All human Na(+)-K(+)-ATPase alpha-subunit isoforms have a similar affinity for cardiac glycosides.

PubMed

Wang, J; Velotta, J B; McDonough, A A; Farley, R A

2001-10-01

Three alpha-subunit isoforms of the sodium pump, which is the receptor for cardiac glycosides, are expressed in human heart. The aim of this study was to determine whether these isoforms have distinct affinities for the cardiac glycoside ouabain. Equilibrium ouabain binding to membranes from a panel of different human tissues and cell lines derived from human tissues was compared by an F statistic to determine whether a single population of binding sites or two populations of sites with different affinities would better fit the data. For all tissues, the single-site model fit the data as well as the two-site model. The mean equilibrium dissociation constant (K(d)) for all samples calculated using the single-site model was 18 +/- 6 nM (mean +/- SD). No difference in K(d) was found between nonfailing and failing human heart samples, although the maximum number of binding sites in failing heart was only approximately 50% of the number of sites in nonfailing heart. Measurement of association rate constants and dissociation rate constants confirmed that the binding affinities of the different human alpha-isoforms are similar to each other, although calculated K(d) values were lower than those determined by equilibrium binding. These results indicate both that the affinity of all human alpha-subunit isoforms for ouabain is similar and that the increased sensitivity of failing human heart to cardiac glycosides is probably due to a reduction in the number of pumps in the heart rather than to a selective inhibition of a subset of pumps with different affinities for the drugs.

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.

Enrichment and Analysis of Non-enzymatically Glycated Peptides: Boronate Affinity Chromatography Coupled with Electron Transfer Dissociation Mass Spectrometry

PubMed Central

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

2008-01-01

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. ETD fragmentation mode permitted identification of a significantly higher number of glycated peptides (87.6% of all identified peptides) versus CID mode (17.0% of all identified peptides), when utilizing enrichment on first the protein and then the peptide level. This study illustrates that phenylboronate affinity chromatography coupled with LC-MS/MS and using ETD as the fragmentation mode is an efficient approach for analysis of glycated proteins and may have broad application in studies of diabetes mellitus. PMID:17488106

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 ...

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

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tiwari, Manisha; Mikuni, Shintaro; Muto, Hideki

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 inmore » 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.« less

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.

High-Affinity Binding of Remyelinating Natural Autoantibodies to Myelin-Mimicking Lipid Bilayers Revealed by Nanohole Surface Plasmon Resonance

PubMed Central

Wittenberg, Nathan J.; Im, Hyungsoon; Xu, Xiaohua; Wootla, Bharath; Watzlawik, Jens; Warrington, Arthur E.; Rodriguez, Moses; Oh, Sang-Hyun

2012-01-01

Multiple sclerosis is a progressive neurological disorder that results in the degradation of myelin sheaths that insulate axons in the central nervous system. Therefore promotion of myelin repair is a major thrust of multiple sclerosis treatment research. Two mouse monoclonal natural autoantibodies, O1 and O4, promote myelin repair in several mouse models of multiple sclerosis. Natural autoantibodies are generally polyreactive and predominantly of the IgM isotype. The prevailing paradigm is that because they are polyreactive, these antibodies bind antigens with low affinities. Despite their wide use in neuroscience and glial cell research, however, the affinities and kinetic constants of O1 and O4 antibodies have not been measured to date. In this work, we developed a membrane biosensing platform based on surface plasmon resonance in gold nanohole arrays with a series of surface modification techniques to form myelin-mimicking lipid bilayer membranes to measure both the association and dissociation rate constants for O1 and O4 antibodies binding to their myelin lipid antigens. The ratio of rate constants shows that O1 and O4 bind to galactocerebroside and sulfated galactocerebroside, respectively, with unusually small apparent dissociation constants (KD ~0.9 nM) for natural autoantibodies. This is approximately one to two orders of magnitude lower than typically observed for the highest affinity natural autoantibodies. We propose that the unusually high affinity of O1 and O4 to their targets in myelin contributes to the mechanism by which they signal oligodendrocytes and induce central nervous system repair. PMID:22762372

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.

Analysis of Protein Interactions with Picomolar Binding Affinity by Fluorescence-Detected Sedimentation Velocity

PubMed Central

2014-01-01

The study of high-affinity protein interactions with equilibrium dissociation constants (KD) in the picomolar range is of significant interest in many fields, but the characterization of stoichiometry and free energy of such high-affinity binding can be far from trivial. Analytical ultracentrifugation has long been considered a gold standard in the study of protein interactions but is typically applied to systems with micromolar KD. Here we present a new approach for the study of high-affinity interactions using fluorescence detected sedimentation velocity analytical ultracentrifugation (FDS-SV). Taking full advantage of the large data sets in FDS-SV by direct boundary modeling with sedimentation coefficient distributions c(s), we demonstrate detection and hydrodynamic resolution of protein complexes at low picomolar concentrations. We show how this permits the characterization of the antibody–antigen interactions with low picomolar binding constants, 2 orders of magnitude lower than previously achieved. The strongly size-dependent separation and quantitation by concentration, size, and shape of free and complex species in free solution by FDS-SV has significant potential for studying high-affinity multistep and multicomponent protein assemblies. PMID:24552356

Studies on lectins. XXXII. Application of affinity electrophoresis to the study of the interaction of lectins and their derivatives with sugars.

PubMed

Horejsí, V; Tichá, M; Kocourek, J

1977-09-29

Affinity electrophoresis was used to study the sugar binding heterogeneity of lectins or their derivatives. Commercial and demetallized preparations of concanavalin A could be resolved by affinity electrophoresis into three components with different affinity to immobilized sugar. Similarly the Vicia cracca lectin obtained by affinity chromatