,6-naphthalenedisulfonate, 4-sulfobenzoate, 2,6-naphthalenedicarboxylate, and terephthalate dianions). The molecules were chosen because of the electronegativity of the CO(2) and SO(3) moieties along with their varied spatial separation in order to investigate the effect of the repulsive Coulomb barrier (RCB) on the dianions' respective dissociation pathways. Density functional theory calculations of the structures, electron detachment and dissociation energies, as well as vibrational frequencies are performed. Calculated infrared active vibrational frequencies are largely in agreement with the IRMPD spectra which provide support for interpretations based upon computed energies. Calculated and experimental results show that fragmentation dominates over electron detachment as the lowest energy dissociation pathway for these systems and the nature of this dissociation is dictated by properties of the substituent group. CID and IRMPD of dianions with two sulfonate groups (SO(3)(-)) resulted in a single dissociation channel leading to observation of SO(3)(-) and its anion conjugate pair, whereas the carboxylate (CO(2)(-)) containing dianions dissociated via loss of one or both CO(2) molecules and an electron. The SO(3)(-) collisional dissociation exhibited a clear energetic threshold toward ionic fragmentation with an isomeric dependence that is in reasonable agreement with a simple electrostatic model of the RCB, as well as with published reports on electron photodetachment. The loss of one or both CO(2) units and an electron from CID of the carboxylate dianions appeared with no threshold (dissociation occurs with no collision gas), implying these dianions to be

  20. Models for direct Monte Carlo simulation of coupled vibration-dissociation

    NASA Technical Reports Server (NTRS)

    Haas, Brian L.; Boyd, Iain D.

    1993-01-01

    A new model for reactive collisions is developed within the framework of a particle method, which simulates coupled vibration-dissociation (CVD) behavior in high-temperature gases. The fundamental principles of particle simulation methods are introduced with particular attention given to the probability functions employed to select thermal and reactive collisions. Reaction probability functions are derived which favor vibrationally excited molecules as reaction candidates. The new models derived here are used to simulate CVD behavior during thermochemical relaxation of constant-volume O2 reservoirs, as well as the dissociation incubation behavior of postshock N2 flows for comparisons with previous models and experimental data.

  1. Binding affinities of anti-acetylcholine receptor autoantibodies in myasthenia gravis

    SciTech Connect

    Bray, J.J.; Drachman, D.B.

    1982-01-01

    Antibodies directed against acetylcholine (ACh) receptors are present in the sera of nearly 90% of patients with myasthenia gravis (MG), and are involved in the pathogenesis of this autoimmune disease. However, the antibody titers measured by the standard radioimmunoassay correspond poorly with the clinical severity of the disease. To determine whether this disparity could be accounted for by differences in the binding affinities of anti-ACh receptor antibodies in different patients, we have measured the binding affinities of these autoantibodies in 15 sera from MG patients. The affinity constants (K/sub o/), as determined by Scatchard analysis, were all in the range of 10/sup 10/ M/sup -1/, comparable to the highest values reported in immunized animals. The affinity constants were truly representative of the population of autoantibodies detected by the radioimmunoassay, as shown by the remarkable linearity of the Scatchard plots (r/sup 2/>0.90) and the close correlation between the antibody titers determined by extrapolation of the Scatchard plots and by saturation analysis (r = 0.99; p < 0.001). There was only a 6-fold variation in affinity constants measured in this series of patients despite widely differing antibody titers and severity of the disease. Factors other than the titer and affinity of anti-ACh receptor antibodies may correlate better with the clinical manifestations of MG.

  2. The Hubble Constant.

    PubMed

    Jackson, Neal

    2007-01-01

    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. 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.

  3. Unitaxial constant velocity microactuator

    DOEpatents

    McIntyre, Timothy J.

    1994-01-01

    A uniaxial drive system or microactuator capable of operating in an ultra-high vacuum environment. The mechanism includes a flexible coupling having a bore therethrough, and two clamp/pusher assemblies mounted in axial ends of the coupling. The clamp/pusher assemblies are energized by voltage-operated piezoelectrics therewithin to operatively engage the shaft and coupling causing the shaft to move along its rotational axis through the bore. The microactuator is capable of repeatably positioning to sub-manometer accuracy while affording a scan range in excess of 5 centimeters. Moreover, the microactuator generates smooth, constant velocity motion profiles while producing a drive thrust of greater than 10 pounds. The system is remotely controlled and piezoelectrically driven, hence minimal thermal loading, vibrational excitation, or outgassing is introduced to the operating environment.

  4. A Constant Pressure Bomb

    NASA Technical Reports Server (NTRS)

    Stevens, F W

    1924-01-01

    This report describes a new optical method of unusual simplicity and of good accuracy suitable to study the kinetics of gaseous reactions. The device is the complement of the spherical bomb of constant volume, and extends the applicability of the relationship, pv=rt for gaseous equilibrium conditions, to the use of both factors p and v. The method substitutes for the mechanical complications of a manometer placed at some distance from the seat of reaction the possibility of allowing the radiant effects of reaction to record themselves directly upon a sensitive film. It is possible the device may be of use in the study of the photoelectric effects of radiation. The method makes possible a greater precision in the measurement of normal flame velocities than was previously possible. An approximate analysis shows that the increase of pressure and density ahead of the flame is negligible until the velocity of the flame approaches that of sound.

  5. When constants are important

    SciTech Connect

    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.

  6. The Hubble constant.

    PubMed Central

    Tully, R B

    1993-01-01

    Five methods of estimating distances have demonstrated internal reproducibility at the level of 5-20% rms accuracy. The best of these are the cepheid (and RR Lyrae), planetary nebulae, and surface-brightness fluctuation techniques. Luminosity-line width and Dn-sigma methods are less accurate for an individual case but can be applied to large numbers of galaxies. The agreement is excellent between these five procedures. It is determined that Hubble constant H0 = 90 +/- 10 km.s-1.Mpc-1 [1 parsec (pc) = 3.09 x 10(16) m]. It is difficult to reconcile this value with the preferred world model even in the low-density case. The standard model with Omega = 1 may be excluded unless there is something totally misunderstood about the foundation of the distance scale or the ages of stars. PMID:11607391

  7. Unitaxial constant velocity microactuator

    DOEpatents

    McIntyre, T.J.

    1994-06-07

    A uniaxial drive system or microactuator capable of operating in an ultra-high vacuum environment is disclosed. The mechanism includes a flexible coupling having a bore therethrough, and two clamp/pusher assemblies mounted in axial ends of the coupling. The clamp/pusher assemblies are energized by voltage-operated piezoelectrics therewithin to operatively engage the shaft and coupling causing the shaft to move along its rotational axis through the bore. The microactuator is capable of repeatably positioning to sub-nanometer accuracy while affording a scan range in excess of 5 centimeters. Moreover, the microactuator generates smooth, constant velocity motion profiles while producing a drive thrust of greater than 10 pounds. The system is remotely controlled and piezoelectrically driven, hence minimal thermal loading, vibrational excitation, or outgassing is introduced to the operating environment. 10 figs.

  8. From state dissociation to status dissociatus.

    PubMed

    Antelmi, Elena; Ferri, Raffaele; Iranzo, Alex; Arnulf, Isabelle; Dauvilliers, Yves; Bhatia, Kailash P; Liguori, Rocco; Schenck, Carlos H; Plazzi, Giuseppe

    2016-08-01

    The states of being are conventionally defined by the simultaneous occurrence of behavioral, neurophysiological and autonomic descriptors. State dissociation disorders are due to the intrusion of features typical of a different state into an ongoing state. Disorders related to these conditions are classified according to the ongoing main state and comprise: 1) Dissociation from prevailing wakefulness as seen in hypnagogic or hypnopompic hallucinations, automatic behaviors, sleep drunkenness, cataplexy and sleep paralysis 2) Dissociation from rapid eye movement (REM) sleep as seen in REM sleep behavior disorder and lucid dreaming and 3) Dissociation from NREM sleep as seen in the disorders of arousal. The extreme expression of states dissociation is characterized by the asynchronous occurrence of the various components of the different states that prevents the recognition of any state of being. This condition has been named status dissociatus. According to the underlying disorders/diseases and to their severity, among status dissociatus we may recognize disorders in which such an extreme dissociation occurs only at night time or intermittently (i.e., autoimmune encephalopathies, narcolepsy type 1 and IgLON5 parasomnia), and others in which it occurs nearly continuously with complete loss of any conventionally defined state of being, and of the circadian pattern (agrypnia excitata). Here, we render a comprehensive review of all diseases/disorders associated with state dissociation and status dissociatus and propose a critical classification of this complex scenario.

  9. Objective assessment of peritraumatic dissociation: psychophysiological indicators.

    PubMed

    Griffin, M G; Resick, P A; Mechanic, M B

    1997-08-01

    This study aims to investigate psychophysiological changes associated with peritraumatic dissociation in female victims of recent rape and assess the relation between these changes and symptoms of post-traumatic stress disorder (PTSD). Samples included 85 rape victims from local police departments (84%), plus victim assistance agencies (15%), and hospitals (1%). Results indicate that there is a relatively normal distribution of peritraumatic dissociative symptoms during rape-related trauma. The Peritraumatic Dissociation Index demonstrated good internal consistency in a group of recent rape victims, which supports the use of this type of scale for assessing peritraumatic dissociation in trauma victims. Individuals in the high peritraumatic dissociation group showed a significantly different pattern of physiological responses from those of the low dissociation group. These responses are due to the suppression of autonomic physiological responses. Moreover, this group also contained a larger proportion of subjects (94%) identified as meeting PTSD symptom criteria. Thus, these findings support the idea that there is a dissociative subtype of persons with PTSD who may not process traumatic information, which may lead to greater levels of PTSD symptoms.

  10. Expression and affinity purification of recombinant proteins from plants

    NASA Technical Reports Server (NTRS)

    Desai, Urvee A.; Sur, Gargi; Daunert, Sylvia; Babbitt, Ruth; Li, Qingshun

    2002-01-01

    With recent advances in plant biotechnology, transgenic plants have been targeted as an inexpensive means for the mass production of proteins for biopharmaceutical and industrial uses. However, the current plant purification techniques lack a generally applicable, economic, large-scale strategy. In this study, we demonstrate the purification of a model protein, beta-glucuronidase (GUS), by employing the protein calmodulin (CaM) as an affinity tag. In the proposed system, CaM is fused to GUS. In the presence of calcium, the calmodulin fusion protein binds specifically to a phenothiazine-modified surface of an affinity column. When calcium is removed with a complexing agent, e.g., EDTA, calmodulin undergoes a conformational change allowing the dissociation of the calmodulin-phenothiazine complex and, therefore, permitting the elution of the GUS-CaM fusion protein. The advantages of this approach are the fast, efficient, and economical isolation of the target protein under mild elution conditions, thus preserving the activity of the target protein. Two types of transformation methods were used in this study, namely, the Agrobacterium-mediated system and the viral-vector-mediated transformation system. Copyright 2002 Elsevier Science (USA).

  11. Dissociation in families experiencing Intimate partner violence.

    PubMed

    Miller, Alison

    2017-02-16

    This paper, using an illustrative case study, presents the hypothesis that cyclical spouse abusers suffer from a dissociative condition (or perhaps a personality disorder in which dissociation is a prominent feature) that results from disorganized attachment. The partner of the spouse abuser tries various unsuccessful strategies to appease her spouse in order to change his behavior. If the relationship lasts for years, she adapts by developing a milder but parallel dissociative process, developing chains of state-dependent memory and resultant ego states for the different phases of the domestic abuse cycle. The children suffer from attachment disruption which can potentially continue the process to the next generation.

  12. Products of Dissociative Recombination in the Ionosphere

    NASA Technical Reports Server (NTRS)

    Cosby, Philip

    1996-01-01

    SRI International undertook a novel experimental measurement of the product states formed by dissociative ro-combination (DR) of C2(+), NO(+), and N2(+) as a function of both electron energy and reactant ion vibrational level. For these measurements we used a recently developed experimental technique for measuring dissociation product distributions that allows both the branching ratios to be accurately determined and the electronic and ro-vibrational state composition of the reactant ions to be specified. DR is the dominant electron loss mechanism in all regions of the ionosphere. In this process, electron attachment to the molecular ion produces an unstable neutral molecule that rapidly dissociates.

  13. Wavepacket theory of collisional dissociation in molecules

    SciTech Connect

    Kulander, K.

    1980-01-01

    An explicit integration scheme is used to solve the time dependent Schroedinger equation for wavepackets which model collisions in the collinear H + H/sub 2/ system. A realistic LEPS-type potential energy surface is used. Collision energies considered are above the dissociation threshold and probabilities for collision induced dissociation are reported. Also quantum mechanical state-to-state transition probabilities are generated. These results are compared to extensive classical trajectory calculations performed on this same system. The time evolution of the wavepacket densities is studied to understand the dynamics of the collinear collisional dissociation process.

  14. Dissociative Ionization of Pyridine by Electron Impact

    NASA Technical Reports Server (NTRS)

    Dateo, Christopher; Huo, Winifred; Kwak, Dochan (Technical Monitor)

    2002-01-01

    In order to understand the damage of biomolecules by electrons, a process important in radiation damage, we undertake a study of the dissociative ionization (DI) of pyridine (C5H5N) from the low-lying ionization channels. The methodology used is the same as in the benzene study. While no experimental DI data are available, we compare the dissociation products from our calculations with the dissociative photoionization measurements of Tixier et al. using dipole (e, e(+) ion) coincidence spectroscopy. Comparisons with the DI of benzene is also made so as to understand the difference in DI between a heterocyclic and an aromatic molecule.

  15. Dissociative identity disorder: a controversial diagnosis.

    PubMed

    Gillig, Paulette Marie

    2009-03-01

    A brief description of the controversies surrounding the diagnosis of dissociative identity disorder is presented, followed by a discussion of the proposed similarities and differences between dissociative identity disorder and borderline personality disorder. The phenomenon of autohypnosis in the context of early childhood sexual trauma and disordered attachment is discussed, as is the meaning of alters or alternate personalities. The author describes recent neurosciences research that may relate the symptoms of dissociative identity disorder to demonstrable disordered attention and memory processes. A clinical description of a typical patient presentation is included, plus some recommendations for approaches to treatment.

  16. Theory of dissociative tunneling ionization

    NASA Astrophysics Data System (ADS)

    Svensmark, Jens; Tolstikhin, Oleg I.; Madsen, Lars Bojer

    2016-05-01

    We present a theoretical study of the dissociative tunneling ionization process. Analytic expressions for the nuclear kinetic energy distribution of the ionization rates are derived. A particularly simple expression for the spectrum is found by using the Born-Oppenheimer (BO) approximation in conjunction with the reflection principle. These spectra are compared to exact non-BO ab initio spectra obtained through model calculations with a quantum mechanical treatment of both the electronic and nuclear degrees of freedom. In the regime where the BO approximation is applicable, imaging of the BO nuclear wave function is demonstrated to be possible through reverse use of the reflection principle, when accounting appropriately for the electronic ionization rate. A qualitative difference between the exact and BO wave functions in the asymptotic region of large electronic distances is shown. Additionally, the behavior of the wave function across the turning line is seen to be reminiscent of light refraction. For weak fields, where the BO approximation does not apply, the weak-field asymptotic theory describes the spectrum accurately.

  17. Complex high affinity interactions occur between MHCI and superantigens

    NASA Technical Reports Server (NTRS)

    Chapes, S. K.; Herpich, A. R.; Spooner, B. S. (Principal Investigator)

    1998-01-01

    Staphylococcal enterotoxins A and C1 (SEA or SEC1) bound to major histocompatibility-I (MHCI) molecules with high affinity (binding constants ranging from 1.1 microM to 79 nM). SEA and SEC1 directly bound MHCI molecules that had been captured by monoclonal antibodies specific for H-2Kk, H-2Dk, or both. In addition, MHCI-specific antibodies inhibited the binding of SEC1 to LM929 cells and SEA competitively inhibited SEC1 binding; indicating that the superantigens bound to MHCI on the cell surface. The affinity and number of superantigen binding sites differed depending on whether MHCI was expressed in the membrane of LM929 cells or whether it was captured. These data support the hypothesis that MHCI molecules can serve as superantigen receptors.

  18. Polysulfone affinity membranes for the treatment of amino acid mixtures.

    PubMed

    Rodemann, K; Staude, E

    1995-06-20

    Affinity membranes for the treatment of solutions containing amino acids were obtained via lithiating polysulfone that was subsequently converted with glycidylether. From this polymer asymmetric ultrafiltration membranes were cast. The membranes were reacted with iminodiacetic acid yielding membranes fitted out with bidentate chelates. The same reaction path was applied to commercially available symmetric microfiltration membranes. The chelate-bearing membranes were complexed with Cu, Ni, and Zn ions. For the experiments with amino acids only the Cu-complexed membranes were used. The complexation constants for histidine and tryptophan for six different membranes were determined. Because of the affinity of these two amino acids for the complexed Cu ions, they could easily be separated from solutions containing amino acids such as alanine, glycine, and valine. Also, concentrating very dilute amino acid solutions was carried out successfully.

  19. Probing the electrostatics and pharmacological modulation of sequence-specific binding by the DNA-binding domain of the ETS family transcription factor PU.1: a binding affinity and kinetics investigation.

    PubMed

    Munde, Manoj; Poon, Gregory M K; Wilson, W David

    2013-05-27

    Members of the ETS family of transcription factors regulate a functionally diverse array of genes. All ETS proteins share a structurally conserved but sequence-divergent DNA-binding domain, known as the ETS domain. Although the structure and thermodynamics of the ETS-DNA complexes are well known, little is known about the kinetics of sequence recognition, a facet that offers potential insight into its molecular mechanism. We have characterized DNA binding by the ETS domain of PU.1 by biosensor-surface plasmon resonance (SPR). SPR analysis revealed a striking kinetic profile for DNA binding by the PU.1 ETS domain. At low salt concentrations, it binds high-affinity cognate DNA with a very slow association rate constant (≤10(5)M(-)(1)s(-)(1)), compensated by a correspondingly small dissociation rate constant. The kinetics are strongly salt dependent but mutually balance to produce a relatively weak dependence in the equilibrium constant. This profile contrasts sharply with reported data for other ETS domains (e.g., Ets-1, TEL) for which high-affinity binding is driven by rapid association (>10(7)M(-)(1)s(-)(1)). We interpret this difference in terms of the hydration properties of ETS-DNA binding and propose that at least two mechanisms of sequence recognition are employed by this family of DNA-binding domain. Additionally, we use SPR to demonstrate the potential for pharmacological inhibition of sequence-specific ETS-DNA binding, using the minor groove-binding distamycin as a model compound. Our work establishes SPR as a valuable technique for extending our understanding of the molecular mechanisms of ETS-DNA interactions as well as developing potential small-molecule agents for biotechnological and therapeutic purposes.

  20. Investigation of the effect of mutations of rat albumin on the binding affinity to the alpha(4)beta(1) integrin antagonist, 4-[1-[3-chloro-4-[N'-(2-methylphenyl)ureido]phenylacetyl]-(4S)-fluoro-(2S)-pyrrolidine-2-yl]methoxybenzoic acid (D01-4582), using recombinant rat albumins.

    PubMed

    Ito, Takashi; Takahashi, Masayuki; Okazaki, Osamu; Sugiyama, Yuichi

    2010-08-02

    The authors reported previously rat strain differences in plasma protein binding to alpha(4)beta(1) antagonist D01-4582, resulting in a great strain difference in its pharmacokinetics (19-fold differences in the AUC). The previous study suggested that amino acid changes of V238L and/or T293I in albumin reduced the binding affinity. In order to elucidate the relative significance of these mutations, an expression system was developed to obtain recombinant rat albumins (rRSA) using Pichia pastoris, followed by a binding analysis of four rRSAs by the ultracentrifugation method. The equilibrium dissociation constant (K(d)) of wild-type rRSA was 210 nM, while K(d) of rRSA that carried both V238L and T293I mutations was 974 nM. K(d) of artificial rRSA that carried only V238L was 426 nM, and K(d) of artificial rRSA that carried only T293I was 191 nM. These results suggested that V238L would be more important in the alteration of K(d). However, since none of the single mutations were sufficient to explain the reduction of affinity, the possibility was also suggested that T293I interacted cooperatively to reduce the binding affinity of rat albumin to D01-4582. Further investigation is required to elucidate the mechanism of the possible cooperative interaction.

  1. Non-dissociative and dissociative ionization of a CO+ beam in intense ultrashort laser pulses

    NASA Astrophysics Data System (ADS)

    Gaire, B.; Ablikim, U.; Zohrabi, M.; Roland, S.; Carnes, K. D.; Ben-Itzhak, I.

    2011-05-01

    We have investigated the ionization of CO+ beams in intense ultrashort laser pulses. With the recent upgrades to our coincidence three-dimensional momentum imaging method we are able to measure both non-dissociative and dissociative ionization of the molecular-ion beam targets. Using CO+ as an example, we have found that non-dissociative ionization (leading to the metastable dication CO2+) involves a direct transition, i.e. the molecule is ionized with little or no internuclear distance stretch. Dissociative ionization (C+ + O+) occurs both directly and indirectly, stretching first and then ionizing. Our results show that the yield of dissociative ionization is higher than that of non-dissociative ionization and can be manipulated with the laser pulse duration by suppressing the indirect ionization path using ultrashort pulses (<=10 fs). Supported by the Chemical Sciences, Geosciences and Biosciences Division, Office of Basic Energy Sciences, Office of Science, U.S. Department of Energy.

  2. Electron attachment and detachment: Electron affinities of isomers of trifluoromethylbenzonitrile

    NASA Astrophysics Data System (ADS)

    Miller, Thomas M.; Viggiano, A. A.; Friedman, Jeffrey F.; Van Doren, Jane M.

    2004-11-01

    Rate constants for electron attachment to the three isomers of trifluoromethylbenzonitrile [(CF3)(CN)C6H4, or TFMBN] were measured over the temperature range of 303-463 K in a 133-Pa He buffer gas, using a flowing-afterglow Langmuir-probe apparatus. At 303 K, the measured attachment rate constants are 9.0×10-8 (o-TFMBN), 5.5×10-8 (m-TFMBN), and 8.9×10-8 cm3 s-1 (p-TFMBN), estimated accurate to ±25%. The attachment process formed only the parent anion in all three cases. Thermal electron detachment was observed for all three anion isomers, and rate constants for this reverse process were also measured. From the attachment and detachment results, the electron affinities of the three isomers of TFMBN were determined to be 0.70(o-TFMBN), 0.67(m-TFMBN), and 0.83 eV (p-TFMBN), all ±0.05 eV. G3(MP2) [Gaussian-3 calculations with reduced Møller-Plesset orders (MP2)] calculations were carried out for the neutrals and anions. Electron affinities derived from these calculations are in good agreement with the experimental values.

  3. Electron attachment and detachment: electron affinities of isomers of trifluoromethylbenzonitrile.

    PubMed

    Miller, Thomas M; Viggiano, A A; Friedman, Jeffrey F; Van Doren, Jane M

    2004-11-22

    Rate constants for electron attachment to the three isomers of trifluoromethylbenzonitrile [(CF(3))(CN)C(6)H(4), or TFMBN] were measured over the temperature range of 303-463 K in a 133-Pa He buffer gas, using a flowing-afterglow Langmuir-probe apparatus. At 303 K, the measured attachment rate constants are 9.0 x 10(-8) (o-TFMBN), 5.5 x 10(-8) (m-TFMBN), and 8.9 x 10(-8) cm(3) s(-1) (p-TFMBN), estimated accurate to +/-25%. The attachment process formed only the parent anion in all three cases. Thermal electron detachment was observed for all three anion isomers, and rate constants for this reverse process were also measured. From the attachment and detachment results, the electron affinities of the three isomers of TFMBN were determined to be 0.70(o-TFMBN), 0.67(m-TFMBN), and 0.83 eV (p-TFMBN), all +/-0.05 eV. G3(MP2) [Gaussian-3 calculations with reduced Møller-Plesset orders (MP2)] calculations were carried out for the neutrals and anions. Electron affinities derived from these calculations are in good agreement with the experimental values.

  4. Higher Nucleoporin-Importinβ Affinity at the Nuclear Basket Increases Nucleocytoplasmic Import

    PubMed Central

    Azimi, Mohammad; Mofrad, Mohammad R. K.

    2013-01-01

    Several in vitro studies have shown the presence of an affinity gradient in nuclear pore complex proteins for the import receptor Importinβ, at least partially contributing to nucleocytoplasmic transport, while others have historically argued against the presence of such a gradient. Nonetheless, the existence of an affinity gradient has remained an uncharacterized contributing factor. To shed light on the affinity gradient theory and better characterize how the existence of such an affinity gradient between the nuclear pore and the import receptor may influence the nucleocytoplasmic traffic, we have developed a general-purpose agent based modeling (ABM) framework that features a new method for relating rate constants to molecular binding and unbinding probabilities, and used our ABM approach to quantify the effects of a wide range of forward and reverse nucleoporin-Importinβ affinity gradients. Our results indicate that transport through the nuclear pore complex is maximized with an effective macroscopic affinity gradient of 2000 µM, 200 µM and 10 µM in the cytoplasmic, central channel and nuclear basket respectively. The transport rate at this gradient is approximately 10% higher than the transport rate for a comparable pore lacking any affinity gradient, which has a peak transport rate when all nucleoporins have an affinity of 200 µM for Importinβ. Furthermore, this optimal ratio of affinity gradients is representative of the ratio of affinities reported for the yeast nuclear pore complex – suggesting that the affinity gradient seen in vitro is highly optimized. PMID:24282617

  5. A review of dissociative disorders treatment studies.

    PubMed

    Brand, Bethany L; Classen, Catherine C; McNary, Scot W; Zaveri, Parin

    2009-09-01

    This review examines empirical reports of treatment for Dissociative Disorders (DD), including 16 DD treatment outcome studies and 4 case studies that used standardized measures. Collectively, these reports suggest that treatment for DD is associated with decreased symptoms of dissociation, depression, posttraumatic stress disorder, distress, and suicidality. Effect sizes, based on pre/post measures, are in the medium to large range across studies. Patients with dissociative disorder who integrated their dissociated self states were found to have reduced symptomatology compared with those who did not integrate. The magnitude of pre/post effect sizes for these DD studies are comparable to pre/post effect sizes in treatment studies of complex PTSD. There are significant methodological limitations in the current DD treatment outcome literature that reduce internal and external validity including regression towards the mean, limited sample sizes, and nonrandomized research designs. Implications for future research and treatment planning for patients suffering from DD are discussed.

  6. Collision-induced gas phase dissociation rates

    NASA Technical Reports Server (NTRS)

    Hansen, C. Frederick

    1990-01-01

    The Landau-Zener theory of reactive cross sections was applied to diatomic molecules dissociating from a ladder of vibrational states. The result predicts a dissociation rate that is quite well duplicated by an Arrhenius function having a preexponential temperature dependence of about T(sub -1/2), at least for inert collision partners. This relation fits experimental data reasonably well. The theory is then used to calculate the effect of vibrational nonequilibrium on dissociation rate. For Morse oscillators, the results are about the same as given by Hammerling, Kivel, and Teare in their analytic approximation for harmonic oscillators, though at very high temperature a correction for the partition function limit is included. The empirical correction for vibration nonequilibrium proposed by Park, which is a convenient algorithm for CFD calculations, is modified to prevent a drastic underestimation of dissociation rates that occurs with this method when vibrational temperature is much smaller than the kinetic temperature of the gas.

  7. ION PAIR DISSOCIATION: Spectroscopy and Dynamics

    NASA Astrophysics Data System (ADS)

    Suits, Arthur G.; Hepburn, John W.

    2006-05-01

    Ion pair dissociation processes may be studied using coherent vacuum ultraviolet laser sources in a manner entirely analogous to photoelectron spectroscopy, albeit with the anion playing the role of a heavy electron. If the excitation energy is above the dissociation energy and the kinetic energy of the fragment is measured using ion imaging, this approach is termed ion pair imaging spectroscopy (IPIS) and is related to conventional photoelectron spectroscopy. If the excitation energy is just below the dissociation energy and pulsed-field dissociation is employed, this approach is analogous to mass analyzed threshold ionization (MATI) spectroscopy and is termed threshold ion pair production spectroscopy (TIPPS). These approaches provide a novel means of investigating ion thermochemistry and spectroscopy and superexcited state decay dynamics at high resolution.

  8. Characterization of Tyrosine Nitration and Cysteine Nitrosylation Modifications by Metastable Atom-Activation Dissociation Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Cook, Shannon L.; Jackson, Glen P.

    2011-02-01

    The fragmentation behavior of nitrated and S-nitrosylated peptides were studied using collision induced dissociation (CID) and metastable atom-activated dissociation mass spectrometry (MAD-MS). Various charge states, such as 1+, 2+, 3+, 2-, of modified and unmodified peptides were exposed to a beam of high kinetic energy helium (He) metastable atoms resulting in extensive backbone fragmentation with significant retention of the post-translation modifications (PTMs). Whereas the high electron affinity of the nitrotyrosine moiety quenches radical chemistry and fragmentation in electron capture dissociation (ECD) and electron transfer dissociation (ETD), MAD does produce numerous backbone cleavages in the vicinity of the modification. Fragment ions of nitrosylated cysteine modifications typically exhibit more abundant neutral losses than nitrated tyrosine modifications because of the extremely labile nature of the nitrosylated cysteine residues. However, compared with CID, MAD produced between 66% and 86% more fragment ions, which preserved the labile -NO modification. MAD was also able to differentiate I/L residues in the modified peptides. MAD is able to induce radical ion chemistry even in the presence of strong radical traps and therefore offers unique advantages to ECD, ETD, and CID for determination of PTMs such as nitrated and S-nitrosylated peptides.

  9. Differential Mobility Spectrometry: Preliminary Findings on Determination of Fundamental Constants

    NASA Technical Reports Server (NTRS)

    Limero, Thomas; Cheng, Patti; Boyd, John

    2007-01-01

    The electron capture detector (ECD) has been used for 40+ years (1) to derive fundamental constants such as a compound's electron affinity. Given this historical perspective, it is not surprising that differential mobility spectrometry (DMS) might be used in a like manner. This paper will present data from a gas chromatography (GC)-DMS instrument that illustrates the potential capability of this device to derive fundamental constants for electron-capturing compounds. Potential energy curves will be used to provide possible explanation of the data.

  10. Paranormal belief and proneness to dissociation.

    PubMed

    Irwin, H J

    1994-12-01

    The study investigated the relationship between scores on paranormal belief and proneness to dissociation in a sample of 100 Australian students of psychology. Scores on dissociation were positively correlated with those on global paranormal belief and with belief in psi, precognition, spiritualism, and extraordinary life-forms. It is suggested that in some instances paranormal beliefs may be a component of a complex defensive framework constructed in the face of the perceived uncontrollability of life.

  11. Dissociation energy of molecules in dense gases

    NASA Technical Reports Server (NTRS)

    Kunc, J. A.

    1992-01-01

    A general approach is presented for calculating the reduction of the dissociation energy of diatomic molecules immersed in a dense (n = less than 10 exp 22/cu cm) gas of molecules and atoms. The dissociation energy of a molecule in a dense gas differs from that of the molecule in vacuum because the intermolecular forces change the intramolecular dynamics of the molecule, and, consequently, the energy of the molecular bond.

  12. Dissociation energies of PH and PH+.

    NASA Astrophysics Data System (ADS)

    Reddy, R. R.; Nazeer Ahammed, Y.; Srinivasa Rao, A.; Rao, T. V. R.

    1995-12-01

    Dissociation energies for the ground electronic states of diatomic PH and PH+ are determined by fitting empirical potential functions to the respective RKRV curves using correlation coefficients. The estimated ground state dissociation energies of PH and PH+ are 3.10 and 3.20 eV respectively by the curve fitting procedure using the Lippincott potential function. The computed values are in good agreement with experimental values.

  13. A novel technique for measurement of thermal rate constants and temperature dependences of dissociative recombination: CO{sub 2}{sup +}, CF{sub 3}{sup +}, N{sub 2}O{sup +}, C{sub 7}H{sub 8}{sup +}, C{sub 7}H{sub 7}{sup +}, C{sub 6}H{sub 6}{sup +}, C{sub 6}H{sub 5}{sup +}, C{sub 5}H{sub 6}{sup +}, C{sub 4}H{sub 4}{sup +}, and C{sub 3}H{sub 3}{sup +}

    SciTech Connect

    Fournier, Joseph A.; Shuman, Nicholas S.; Melko, Joshua J.; Ard, Shaun G.; Viggiano, Albert A.

    2013-04-21

    A novel technique using a flowing afterglow-Langmuir probe apparatus for measurement of temperature dependences of rate constants for dissociative recombination (DR) is presented. Low ({approx}10{sup 11} cm{sup -3}) concentrations of a neutral precursor are added to a noble gas/electron afterglow plasma thermalized at 300-500 K. Charge exchange yields one or many cation species, each of which may undergo DR. Relative ion concentrations are monitored at a fixed reaction time while the initial plasma density is varied between 10{sup 9} and 10{sup 10} cm{sup -3}. Modeling of the decrease in concentration of each cation relative to the non-recombining noble gas cation yields the rate constant for DR. The technique is applied to several species (O{sub 2}{sup +}, CO{sub 2}{sup +}, CF{sub 3}{sup +}, N{sub 2}O{sup +}) with previously determined 300 K values, showing excellent agreement. The measurements of those species are extended to 500 K, with good agreement to literature values where they exist. Measurements are also made for a range of C{sub n}H{sub m}{sup +} (C{sub 7}H{sub 7}{sup +}, C{sub 7}H{sub 8}{sup +}, C{sub 5}H{sub 6}{sup +}, C{sub 4}H{sub 4}{sup +}, C{sub 6}H{sub 5}{sup +}, C{sub 3}H{sub 3}{sup +}, and C{sub 6}H{sub 6}{sup +}) derived from benzene and toluene neutral precursors. C{sub n}H{sub m}{sup +} DR rate constants vary from 8-12 Multiplication-Sign 10{sup -7} cm{sup 3} s{sup -1} at 300 K with temperature dependences of approximately T{sup -0.7}. Where prior measurements exist these results are in agreement, with the exception of C{sub 3}H{sub 3}{sup +} where the present results disagree with a previously reported flat temperature dependence.

  14. Water binding energies of [Pb(amino acid-H)H2O]+ complexes determined by blackbody infrared radiative dissociation.

    PubMed

    Burt, Michael B; Decker, Sarah G A; Fridgen, Travis D

    2012-11-21

    The water binding energies (E(0)) of eight deprotonated Pb(2+)-amino acid (Aa) complexes of the form [Pb(Aa-H)H(2)O](+) (Aa = Gly, Ala, Val, Leu, Ile, Phe, Glu, and Lys) were determined using blackbody infrared radiative dissociation (BIRD). A Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometer was used to trap ions generated by electrospray ionization (ESI) in a "zero"-pressure (~10(-10) torr) cell where dissociation can only occur by absorption of thermally generated photons. Since the [Pb(Aa-H)H(2)O](+) complexes have relatively few vibrational degrees of freedom (36-78) and are within the slow-exchange kinetic limit, the master equation was solved to extract meaningful threshold dissociation energies and thermal unimolecular dissociation rate constants (k(uni)). The master equation analysis uses variable reaction coordinate transition state theory (VRC-TST) to minimize the Rice-Ramsperger-Kassel-Marcus (RRKM) dissociation rate constants. The determined water binding energies range from 76.6 to 113.6 kJ mol(-1), and agree well with 0 K dissociation energies calculated using the B3LYP/6-31+G(d,p) and MP2(full)/6-311++G(2d,2p)//B3LYP/6-31+G(d,p) methods. The relative strengths of the binding energies reflect the known structural isomers (A-, B-, C-, and D-type) of these [Pb(Aa-H)H(2)O](+) complexes.

  15. Dissociative Ionization of Benzene by Electron Impact

    NASA Technical Reports Server (NTRS)

    Huo, Winifred; Dateo, Christopher; Kwak, Dochan (Technical Monitor)

    2002-01-01

    We report a theoretical study of the dissociative ionization (DI) of benzene from the low-lying ionization channels. Our approach makes use of the fact that electron motion is much faster than nuclear motion and DI is treated as a two-step process. The first step is electron-impact ionization resulting in an ion with the same nuclear geometry as the neutral molecule. In the second step the nuclei relax from the initial geometry and undergo unimolecular dissociation. For the ionization process we use the improved binary-encounter dipole (iBED) model. For the unimolecular dissociation step, we study the steepest descent reaction path to the minimum of the ion potential energy surface. The path is used to analyze the probability of unimolecular dissociation and to determine the product distributions. Our analysis of the dissociation products and the thresholds of the productions are compared with the result dissociative photoionization measurements of Feng et al. The partial oscillator strengths from Feng et al. are then used in the iBED cross section calculations.

  16. Selective export of MHC class I molecules from the ER after their dissociation from TAP.

    PubMed

    Spiliotis, E T; Manley, H; Osorio, M; Zúñiga, M C; Edidin, M

    2000-12-01

    It has been assumed that upon dissociation from TAP, MHC class I molecules exit the ER by nonselective bulk flow. We now show that exit must occur by association with cargo receptors. Inconsistent with exit by bulk flow, loading of MHC class I molecules with high-affinity peptides triggers dissociation from TAP but has no effect on rates of ER-to-Golgi transport. Moreover, peptide-loaded MHC class I molecules accumulate at ER exit sites from which TAP molecules are excluded. Consistent with receptor-mediated exit, ER-to-Golgi transport of MHC class I molecules is independent of their cytoplasmic tails, which themselves lack ER export motifs. In addition, we show that MHC class I molecules associate with the putative cargo receptor BAP31.

  17. Hyphenated affinity capillary electrophoresis with a high-sensitivity cell for the simultaneous binding study of retinol and retinoic acid in nanomolars with serum albumins.

    PubMed

    El-Hady, D Abd; Albishri, H M

    2012-12-12

    Retinol and retinoic acid are Vitamin A components that are critical for many biological processes. Both of them are strongly complexing with serum albumins giving constants of the order of 10(5)Lmol(-1) or higher. With respect to this fact, affinity capillary electrophoresis (ACE) is not applicable in its commonly used form. Therefore, for the first time, the hyphenated ACE with a high-sensitivity cell was developed and employed to investigate the binding of retinol and retinoic acid in nanomolars with human serum albumin (HSA) and bovine serum albumin (BSA) under physiological conditions. ACE/high-sensitivity coupled cell had contributed to fast the association and dissociation rates of the complexes in nanomolar scale of analytes ensuring the establishment of a dynamic equilibrium within a short electrophoresis time. In addition, this hyphenation led to reduce the concentrations of serum albumins as additives in background electrolyte making a sense beside the proper rinsing protocol for the negligible possibility of their adsorption. The mobility ratio based on nonlinear regression analysis was used to deduce precise binding constants of analytes with serum albumins. The binding constants (K, Lmol(-1)) of retinol were 1.28×10(5) and 5.25×10(6) and retinoic acid were 3.29×10(5) and 2.27×10(6) with HSA and BSA, respectively. The displacement and reciprocal competitive binding of analytes were investigated and indicated that retinoic acid was able to replace retinol from HSA and vice versa in the case of BSA.

  18. Affinity membrane introduction mass spectrometry

    SciTech Connect

    Xu, C.; Patrick, J.S.; Cooks, R.G. )

    1995-02-15

    A new technique, affinity membrane introduction mass spectrometry, is described. In this method, a chemically modified membrane is used to selectively adsorb analytes bearing a particular functional group and concentrate them from solution. Release of the bound analyte results in its transfer across the membrane and allows it to be monitored mass spectrometrically, using, in the present case, a benchtop ion trap instrument. Alkylamine-modified cellulose membranes are used to bind substituted benzaldehydes through imine formation at high pH. Release of the bound aldehyde is achieved by acid hydrolysis of the surface-bound imine. Benzaldehyde is detected with excellent specificity at 10 ppm in a complex mixture using this method. Using the enrichment capability of the membrane, a full mass spectrum of benzaldehyde can be measured at a concentration of 10 ppb. The behavior of a variety of other aldehydes is also discussed to illustrate the capabilities of the method. 21 refs., 5 figs., 2 tabs.

  19. An improved potential energy surface and multi-temperature quasiclassical trajectory calculations of N2 + N2 dissociation reactions

    NASA Astrophysics Data System (ADS)

    Bender, Jason D.; Valentini, Paolo; Nompelis, Ioannis; Paukku, Yuliya; Varga, Zoltan; Truhlar, Donald G.; Schwartzentruber, Thomas; Candler, Graham V.

    2015-08-01

    Accurate modeling of high-temperature hypersonic flows in the atmosphere requires consideration of collision-induced dissociation of molecular species and energy transfer between the translational and internal modes of the gas molecules. Here, we describe a study of the N2 + N2⟶N2 + 2N and N2 + N2⟶4N nitrogen dissociation reactions using the quasiclassical trajectory (QCT) method. The simulations used a new potential energy surface for the N4 system; the surface is an improved version of one that was presented previously. In the QCT calculations, initial conditions were determined based on a two-temperature model that approximately separates the translational-rotational temperature from the vibrational temperature of the N2 diatoms. Five values from 8000 K to 30 000 K were considered for each of the two temperatures. Over 2.4 × 109 trajectories were calculated. We present results for ensemble-averaged dissociation rate constants as functions of the translational-rotational temperature T and the vibrational temperature Tv. The rate constant depends more strongly on T when Tv is low, and it depends more strongly on Tv when T is low. Quasibound reactant states contribute significantly to the rate constants, as do exchange processes at higher temperatures. We discuss two sets of runs in detail: an equilibrium test set in which T = Tv and a nonequilibrium test set in which Tv < T. In the equilibrium test set, high-v and moderately-low-j molecules contribute most significantly to the overall dissociation rate, and this state specificity becomes stronger as the temperature decreases. Dissociating trajectories tend to result in a major loss of vibrational energy and a minor loss of rotational energy. In the nonequilibrium test set, as Tv decreases while T is fixed, higher-j molecules contribute more significantly to the dissociation rate, dissociating trajectories tend to result in a greater rotational energy loss, and the dissociation probability's dependence on v

  20. An improved potential energy surface and multi-temperature quasiclassical trajectory calculations of N2 + N2 dissociation reactions.

    PubMed

    Bender, Jason D; Valentini, Paolo; Nompelis, Ioannis; Paukku, Yuliya; Varga, Zoltan; Truhlar, Donald G; Schwartzentruber, Thomas; Candler, Graham V

    2015-08-07

    Accurate modeling of high-temperature hypersonic flows in the atmosphere requires consideration of collision-induced dissociation of molecular species and energy transfer between the translational and internal modes of the gas molecules. Here, we describe a study of the N2 + N2⟶N2 + 2N and N2 + N2⟶4N nitrogen dissociation reactions using the quasiclassical trajectory (QCT) method. The simulations used a new potential energy surface for the N4 system; the surface is an improved version of one that was presented previously. In the QCT calculations, initial conditions were determined based on a two-temperature model that approximately separates the translational-rotational temperature from the vibrational temperature of the N2 diatoms. Five values from 8000 K to 30,000 K were considered for each of the two temperatures. Over 2.4 × 10(9) trajectories were calculated. We present results for ensemble-averaged dissociation rate constants as functions of the translational-rotational temperature T and the vibrational temperature T(v). The rate constant depends more strongly on T when T(v) is low, and it depends more strongly on T(v) when T is low. Quasibound reactant states contribute significantly to the rate constants, as do exchange processes at higher temperatures. We discuss two sets of runs in detail: an equilibrium test set in which T = T(v) and a nonequilibrium test set in which T(v) < T. In the equilibrium test set, high-v and moderately-low-j molecules contribute most significantly to the overall dissociation rate, and this state specificity becomes stronger as the temperature decreases. Dissociating trajectories tend to result in a major loss of vibrational energy and a minor loss of rotational energy. In the nonequilibrium test set, as T(v) decreases while T is fixed, higher-j molecules contribute more significantly to the dissociation rate, dissociating trajectories tend to result in a greater rotational energy loss, and the dissociation probability

  1. A Simple Method for the Consecutive Determination of Protonation Constants through Evaluation of Formation Curves

    ERIC Educational Resources Information Center

    Hurek, Jozef; Nackiewicz, Joanna

    2013-01-01

    A simple method is presented for the consecutive determination of protonation constants of polyprotic acids based on their formation curves. The procedure is based on generally known equations that describe dissociation equilibria. It has been demonstrated through simulation that the values obtained through the proposed method are sufficiently…

  2. Ab initio state-specific N2 + O dissociation and exchange modeling for molecular simulations

    NASA Astrophysics Data System (ADS)

    Luo, Han; Kulakhmetov, Marat; Alexeenko, Alina

    2017-02-01

    Quasi-classical trajectory (QCT) calculations are used in this work to calculate state-specific N2(X1Σ ) +O(3P ) →2 N(4S ) +O(3P ) dissociation and N2(X1Σ ) +O(3P ) →NO(X2Π ) +N(4S ) exchange cross sections and rates based on the 13A″ and 13A' ab initio potential energy surface by Gamallo et al. [J. Chem. Phys. 119, 2545-2556 (2003)]. The calculations consider translational energies up to 23 eV and temperatures between 1000 K and 20 000 K. Vibrational favoring is observed for dissociation reaction at the whole range of collision energies and for exchange reaction around the dissociation limit. For the same collision energy, cross sections for v = 30 are 4 to 6 times larger than those for the ground state. The exchange reaction has an effective activation energy that is dependent on the initial rovibrational level, which is different from dissociation reaction. In addition, the exchange cross sections have a maximum when the total collision energy (TCE) approaches dissociation energy. The calculations are used to generate compact QCT-derived state-specific dissociation (QCT-SSD) and QCT-derived state-specific exchange (QCT-SSE) models, which describe over 1 × 106 cross sections with about 150 model parameters. The models can be used directly within direct simulation Monte Carlo and computational fluid dynamics simulations. Rate constants predicted by the new models are compared to the experimental measurements, direct QCT calculations and predictions by other models that include: TCE model, Bose-Candler QCT-based exchange model, Macheret-Fridman dissociation model, Macheret's exchange model, and Park's two-temperature model. The new models match QCT-calculated and experimental rates within 30% under nonequilibrium conditions while other models under predict by over an order of magnitude under vibrationally-cold conditions.

  3. Ab initio state-specific N2 + O dissociation and exchange modeling for molecular simulations.

    PubMed

    Luo, Han; Kulakhmetov, Marat; Alexeenko, Alina

    2017-02-21

    Quasi-classical trajectory (QCT) calculations are used in this work to calculate state-specific N2(X(1)Σ)+O((3)P)→2N((4)S)+O((3)P) dissociation and N2(X(1)Σ)+O((3)P)→NO(X(2)Π)+N((4)S) exchange cross sections and rates based on the 1(3)A″ and 1(3)A' ab initio potential energy surface by Gamallo et al. [J. Chem. Phys. 119, 2545-2556 (2003)]. The calculations consider translational energies up to 23 eV and temperatures between 1000 K and 20 000 K. Vibrational favoring is observed for dissociation reaction at the whole range of collision energies and for exchange reaction around the dissociation limit. For the same collision energy, cross sections for v = 30 are 4 to 6 times larger than those for the ground state. The exchange reaction has an effective activation energy that is dependent on the initial rovibrational level, which is different from dissociation reaction. In addition, the exchange cross sections have a maximum when the total collision energy (TCE) approaches dissociation energy. The calculations are used to generate compact QCT-derived state-specific dissociation (QCT-SSD) and QCT-derived state-specific exchange (QCT-SSE) models, which describe over 1 × 10(6) cross sections with about 150 model parameters. The models can be used directly within direct simulation Monte Carlo and computational fluid dynamics simulations. Rate constants predicted by the new models are compared to the experimental measurements, direct QCT calculations and predictions by other models that include: TCE model, Bose-Candler QCT-based exchange model, Macheret-Fridman dissociation model, Macheret's exchange model, and Park's two-temperature model. The new models match QCT-calculated and experimental rates within 30% under nonequilibrium conditions while other models under predict by over an order of magnitude under vibrationally-cold conditions.

  4. Facilitated Dissociation of a Nucleoid Protein from the Bacterial Chromosome

    PubMed Central

    Hadizadeh, Nastaran; Johnson, Reid C.

    2016-01-01

    ABSTRACT Off-rates of proteins from the DNA double helix are widely considered to be dependent only on the interactions inside the initially bound protein-DNA complex and not on the concentration of nearby molecules. However, a number of recent single-DNA experiments have shown off-rates that depend on solution protein concentration, or “facilitated dissociation.” Here, we demonstrate that this effect occurs for the major Escherichia coli nucleoid protein Fis on isolated bacterial chromosomes. We isolated E. coli nucleoids and showed that dissociation of green fluorescent protein (GFP)-Fis is controlled by solution Fis concentration and exhibits an “exchange” rate constant (kexch) of ≈104 M−1 s−1, comparable to the rate observed in single-DNA experiments. We also show that this effect is strongly salt dependent. Our results establish that facilitated dissociation can be observed in vitro on chromosomes assembled in vivo. IMPORTANCE Bacteria are important model systems for the study of gene regulation and chromosome dynamics, both of which fundamentally depend on the kinetics of binding and unbinding of proteins to DNA. In experiments on isolated E. coli chromosomes, this study showed that the prolific transcription factor and chromosome packaging protein Fis displays a strong dependence of its off-rate from the bacterial chromosome on Fis concentration, similar to that observed in in vitro experiments. Therefore, the free cellular DNA-binding protein concentration can strongly affect lifetimes of proteins bound to the chromosome and must be taken into account in quantitative considerations of gene regulation. These results have particularly profound implications for transcription factors where DNA binding lifetimes can be a critical determinant of regulatory function. PMID:27044624

  5. Calculation of rates of exciton dissociation into hot charge-transfer states in model organic photovoltaic interfaces

    NASA Astrophysics Data System (ADS)

    Vázquez, Héctor; Troisi, Alessandro

    2013-11-01

    We investigate the process of exciton dissociation in ordered and disordered model donor/acceptor systems and describe a method to calculate exciton dissociation rates. We consider a one-dimensional system with Frenkel states in the donor material and states where charge transfer has taken place between donor and acceptor. We introduce a Green's function approach to calculate the generation rates of charge-transfer states. For disorder in the Frenkel states we find a clear exponential dependence of charge dissociation rates with exciton-interface distance, with a distance decay constant β that increases linearly with the amount of disorder. Disorder in the parameters that describe (final) charge-transfer states has little effect on the rates. Exciton dissociation invariably leads to partially separated charges. In all cases final states are “hot” charge-transfer states, with electron and hole located far from the interface.

  6. Modeling electrostatic and heterogeneity effects on proton dissociation from humic substances

    USGS Publications Warehouse

    Tipping, E.; Reddy, M.M.; Hurley, M.A.

    1990-01-01

    The apparent acid dissociation constant of humic substances increases by 2-4 pK units as ionization of the humic carboxylate groups proceeds. This change in apparent acid strength is due in part to the increase in electrical charge on the humic molecules as protons are shed. In addition, proton dissociation reactions are complicated because humic substances are heterogeneous with respect to proton dissociating groups and molecular size. In this paper, we use the Debye-Hu??ckel theory to describe the effects of electrostatic interactions on proton dissociation of humic substances. Simulations show that, for a size-heterogeneous system of molecules, the weight-average molecular weight is preferable to the number-average value for averaging the effects of electrostatic interactions. Analysis of published data on the proton dissociation of fulvic acid from the Suwannee River shows that the electrostatic interactions can be satisfactorily described by a hypothetical homogeneous compound having a molecular weight of 1000 (similar to the experimentally determined weight-average value). Titration data at three ionic strengths, for several fulvic acid concentrations, and in the pH range from 2.9 to 6.4 can be fitted with three adjustable parameters (pK??int values), given information on molecular size and carboxylate group content. ?? 1990 American Chemical Society.

  7. Direct molecular simulation of nitrogen dissociation based on an ab initio potential energy surface

    SciTech Connect

    Valentini, Paolo Schwartzentruber, Thomas E. Bender, Jason D. Nompelis, Ioannis Candler, Graham V.

    2015-08-15

    The direct molecular simulation (DMS) approach is used to predict the internal energy relaxation and dissociation dynamics of high-temperature nitrogen. An ab initio potential energy surface (PES) is used to calculate the dynamics of two interacting nitrogen molecules by providing forces between the four atoms. In the near-equilibrium limit, it is shown that DMS reproduces the results obtained from well-established quasiclassical trajectory (QCT) analysis, verifying the validity of the approach. DMS is used to predict the vibrational relaxation time constant for N{sub 2}–N{sub 2} collisions and its temperature dependence, which are in close agreement with existing experiments and theory. Using both QCT and DMS with the same PES, we find that dissociation significantly depletes the upper vibrational energy levels. As a result, across a wide temperature range, the dissociation rate is found to be approximately 4–5 times lower compared to the rates computed using QCT with Boltzmann energy distributions. DMS calculations predict a quasi-steady-state distribution of rotational and vibrational energies in which the rate of depletion of high-energy states due to dissociation is balanced by their rate of repopulation due to collisional processes. The DMS approach simulates the evolution of internal energy distributions and their coupling to dissociation without the need to precompute rates or cross sections for all possible energy transitions. These benchmark results could be used to develop new computational fluid dynamics models for high-enthalpy flow applications.

  8. Thermal dissociation of chemisorbed oxygen molecules on Ag(110): an investigation by scanning tunnelling microscopy

    NASA Astrophysics Data System (ADS)

    Zambelli, T.; Barth, J. V.; Wintterlin, J.

    2002-04-01

    The thermal dissociation of oxygen molecules on Ag(110) was studied by means of scanning tunnelling microscopy. At temperatures around 170 K a fraction of the molecules dissociate to form pairs of O atoms on the unreconstructed surface. The atoms lie in the trenches between the close-packed Ag rows; the intrapair distance is two lattice constants. The O atoms are very reactive with CO at temperatures as low as 70 K. The preferential orientation of the pairs in the [001] direction contrasts with the finding in a previous study (Hahn J R, Lee H J and Ho W 2000 Phys. Rev. Lett. Vol. 85 1914) that electron-induced dissociation exclusively leads to [110]-oriented pairs. The trajectories of thermal and electron-induced dissociations must therefore be different. In the temperature range of the dissociation, several additional oxygen features occur on the surface. These include unreacted O2 molecules, the first nuclei of the added-row reconstruction, and two unidentified O-induced structures.

  9. Antisymmetric tensor generalizations of affine vector fields

    PubMed Central

    Morisawa, Yoshiyuki; Tomoda, Kentaro

    2016-01-01

    Tensor generalizations of affine vector fields called symmetric and antisymmetric affine tensor fields are discussed as symmetry of spacetimes. We review the properties of the symmetric ones, which have been studied in earlier works, and investigate the properties of the antisymmetric ones, which are the main theme in this paper. It is shown that antisymmetric affine tensor fields are closely related to one-lower-rank antisymmetric tensor fields which are parallelly transported along geodesics. It is also shown that the number of linear independent rank-p antisymmetric affine tensor fields in n-dimensions is bounded by (n + 1)!/p!(n − p)!. We also derive the integrability conditions for antisymmetric affine tensor fields. Using the integrability conditions, we discuss the existence of antisymmetric affine tensor fields on various spacetimes. PMID:26858463

  10. Conformal field theory on affine Lie groups

    SciTech Connect

    Clubok, Kenneth Sherman

    1996-04-01

    Working directly on affine Lie groups, we construct several new formulations of the WZW model, the gauged WZW model, and the generic affine-Virasoro action. In one formulation each of these conformal field theories (CFTs) is expressed as a one-dimensional mechanical system whose variables are coordinates on the affine Lie group. When written in terms of the affine group element, this formulation exhibits a two-dimensional WZW term. In another formulation each CFT is written as a two-dimensional field theory, with a three- dimensional WZW term, whose fields are coordinates on the affine group. On the basis of these equivalent formulations, we develop a translation dictionary in which the new formulations on the affine Lie group are understood as mode formulations of the conventional formulations on the Lie group. Using this dictionary, we also express each CFT as a three-dimensional field theory on the Lie group with a four-dimensional WZW term. 36 refs.

  11. Dissociation and borderline personality disorder: an update for clinicians.

    PubMed

    Korzekwa, Marilyn I; Dell, Paul F; Pain, Clare

    2009-02-01

    Dissociation occurs in about two thirds of people with borderline personality disorder (BPD) but is still not well understood by clinicians. In the past decade, however, research has used new measures of dissociation that provide some of the detail that clinicians need to understand and treat the dissociative symptoms of patients with BPD. In particular, this review examines BPD's comorbidity with the dissociative disorders, the neurobiology of dissociation in BPD, the role of trauma and disorganized attachment in the etiology of dissociation in BPD, and the clinical assessment and treatment of dissociation in BPD.

  12. Efficient calculation of low energy statistical rates for gas phase dissociation using umbrella sampling

    NASA Astrophysics Data System (ADS)

    Mella, Massimo

    2006-03-01

    Monte Carlo (MC) simulations can be used to compute microcanonical statistical rates of gas phase dissociation reactions. Unfortunately, the MC approach may suffer from a slow convergence and large statistical errors for energies just above the dissociation threshold. In this work, umbrella sampling is proposed as a device to reduce the statistical error of MC rate constants. The method is tested by computing the classical dissociation rate for the reaction [H5O2+]*→H2O+H3O+ over the range of internal energy 38constants and classical Rice-Ramsperberg-Kassel harmonic theory shows that anharmonicity plays an important role in the dissociation process of the Zundel cation (H5O2+) at all energies.

  13. A Novel Vertex Affinity for Community Detection

    SciTech Connect

    Yoo, Andy; Sanders, Geoffrey; Henson, Van; Vassilevski, Panayot

    2015-10-05

    We propose a novel vertex affinity measure in this paper. The new vertex affinity quantifies the proximity between two vertices in terms of their clustering strength and is ideal for such graph analytics applications as community detection. We also developed a framework that combines simple graph searches and resistance circuit formulas to compute the vertex affinity efficiently. We study the properties of the new affinity measure empirically in comparison to those of other popular vertex proximity metrics. Our results show that the existing metrics are ill-suited for community detection due to their lack of fundamental properties that are essential for correctly capturing inter- and intra-cluster vertex proximity.

  14. Structural determinants of sigma receptor affinity

    SciTech Connect

    Largent, B.L.; Wikstroem, H.G.; Gundlach, A.L.; Snyder, S.H.

    1987-12-01

    The structural determinants of sigma receptor affinity have been evaluated by examining a wide range of compounds related to opioids, neuroleptics, and phenylpiperidine dopaminergic structures for affinity at sigma receptor-binding sites labeled with (+)-(/sup 3/H)3-PPP. Among opioid compounds, requirements for sigma receptor affinity differ strikingly from the determinants of affinity for conventional opiate receptors. Sigma sites display reverse stereoselectivity to classical opiate receptors. Multi-ringed opiate-related compounds such as morphine and naloxone have negligible affinity for sigma sites, with the highest sigma receptor affinity apparent for benzomorphans which lack the C ring of opioids. Highest affinity among opioids and other compounds occurs with more lipophilic N-substituents. This feature is particularly striking among the 3-PPP derivatives as well as the opioids. The butyrophenone haloperidol is the most potent drug at sigma receptors we have detected. Among the series of butyrophenones, receptor affinity is primarily associated with the 4-phenylpiperidine moiety. Conformational calculations for various compounds indicate a fairly wide range of tolerance for distances between the aromatic ring and the amine nitrogen, which may account for the potency at sigma receptors of structures of considerable diversity. Among the wide range of structures that bind to sigma receptor-binding sites, the common pharmacophore associated with high receptor affinity is a phenylpiperidine with a lipophilic N-substituent.

  15. Constant domain-regulated antibody catalysis.

    PubMed

    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.

  16. Base-sequence specificity of Hoechst 33258 and DAPI binding to five (A/T)4 DNA sites with kinetic evidence for more than one high-affinity Hoechst 33258-AATT complex.

    PubMed

    Breusegem, Sophia Y; Clegg, Robert M; Loontiens, Frank G

    2002-02-01

    The binding of Hoechst 33258 and DAPI to five different (A/T)4 sequences in a stable DNA hairpin was studied exploiting the substantial increase in dye fluorescence upon binding. The two dyes have comparable affinities for the AATT site (e.g. association constant K(a)=5.5 x 10(8) M(-1) for DAPI), and their affinities decrease in the series AATT > TAAT approximately equal to ATAT > TATA approximately equal to TTAA. The extreme values of K(a) differ by a factor of 200 for Hoechst 33258 but only 30 for DAPI. The binding kinetics of Hoechst 33258 were measured by stopped-flow under pseudo-first order conditions with an (A/T)4 site in excess. The lower-resolution experiments can be well represented by single exponential processes, corresponding to a single-step binding mechanism. The calculated association-rate parameters for the five (A/T)4 sites are similar (2.46 x 10(8) M(-1) s(-1) to 0.86 x 10(8) M(-1) s(-1)) and nearly diffusion-controlled, while the dissociation-rate parameters vary from 0.42 s(-1) to 96 s(-1). Thus the association constants are kinetically controlled and are close to their equilibrium-determined values. However, when obtained with increased signal-to-noise ratio, the kinetic traces for Hoechst 33258 binding at the AATT site reveal two components. The concentration dependencies of the two time constants and amplitudes are consistent with two different kinetically equivalent two-step models. In the first model, fast bimolecular binding is followed by an isomerization of the initial complex. In the second model, two single-step associations form two complexes that mutually exclude each other. For both models the four reaction-rate parameters are calculated. Finally, specific dissociation kinetics, using poly[d(A-5BrU)], show that the kinetics are even more complex than either two-step model. We correlate our results with the different binding orientations and locations of Hoechst 33258 in the DNA minor groove found in several structural studies in

  17. Dissociations between developmental dyslexias and attention deficits

    PubMed Central

    Lukov, Limor; Friedmann, Naama; Shalev, Lilach; Khentov-Kraus, Lilach; Shalev, Nir; Lorber, Rakefet; Guggenheim, Revital

    2014-01-01

    We examine whether attention deficits underlie developmental dyslexia, or certain types of dyslexia, by presenting double dissociations between the two. We took into account the existence of distinct types of dyslexia and of attention deficits, and focused on dyslexias that may be thought to have an attentional basis: letter position dyslexia (LPD), in which letters migrate within words, attentional dyslexia (AD), in which letters migrate between words, neglect dyslexia, in which letters on one side of the word are omitted or substituted, and surface dyslexia, in which words are read via the sublexical route. We tested 110 children and adults with developmental dyslexia and/or attention deficits, using extensive batteries of reading and attention. For each participant, the existence of dyslexia and the dyslexia type were tested using reading tests that included stimuli sensitive to the various dyslexia types. Attention deficit and its type was established through attention tasks assessing sustained, selective, orienting, and executive attention functioning. Using this procedure, we identified 55 participants who showed a double dissociation between reading and attention: 28 had dyslexia with normal attention and 27 had attention deficits with normal reading. Importantly, each dyslexia with suspected attentional basis dissociated from attention: we found 21 individuals with LPD, 13 AD, 2 neglect dyslexia, and 12 surface dyslexia without attention deficits. Other dyslexia types (vowel dyslexia, phonological dyslexia, visual dyslexia) also dissociated from attention deficits. Examination of 55 additional individuals with both a specific dyslexia and a certain attention deficit found no attention function that was consistently linked with any dyslexia type. Specifically, LPD and AD dissociated from selective attention, neglect dyslexia dissociated from orienting, and surface dyslexia dissociated from sustained and executive attention. These results indicate that

  18. Dissociations between developmental dyslexias and attention deficits.

    PubMed

    Lukov, Limor; Friedmann, Naama; Shalev, Lilach; Khentov-Kraus, Lilach; Shalev, Nir; Lorber, Rakefet; Guggenheim, Revital

    2014-01-01

    We examine whether attention deficits underlie developmental dyslexia, or certain types of dyslexia, by presenting double dissociations between the two. We took into account the existence of distinct types of dyslexia and of attention deficits, and focused on dyslexias that may be thought to have an attentional basis: letter position dyslexia (LPD), in which letters migrate within words, attentional dyslexia (AD), in which letters migrate between words, neglect dyslexia, in which letters on one side of the word are omitted or substituted, and surface dyslexia, in which words are read via the sublexical route. We tested 110 children and adults with developmental dyslexia and/or attention deficits, using extensive batteries of reading and attention. For each participant, the existence of dyslexia and the dyslexia type were tested using reading tests that included stimuli sensitive to the various dyslexia types. Attention deficit and its type was established through attention tasks assessing sustained, selective, orienting, and executive attention functioning. Using this procedure, we identified 55 participants who showed a double dissociation between reading and attention: 28 had dyslexia with normal attention and 27 had attention deficits with normal reading. Importantly, each dyslexia with suspected attentional basis dissociated from attention: we found 21 individuals with LPD, 13 AD, 2 neglect dyslexia, and 12 surface dyslexia without attention deficits. Other dyslexia types (vowel dyslexia, phonological dyslexia, visual dyslexia) also dissociated from attention deficits. Examination of 55 additional individuals with both a specific dyslexia and a certain attention deficit found no attention function that was consistently linked with any dyslexia type. Specifically, LPD and AD dissociated from selective attention, neglect dyslexia dissociated from orienting, and surface dyslexia dissociated from sustained and executive attention. These results indicate that

  19. Development of new peptide-based receptor of fluorescent probe with femtomolar affinity for Cu(+) and detection of Cu(+) in Golgi apparatus.

    PubMed

    Jung, Kwan Ho; Oh, Eun-Taex; Park, Heon Joo; Lee, Keun-Hyeung

    2016-11-15

    Developing fluorescent probes for monitoring intracellular Cu(+) is important for human health and disease, whereas a few types of their receptors showing a limited range of binding affinities for Cu(+) have been reported. In the present study, we first report a novel peptide receptor of a fluorescent probe for the detection of Cu(+). Dansyl-labeled tripeptide probe (Dns-LLC) formed a 1:1 complex with Cu(+) and showed a turn-on fluorescent response to Cu(+) in aqueous buffered solutions. The dissociation constant of Dns-LLC for Cu(+) was determined to be 12 fM, showing that Dns-LLC had more potent binding affinity for Cu(+) than those of previously reported chemical probes for Cu(+). The binding mode study showed that the thiol group of the peptide receptor plays a critical role in potent binding with Cu(+) and the sulfonamide and amide groups of the probe might cooperate to form a complex with Cu(+). Dns-LLC detected Cu(+) selectively by a turn-on response among various biologically relevant metal ions, including Cu(2+) and Zn(2+). The selectivity of the peptide-based probe for Cu(+) was strongly dependent on the position of the cysteine residue in the peptide receptor part. The fluorescent peptide-based probe penetrated the living RKO cells and successfully detected Cu(+) in the Golgi apparatus in live cells by a turn-on response. Given the growing interest in imaging Cu(+) in live cells, a novel peptide receptor of Cu(+) will offer the potential for developing a variety of fluorescent probes for Cu(+) in the field of copper biochemistry.

  20. Combinatorial de novo design and application of a biomimetic affinity ligand for the purification of human anti-HIV mAb 4E10 from transgenic tobacco.

    PubMed

    Platis, Dimitris; Maltezos, Anastasios; Ma, Julian K-C; Labrou, Nikolaos E

    2009-01-01

    Monoclonal anti-HIV antibody 4E10 (mAb 4E10) is one of the most broadly neutralizing antibodies against HIV, directed against a specific epitope on envelope protein gp41. In the present study, a combinatorial de novo design approach was used for the development of a biomimetic ligand for the affinity purification of mAb 4E10 from tobacco transgenic extract in a single chromatographic step. The biomimetic ligand (4E10lig) was based on a L-Phe/beta-Ala bi-substituted 1,3,5-triazine (Trz) scaffold (beta-Ala-Trz-L-Phe, 4E10lig) which potentially mimics the more pronounced electrostatic and hydrophobic interactions of mAb 4E10-binding sequence determined by screening of a random peptide library. This library was comprised of Escherichia coli cells harboring a plasmid (pFlitrx) engineered to express a fusion protein containing random dodecapeptides that were inserted into the active loop of thioredoxin, which itself was inserted into the dispensable region of the flagellin gene. Adsorption equilibrium studies with this biomimetic ligand and mAb 4E10 determined a dissociation constant (K(D)) of 0.41 +/- 0.05 microM. Molecular modeling studies of the biomimetic ligand revealed that it can potentially occupy the same binding site as the natural binding core peptide epitope. The biomimetic affinity adsorbent was exploited in the development of a facile mAb 4E10 purification protocol, affording mAb 4E10 of high purity (approximately 95%) with good overall yield (60-80%). Analysis of the antibody preparation by SDS-PAGE, enzyme-linked immunosorbent assays (ELISA), and western blot showed that the mAb 4E10 was fully active and free of degraded variants, polyphenols, and alkaloids.

  1. Dissociative experiences in epilepsy: effects of epilepsy-related factors on pathological dissociation.

    PubMed

    Hara, Koichiro; Adachi, Naoto; Akanuma, Nozomi; Ito, Masumi; Okazaki, Mitsutoshi; Matsubara, Ryoji; Adachi, Takuya; Ishii, Ryouhei; Kanemoto, Kousuke; Matsuura, Masato; Hara, Eriko; Kato, Masaaki; Onuma, Teiichi

    2015-03-01

    Psychogenic nonepileptic seizures (PNESs) in patients with epilepsy can be categorized as dissociative disorders. The prevalence of PNESs in patients with epilepsy appears to be much higher than that of dissociative experiences in nonclinical subjects. In order to clarify as to whether epilepsy-related factors were associated with pathological dissociation, we conducted a controlled study with 225 patients with epilepsy and 334 nonclinically matched individuals. All participants completed the Japanese version of the Dissociative Experiences Scale (DES). There was no significant difference in the DES score (DES-S) between the group with epilepsy and the control group. The group with epilepsy showed a significantly higher DES taxon (DES-T; a subset of DES-S and an index of pathological dissociation) than the control group. Thirty-one out of the 225 patients with epilepsy (13.8%) had PNESs. Because of its strong association with the DES-S and DES-T, PNESs can be regarded as a symptom of dissociation. With multiple regression analysis, the patients with a shorter duration of epilepsy, higher seizure frequency, or shorter period in education tend to suffer from pathological dissociation. These findings demonstrate that patients with epilepsy are more prone to experiencing pathological dissociation when having certain clinical factors.

  2. Is the dissociative adult suggestible? A test of the trauma and fantasy models of dissociation.

    PubMed

    Kluemper, Nicole S; Dalenberg, Constance

    2014-01-01

    Psychologists have long assumed a connection between traumatic experience and psychological dissociation. This hypothesis is referred to as the trauma model of dissociation. In the past decade, a series of papers have been published that question this traditional causal link, proposing an alternative fantasy model of dissociation. In the present research, the relationship among dissociation, suggestibility, and fantasy proneness was examined. Suggestibility was measured through the Gudjonsson Scale of Interrogative Suggestibility (GSS) as well as an autobiographically based version of this measure based on the events of September 11, 2001. Consistent with prior research and with the trauma model, dissociation correlated positively with trauma severity (r = .32, p < .01) and fantasy proneness (r = .60, p < .01). Inconsistent with the fantasy model, dissociation did not correlate with the neutral form of the GSS and correlated negatively (r = -.24, p < .05) with the trauma-focused form of this suggestibility measure. Although some participants did become quite emotional during the procedure, the risk/benefit ratio was perceived by almost all participants to be positive, with more reactive individuals evaluating the procedure more positively. The results consistently support the trauma model of dissociation and fail to support the fantasy model of dissociation.

  3. Dissociation in borderline personality disorder: a detailed look.

    PubMed

    Korzekwa, Marilyn I; Dell, Paul F; Links, Paul S; Thabane, Lehana; Fougere, Philip

    2009-01-01

    The objective of the present study was to assess in detail the whole spectrum of normal and pathological dissociative experiences and dissociative disorder (DD) diagnoses in borderline personality disorder (BPD) as diagnosed with the Revised Diagnostic Interview for Borderlines. Dissociation was measured comprehensively in 21 BPD outpatients using the Structured Clinical Interview for DSM-IV Dissociative Disorders-Revised, the Multidimensional Inventory of Dissociation (MID), the Dissociative Experiences Scale pathological taxon analysis, and the Somatoform Dissociation Questionnaire. The frequencies of DDs in this BPD sample were as follows: 24% no DD, 29% mild DD (dissociative amnesia and depersonalization disorder), 24% DD Not Otherwise Specified (DDNOS), and 24% dissociative identity disorder. With regard to the dissociative experiences endorsed, almost all patients reported identity confusion, unexplained mood changes, and depersonalization. Even those BPD patients with mild DD reported derealization, depersonalization, and dissociative amnesia. BPD patients with DDNOS reported frequent depersonalization, frequent amnesia, and notable experiences of identity alteration. BPD patients with dissociative identity disorder endorsed severe dissociative symptoms in all categories. Analysis of the MID pathological dissociation items revealed that 32% of the items were endorsed at a clinically significant level of frequency by more than 50% of our BPD patients. In conclusion, the frequencies of Diagnostic and Statistical Manual of Mental Disorders (4th ed.) DDs in these patients with BPD were surprisingly high. Likewise, the "average" BPD patient endorsed a wide variety of recurrent pathological dissociative symptoms.

  4. Dissociative electron attachment to C{sub 2}F{sub 5} radicals

    SciTech Connect

    Haughey, Sean A.; Field, Thomas A.; Langer, Judith; Shuman, Nicholas S.; Miller, Thomas M.; Friedman, Jeffrey F.; Viggiano, A. A.

    2012-08-07

    Dissociative electron attachment to the reactive C{sub 2}F{sub 5} molecular radical has been investigated with two complimentary experimental methods; a single collision beam experiment and a new flowing afterglow Langmuir probe technique. The beam results show that F{sup -} is formed close to zero electron energy in dissociative electron attachment to C{sub 2}F{sub 5}. The afterglow measurements also show that F{sup -} is formed in collisions between electrons and C{sub 2}F{sub 5} molecules with rate constants of 3.7 Multiplication-Sign 10{sup -9} cm{sup 3} s{sup -1} to 4.7 Multiplication-Sign 10{sup -9} cm{sup 3} s{sup -1} at temperatures of 300-600 K. The rate constant increases slowly with increasing temperature, but the rise observed is smaller than the experimental uncertainty of 35%.

  5. The contribution of the methyl groups on thymine bases to binding specificity and affinity by alanine-rich mutants of the bZIP motif.

    PubMed

    Kise, K J; Shin, J A

    2001-09-01

    We have used fluorescence anisotropy to measure in situ the thermodynamics of binding of alanine-rich mutants of the GCN4 basic region/leucine zipper (bZIP) to short DNA duplexes, in which thymines were replaced with uracils, in order to quantify the contributions of the C5 methyl group on thymines with alanine methyl side chains. We simplified the alpha-helical GCN4 bZIP by alanine substitution: 4A, 11A, and 18A contain four, 11, and 18 alanine mutations in their DNA-binding basic regions, respectively. Titration of fluorescein-labeled duplexes with increasing amounts of protein yielded dissociation constants in the low-to-mid nanomolar range for all bZIP mutants in complex with the AP-1 target site (5'-TGACTCA-3'); binding to the nonspecific control duplex was >1000-fold weaker. Small changes of <1 kcal/mol in binding free energies were observed for wild-type bZIP and 4A mutant to uracil-containing AP-1, whereas 11A and 18A bound almost equally well to native AP-1 and uracil-containing AP-1. These modest changes in binding affinities may reflect the multivalent nature of protein-DNA interactions, as our highly mutated proteins still exhibit native-like behavior. These protein mutations may compensate for changes in enthalpic and entropic contributions toward DNA-binding in order to maintain binding free energies similar to that of the native protein-DNA complex.

  6. Rovibrational energy transfer and dissociation in O2-O collisions.

    PubMed

    Andrienko, Daniil A; Boyd, Iain D

    2016-03-14

    A set of state-specific transition rates for each rovibrational level is generated for the O2(X(3)Σ(g)(-))-O(3)P system using the quasi-classical trajectory method at temperatures observed in hypersonic flows. A system of master equations describes the relaxation of the rovibrational ensemble to thermal equilibrium under ideal heat bath conditions at a constant translational temperature. Vibrational and rotational relaxation times, obtained from the average internal energies, exhibit a pattern inherent in a chemically reactive collisional pair. An intrinsic feature of the O3 molecular system with a large attractive potential is a weak temperature dependence of the rovibrational transition rates. For this reason, the quasi-steady vibrational and rotational temperatures experience a maximum at increasing translational temperature. The energy rate coefficients, that characterize the average loss of internal energy due to dissociation, quickly diminish at high temperatures, compared to other molecular systems.

  7. Rovibrational energy transfer and dissociation in O2-O collisions

    NASA Astrophysics Data System (ADS)

    Andrienko, Daniil A.; Boyd, Iain D.

    2016-03-01

    A set of state-specific transition rates for each rovibrational level is generated for the O 2 ( X 3 Σg - ) - O (" separators=" 3 P ) system using the quasi-classical trajectory method at temperatures observed in hypersonic flows. A system of master equations describes the relaxation of the rovibrational ensemble to thermal equilibrium under ideal heat bath conditions at a constant translational temperature. Vibrational and rotational relaxation times, obtained from the average internal energies, exhibit a pattern inherent in a chemically reactive collisional pair. An intrinsic feature of the O3 molecular system with a large attractive potential is a weak temperature dependence of the rovibrational transition rates. For this reason, the quasi-steady vibrational and rotational temperatures experience a maximum at increasing translational temperature. The energy rate coefficients, that characterize the average loss of internal energy due to dissociation, quickly diminish at high temperatures, compared to other molecular systems.

  8. Dissociation energies of some high temperature molecules containing aluminum

    NASA Technical Reports Server (NTRS)

    Stearns, C. A.; Kohl, F. J.

    1972-01-01

    The Knudsen cell mass spectrometric method has been used to investigate the gaseous molecules Al2, AlSi,AlSiO, AlC2, Al2C2, and AlAuC2. Special attention was given to the experimental considerations and techniques needed to identify and to measure ion intensities for very low abundance molecular species. Second- and third-law procedures were used to obtain reaction enthalpies for pressure calibration independent and isomolecular exchange reactions. Dissociation energies for the molecules were derived from the measured ion intensities, free-energy functions obtained from estimated molecular constants, and auxiliary thermodynamic data. The bonding and stability of these aluminum containing molecules are compared with other similar species.

  9. Dissociative recombination coefficient for low temperature equilibrium cesium plasma

    NASA Astrophysics Data System (ADS)

    Momozaki, Yoichi; El-Genk, Mohamed S.

    2002-07-01

    The dissociative recombination (DR) coefficient in decaying low temperature Cs plasma is calculated based on the experimentally measured relaxation time of decaying Cs plasma by L. P. Harris [J. Appl. Phys. 36, 1543 (1965)]. Results showed that DR is the dominant recombination process over three-body recombination at T<1650 K and PCs of 0.5-20 Torr (67-2666 Pa). The estimated DR coefficient for Cs is between 10-12 and 10-13 m3/s at T<1750 K and PCs of 0.5-20 Torr. Although theory predicts that DR coefficient solely depends on temperature, the present results show pressure dependency. For typical operating conditions in thermionic converters (T<1650 K and PCsless-than-or-equal400 Pa), DR is constant and approx5.26 x10-13 m3/s.

  10. Electron capture dissociation in a digital ion trap mass spectrometer.

    PubMed

    Ding, Li; Brancia, Francesco L

    2006-03-15

    Electron capture dissociation was implemented in a digital ion trap without using any magnetic field to focus the electrons. Since rectangular waveforms are employed in the DIT for both trapping and dipole excitation, electrons can be injected into the trap when the electric field is constant. Following deceleration, electrons reach the precursor ion cloud. The fragment ions produced by interactions with the electron beam are subsequently analyzed by resonant ejection. [Glu(1)]-Fibrinopeptide B and substance P were used to evaluate the performance of the current design. Fragmentation efficiency of 5.5% was observed for substance P peptide ions. Additionally, analysis of the monophosphorylated peptide FQ[pS]EEQQQTEDELQDK shows that in the resulting c- and z-type ions, the phosphate group is retained on the phophoserine residue, providing information on which amino acid residue the modification is located.

  11. Dissociation of cerium(III) and neodymium(III) phthalocyanines

    NASA Astrophysics Data System (ADS)

    Lomova, T. N.

    2015-07-01

    The kinetics of dissociation of phthalocyanine complexes with cerium(III) and neodymium(III) (X)LnPc (X = Cl-, Br-, AcO-) under the action of acetic acid in ethanol with isolation of the macrocyclic ligand depending on the temperature was studied. The kinetic equations with the numerical values of rate constants, activation parameters, and the stoichiometric mechanisms with the limiting simple reaction between the nonionized AcOH molecule and (phthalocyaninato)lanthanide(III) in the axially coordinated ((X)LnPc, cerium complexes) or axially ionized ([(AcOH)LnPc]+X-, neodymium complexes) state were derived by solving the direct and inverse problems. As shown by a comparative analysis of quantitative kinetic data, the state is determined by the electronic structure of the metal cation and the mutual effect of the axial and equatorial ligands in the first coordination sphere.

  12. Structure of classical affine and classical affine fractional W-algebras

    SciTech Connect

    Suh, Uhi Rinn

    2015-01-15

    We introduce a classical BRST complex (See Definition 3.2.) and show that one can construct a classical affine W-algebra via the complex. This definition clarifies that classical affine W-algebras can be considered as quasi-classical limits of quantum affine W-algebras. We also give a definition of a classical affine fractional W-algebra as a Poisson vertex algebra. As in the classical affine case, a classical affine fractional W-algebra has two compatible λ-brackets and is isomorphic to an algebra of differential polynomials as a differential algebra. When a classical affine fractional W-algebra is associated to a minimal nilpotent, we describe explicit forms of free generators and compute λ-brackets between them. Provided some assumptions on a classical affine fractional W-algebra, we find an infinite sequence of integrable systems related to the algebra, using the generalized Drinfel’d and Sokolov reduction.

  13. Dissociating motor cortex from the motor

    PubMed Central

    Schieber, Marc H

    2011-01-01

    Abstract During closed-loop control of a brain–computer interface, neurons in the primary motor cortex can be intensely active even though the subject may be making no detectable movement or muscle contraction. How can neural activity in the primary motor cortex become dissociated from the movements and muscles of the native limb that it normally controls? Here we examine circumstances in which motor cortex activity is known to dissociate from movement – including mental imagery, visuo-motor dissociation and instructed delay. Many such motor cortex neurons may be related to muscle activity only indirectly. Furthermore, the integration of thousands of synaptic inputs by individual α-motoneurons means that under certain circumstances even cortico-motoneuronal cells, which make monosynaptic connections to α-motoneurons, can become dissociated from muscle activity. The natural ability of motor cortex neurons under voluntarily control to become dissociated from bodily movement may underlie the utility of this cortical area for controlling brain–computer interfaces. PMID:22005673

  14. Isotope separation by photoselective dissociative electron capture

    DOEpatents

    Stevens, Charles G. [Pleasanton, CA

    1978-08-29

    A method of separating isotopes based on photoselective electron capture dissociation of molecules having an electron capture cross section dependence on the vibrational state of the molecule. A molecular isotope source material is irradiated to selectively excite those molecules containing a desired isotope to a predetermined vibrational state having associated therewith an electron capture energy region substantially non-overlapping with the electron capture energy ranges associated with the lowest vibration states of the molecules. The isotope source is also subjected to electrons having an energy corresponding to the non-overlapping electron capture region whereby the selectively excited molecules preferentially capture electrons and dissociate into negative ions and neutrals. The desired isotope may be in the negative ion product or in the neutral product depending upon the mechanism of dissociation of the particular isotope source used. The dissociation product enriched in the desired isotope is then separated from the reaction system by conventional means. Specifically, .sup.235 UF.sub.6 is separated from a UF.sub.6 mixture by selective excitation followed by dissociative electron capture into .sup.235 UF.sub.5 - and F.

  15. Isotope separation by photoselective dissociative electron capture

    DOEpatents

    Stevens, C.G.

    1978-08-29

    Disclosed is a method of separating isotopes based on photoselective electron capture dissociation of molecules having an electron capture cross section dependence on the vibrational state of the molecule. A molecular isotope source material is irradiated to selectively excite those molecules containing a desired isotope to a predetermined vibrational state having associated therewith an electron capture energy region substantially non-overlapping with the electron capture energy ranges associated with the lowest vibration states of the molecules. The isotope source is also subjected to electrons having an energy corresponding to the non-overlapping electron capture region whereby the selectively excited molecules preferentially capture electrons and dissociate into negative ions and neutrals. The desired isotope may be in the negative ion product or in the neutral product depending upon the mechanism of dissociation of the particular isotope source used. The dissociation product enriched in the desired isotope is then separated from the reaction system by conventional means. Specifically, [sup 235]UF[sub 6] is separated from a UF[sub 6] mixture by selective excitation followed by dissociative electron capture into [sup 235]UF[sub 5]- and F. 2 figs.

  16. Ion-induced dissociation dynamics of acetylene

    SciTech Connect

    De, Sankar; Rajput, Jyoti; Roy, A.; Safvan, C. P.; Ghosh, P. N.

    2008-02-15

    We report on the results of dissociation dynamics of multiple charged acetylene molecules formed in collision with 1.2 MeV Ar{sup 8+} projectiles. Using the coincidence map, we can separate out the different dissociation pathways between carbon and hydrogen ionic fragments as well as complete two-body breakup events. From the measured slopes of the coincidence islands for carbon atomic fragments and theoretical values determined from the charge and momentum distribution of the correlated particles, we observe a diatom like behavior of the C-C charged complex during dissociation of multiply charged C{sub 2}H{sub 2}. We conclude that this behavior in breakup dynamics is a signature of sequentiality in dissociation of this multiply charged molecular species. The shape and orientation of the islands give further information about the momentum balance in the fragmentation process of two- or many-body dissociation pathways. Kinetic energy release of different breakup channels are reported here and compared with values calculated from the pure Coulomb explosion model.

  17. Affinity capillary electrophoresis: the theory of electromigration.

    PubMed

    Dubský, Pavel; Dvořák, Martin; Ansorge, Martin

    2016-12-01

    We focus on the state-of-the-art theory of electromigration under single and multiple complexation equilibrium. Only 1:1 complexation stoichiometry is discussed because of its unique status in the field of affinity capillary electrophoresis (ACE). First, we summarize the formulas for the effective mobility in various ACE systems as they appeared since the pioneering days in 1992 up to the most recent theories till 2015. Disturbing phenomena that do not alter the mobility of the analyte directly but cause an unexpected peak broadening have been studied only recently and are also discussed in this paper. Second, we turn our attention to the viscosity effects in ACE. Change in the background electrolyte viscosity is unavoidable in ACE but numerous observations scattered throughout the literature have not been reviewed previously. This leads to an uncritical employment of correction factors that may or may not be appropriate in practice. Finally, we consider the ionic strength effects in ACE, too. Limitations of the current theories are also discussed and the tasks identified where open problems still prevail. Graphical Abstract A weak base (A) undergoes an acidic-basic equilibria (in blue) and migrates with an electrophoretic mobility of [Formula: see text]. Simultaneously, it interacts with a selector (sel) while the analyte-selector complex migrates with an electrophoretic mobility of [Formula: see text]. The strength of the interaction (in orange) is governed by the binding constant, K A , and the concentration of the selector, c sel . This all gives the analyte an effective mobility of [Formula: see text] and moves it out of the zero position (EOF; right top insert). The interaction of the positively charged analyte with the neutral selector slows down the analyte with increasing selector concentration (right bottom insert).

  18. Simulating the dissociation of CO/sub 2/ in a supersonic nonequilibrium plasma flow

    SciTech Connect

    Grigor'eva, T.M.; Fridman, A.A.; Levitskii, A.A.; Polak, L.S.; Potapkin, B.V.; Rusanov, V.D.

    1986-11-01

    Gas-dynamic and chemical-kinetic equations have been used in simulating CO/sub 2/ dissociation in a supersonic flow of nonequilibrium plasma. The energy performance and the degree of conversion of CO/sub 2/ have been related to the specific energy deposition in the vibrational degrees of freedom. It is shown that the degree of conversion may attain about 30% in a constant-pressure reactor with comparatively small expansion angles.

  19. Is Planck's quantization constant unique?

    NASA Astrophysics Data System (ADS)

    Livadiotis, George

    2016-07-01

    A cornerstone of Quantum Mechanics is the existence of a non-zero least action, the Planck constant. However, the basic concepts and theoretical developments of Quantum Mechanics are independent of its specific numerical value. A different constant h _{*}, similar to the Planck constant h, but ˜12 orders of magnitude larger, characterizes plasmas. The study of >50 different geophysical, space, and laboratory plasmas, provided the first evidence for the universality and the quantum nature of h _{*}, revealing that it is a new quantization constant. The recent results show the diagnostics for determining whether plasmas are characterized by the Planck or the new quantization constant, compounding the challenge to reconcile both quantization constants in quantum mechanics.

  20. Isolation and partial characterization of Bromelia hemisphaerica protease by affinity chromatography.

    PubMed

    Ochoa, N; Agundis, C; Córdoba, F

    1987-01-01

    Hemisphaericin, the protease from Bromelia hemisphaerica fruit juice was isolated by affinity chromatography in one step, using a mercurial sepharose derivative. The enzyme behaves as a single component in immunodifussion, immunoelectrophoresis and polyacrylamide electrophoresis in the presence of SDS and 2-mercaptoethanol. Association and dissociation of active components were evidenced in electrophoresis at pH 3.6 and at pH 8.6. Immunoelectrophoresis analyses also disclosed a certain degree of internal immunological heterogeneity. The results are explained by the presence of an enzyme subunit, of about 8000 daltons, endowed with polymeric properties induced by the pH and oxidative environment.

  1. Differential Effects of TR Ligands on Hormone Dissociation Rates: Evidence for Multiple Ligand Entry/Exit Pathways

    PubMed Central

    Lima, Suzana T. Cunha; Nguyen, Ngoc-Ha; Togashi, Marie; Apriletti, James W.; Nguyen, Phuong; Polikarpov, Igor; Scanlan, Thomas S.; Baxter, John D.; Webb, Paul

    2009-01-01

    Some nuclear receptor (NR) ligands promote dissociation of radiolabeled bound hormone from the buried ligand binding cavity (LBC) more rapidly than excess unlabeled hormone itself. This result was interpreted to mean that challenger ligands bind allosteric sites on the LBD to induce hormone dissociation, and recent findings indicate that ligands bind weakly to multiple sites on the LBD surface. Here, we show that a large fraction of thyroid hormone receptor (TR) ligands promote rapid dissociation (T1/2 <2 hours) of radiolabeled T3 versus T3 (T1/2 ≈5–7 hours). We cannot discern relationships between this effect and ligand size, activity or affinity for TRβ. One ligand, GC-24, binds the TR LBC and (weakly) to the TRβ-LBD surface that mediates dimer/heterodimer interaction, but we cannot link this interaction to rapid T3 dissociation. Instead, several lines of evidence suggest that the challenger ligand must interact with the buried LBC to promote rapid T3 release. Since previous molecular dynamics simulations suggest that TR ligands leave the LBC by several routes, we propose that a subset of challenger ligands binds and stabilizes a partially unfolded intermediate state of TR that arises during T3 release and that this effect enhances hormone dissociation. PMID:19729063

  2. Dissociation in virtual reality: depersonalization and derealization

    NASA Astrophysics Data System (ADS)

    Garvey, Gregory P.

    2010-01-01

    This paper looks at virtual worlds such as Second Life7 (SL) as possible incubators of dissociation disorders as classified by the Diagnostic and Statistical Manual of Mental Disorders, 4th Edition3 (also known as the DSM-IV). Depersonalization is where "a person feels that he or she has changed in some way or is somehow unreal." Derealization when "the same beliefs are held about one's surroundings." Dissociative Identity Disorder (DID), previously known as multiple personality disorder fits users of Second Life who adopt "in-world" avatars and in effect, enact multiple distinct identities or personalities (known as alter egos or alters). Select questions from the Structured Clinical Interview for Depersonalization (SCI-DER)8 will be discussed as they might apply to the user's experience in Second Life. Finally I would like to consider the hypothesis that rather than a pathological disorder, dissociation is a normal response to the "artificial reality" of Second Life.

  3. Psychotherapy and pharmacotherapy for patients with dissociative identity disorder.

    PubMed

    Gentile, Julie P; Dillon, Kristy S; Gillig, Paulette Marie

    2013-02-01

    There is a wide variety of what have been called "dissociative disorders," including dissociative amnesia, dissociative fugue, depersonalization disorder, dissociative identity disorder, and forms of dissociative disorder not otherwise specified. Some of these diagnoses, particularly dissociative identity disorder, are controversial and have been questioned by many clinicians over the years. The disorders may be under-diagnosed or misdiagnosed, but many persons who have experienced trauma report "dissociative" symptoms. Prevalence of dissociative disorders is unknown, but current estimates are higher than previously thought. This paper reviews clinical, phenomenological, and epidemiological data regarding diagnosis in general, and illustrates possible treatment interventions for dissociative identity disorder, with a focus on psychotherapy interventions and a review of current psychopharmacology recommendations as part of a comprehensive multidisciplinary treatment plan.

  4. Improving image segmentation by learning region affinities

    SciTech Connect

    Prasad, Lakshman; Yang, Xingwei; Latecki, Longin J

    2010-11-03

    We utilize the context information of other regions in hierarchical image segmentation to learn new regions affinities. It is well known that a single choice of quantization of an image space is highly unlikely to be a common optimal quantization level for all categories. Each level of quantization has its own benefits. Therefore, we utilize the hierarchical information among different quantizations as well as spatial proximity of their regions. The proposed affinity learning takes into account higher order relations among image regions, both local and long range relations, making it robust to instabilities and errors of the original, pairwise region affinities. Once the learnt affinities are obtained, we use a standard image segmentation algorithm to get the final segmentation. Moreover, the learnt affinities can be naturally unutilized in interactive segmentation. Experimental results on Berkeley Segmentation Dataset and MSRC Object Recognition Dataset are comparable and in some aspects better than the state-of-art methods.

  5. Structure-Based Development of an Affinity Probe for Sirtuin 2.

    PubMed

    Schiedel, Matthias; Rumpf, Tobias; Karaman, Berin; Lehotzky, Attila; Gerhardt, Stefan; Ovádi, Judit; Sippl, Wolfgang; Einsle, Oliver; Jung, Manfred

    2016-02-05

    Sirtuins are NAD(+)-dependent protein deacylases that cleave off acetyl groups, as well as other acyl groups, from the ɛ-amino group of lysines in histones and other substrate proteins. Dysregulation of human Sirt2 activity has been associated with the pathogenesis of cancer, inflammation, and neurodegeneration, thus making Sirt2 a promising target for pharmaceutical intervention. Here, based on a crystal structure of Sirt2 in complex with an optimized sirtuin rearranging ligand (SirReal) that shows improved potency, water solubility, and cellular efficacy, we present the development of the first Sirt2-selective affinity probe. A slow dissociation of the probe/enzyme complex offers new applications for SirReals, such as biophysical characterization, fragment-based screening, and affinity pull-down assays. This possibility makes the SirReal probe an important tool for studying sirtuin biology.

  6. Kinetics of Propargyl Radical Dissociation.

    PubMed

    Klippenstein, Stephen J; Miller, James A; Jasper, Ahren W

    2015-07-16

    Due to the prominent role of the propargyl radical for hydrocarbon growth within combustion environments, it is important to understand the kinetics of its formation and loss. The ab initio transition state theory-based master equation method is used to obtain theoretical kinetic predictions for the temperature and pressure dependence of the thermal decomposition of propargyl, which may be its primary loss channel under some conditions. The potential energy surface for the decomposition of propargyl is first mapped at a high level of theory with a combination of coupled cluster and multireference perturbation calculations. Variational transition state theory is then used to predict the microcanonical rate coefficients, which are subsequently implemented within the multiple-well multiple-channel master equation. A variety of energy transfer parameters are considered, and the sensitivity of the thermal rate predictions to these parameters is explored. The predictions for the thermal decomposition rate coefficient are found to be in good agreement with the limited experimental data. Modified Arrhenius representations of the rate constants are reported for utility in combustion modeling.

  7. Efficacy and kinetics of opioid action on acutely dissociated neurons.

    PubMed

    Ingram, S; Wilding, T J; McCleskey, E W; Williams, J T

    1997-07-01

    Opioids have been shown to cause a potent inhibition of neurons in the locus ceruleus (LC) in vivo in brain slices and isolated neurons; however, the kinetics of opioid action have not been described. In this study, we used acutely isolated LC neurons to examine opioid and alpha2-adrenoceptor action on potassium and calcium currents. [Met]Enkephalin (ME), [D-Ser2,Leu5,Thr6]-enkephalin, etorphine, and [D-Ala2,N-Me-Phe4,Gly-ol5]enkephalin increased potassium conductance, whereas morphine and naloxone were antagonists. The time constant of potassium channel activation was approximately 0.7 sec and was the same for each agonist. The amplitude of the current and the time constant of decay were dependent on the agonist, suggesting that agonist efficacy and affinity, respectively, determined these parameters. The amplitude of potassium current induced by the alpha2-adrenoceptor agonist UK14304 was not significantly different from that induced by ME, but the time constant of current activation was half that of ME, and the decline was more rapid. When potassium conductances were blocked with the combination of internal cesium and external barium, opioid and alpha2 agonists had no effect at potentials more negative than -50 mV and decreased barium currents at potentials between -40 and +20 mV. Both morphine and clonidine caused a small inhibition of barium current. In dorsal root ganglion cells, morphine alone had small and inconsistent effects on the calcium current, but it always competitively antagonized the inhibition caused by [D-Ala2,N-Me-Phe4,Gly-ol5]enkephalin. The results in isolated LC neurons suggest 1) the amplitude and time course of the opioid-induced potassium current depend on agonist efficacy and affinity and 2) the coupling of both mu-opioid and alpha2-adrenoceptors to calcium channels seems to be more efficient than that to potassium channels.

  8. Cognitive behavioral hypnotherapy for dissociative disorders.

    PubMed

    Fine, Catherine G

    2012-04-01

    Dissociative disorders (DD) prevail as sequelae to overwhelming experiences in childhood. These readily formed post-traumatic responses and trance states develop in high hypnotizable subjects whose dysregulations become organized into ego states. A cognitive behavioral hypnotherapeutic treatment model will effectively contain, explore, metabolize, and resolve these life-endangering conditions. This article will detail the cognitive hypnotic world of DD patients, the relational spaces of the ego states, and the triphasic treatment mode to successfully resolve the dissociative pathology. Structured and phase appropriate hypnotic interventions will be described.

  9. Quantum Zeno control of coherent dissociation

    SciTech Connect

    Khripkov, C.; Vardi, A.

    2011-08-15

    We study the effect of dephasing on the coherent dissociation dynamics of an atom-molecule Bose-Einstein condensate. We show that when phase-noise intensity is strong with respect to the inverse correlation time of the stimulated process, dissociation is suppressed via a Bose enhanced quantum Zeno effect. This is complementary to the quantum Zeno control of phase-diffusion in a bimodal condensate by symmetric noise [Phys. Rev. Lett. 100, 220403 (2008)] in that the controlled process here is phase formation and the required decoherence mechanism for its suppression is purely phase noise.

  10. Phenol dissociation on pristine and defective graphene

    NASA Astrophysics Data System (ADS)

    Widjaja, Hantarto; Oluwoye, Ibukun; Altarawneh, Mohammednoor; Hamra, A. A. B.; Lim, H. N.; Huang, N. M.; Yin, Chun-Yang; Jiang, Zhong-Tao

    2017-03-01

    Phenol (C6H5O‒H) dissociation on both pristine and defective graphene sheets in terms of associated enthalpic requirements of the reaction channels was investigated. Here, we considered three common types of defective graphene, namely, Stone-Wales, monovacancy and divacancy configurations. Theoretical results demonstrate that, graphene with monovacancy creates C atoms with dangling bond (unpaired valence electron), which remains particularly useful for spontaneous dissociation of phenol into phenoxy (C6H5O) and hydrogen (H) atom. The reactions studied herein appear barrierless with reaction exothermicity as high as 2.2 eV. Our study offers fundamental insights into another potential application of defective graphene sheets.

  11. Charge Transfer Dissociation of Complex Oligosaccharides: Comparison with Collision-Induced Dissociation and Extreme Ultraviolet Dissociative Photoionization

    NASA Astrophysics Data System (ADS)

    Ropartz, David; Li, Pengfei; Fanuel, Mathieu; Giuliani, Alexandre; Rogniaux, Hélène; Jackson, Glen P.

    2016-10-01

    The structural characterization of oligosaccharides still challenges the field of analytical chemistry. Tandem mass spectrometry offers many advantages toward this aim, although the generic fragmentation method (low-energy collision-induced dissociation) shows clear limitations and is often insufficient to retrieve some essential structural information on these molecules. In this work, we present the first application of helium charge transfer dissociation (He-CTD) to characterize the structure of complex oligosaccharides. We compare this method with low-energy collision-induced dissociation and extreme-ultraviolet dissociative photoionization (XUV-DPI), which was shown previously to ensure the successful characterization of complex glycans. Similarly to what could be obtained by XUV-DPI, He-CTD provides a complete description of the investigated structures by producing many informative cross-ring fragments and no ambiguous fragmentation. Unlike XUV-DPI, which is performed at a synchrotron source, He-CTD has the undeniable advantage of being implementable in a conventional benchtop ion trap in a conventional laboratory setting.

  12. Protein-ligand binding affinity determination by the waterLOGSY method: An optimised approach considering ligand rebinding

    PubMed Central

    Huang, Renjie; Bonnichon, Arnaud; Claridge, Timothy D. W.; Leung, Ivanhoe K. H.

    2017-01-01

    WaterLOGSY is a popular ligand-observed NMR technique to screen for protein-ligand interactions, yet when applied to measure dissociation constants (KD) through ligand titration, the results were found to be strongly dependent on sample conditions. Herein, we show that accurate KDs can be obtained by waterLOGSY with optimised experimental setup. PMID:28256624

  13. Protein-ligand binding affinity determination by the waterLOGSY method: An optimised approach considering ligand rebinding

    NASA Astrophysics Data System (ADS)

    Huang, Renjie; Bonnichon, Arnaud; Claridge, Timothy D. W.; Leung, Ivanhoe K. H.

    2017-03-01

    WaterLOGSY is a popular ligand-observed NMR technique to screen for protein-ligand interactions, yet when applied to measure dissociation constants (KD) through ligand titration, the results were found to be strongly dependent on sample conditions. Herein, we show that accurate KDs can be obtained by waterLOGSY with optimised experimental setup.

  14. Interventions for dissociated vertical deviation

    PubMed Central

    Hatt, Sarah R; Wang, Xue; Holmes, Jonathan M

    2015-01-01

    Background The term “strabismus” describes misalignment of the eyes. One or both eyes may deviate inward, outward, upward, or downward. Dissociated vertical deviation (DVD) is a well-recognized type of upward drifting of one or both eyes, which can occur in children or adults. DVD often develops in the context of infantile- or childhood-onset horizontal strabismus, either esotropia (inward-turning) or exotropia (outward-turning). For some individuals, DVD remains controlled and can only be detected during clinical testing. For others, DVD becomes spontaneously “manifest” and the eye drifts up of its own accord. Spontaneously manifest DVD can be difficult to control and often causes psychosocial concerns. Traditionally, DVD has been thought to be asymptomatic, although some individuals have double vision. More recently it has been suggested that individuals with DVD may also suffer from eyestrain. Treatment for DVD may be sought either due to psychosocial concerns or because of these symptoms. The standard treatment for DVD is a surgical procedure; non-surgical treatments are offered less commonly. Although there are many studies evaluating different management options for the correction of DVD, a lack of clarity remains regarding which treatments are most effective. Objectives The objective of this review was to determine the effectiveness and safety of various surgical and non-surgical interventions in randomized controlled trials of participants with DVD. Search methods We searched CENTRAL (which contains the Cochrane Eyes and Vision Trials Register) (2015, Issue 8), Ovid MEDLINE, Ovid MEDLINE In-Process and Other Non-Indexed Citations, Ovid MEDLINE Daily, Ovid OLDMEDLINE (January 1946 to August 2015), EMBASE (January 1980 to August 2015), PubMed (1948 to August 2015), Latin American and Caribbean Health Sciences Literature Database (LILACS) (1982 to August 2015), the metaRegister of Controlled Trials (mRCT) (www.controlled-trials.com) (last searched 3

  15. QCD coupling constants and VDM

    SciTech Connect

    Erkol, G.; Ozpineci, A.; Zamiralov, V. S.

    2012-10-23

    QCD sum rules for coupling constants of vector mesons with baryons are constructed. The corresponding QCD sum rules for electric charges and magnetic moments are also derived and with the use of vector-meson-dominance model related to the coupling constants. The VDM role as the criterium of reciprocal validity of the sum rules is considered.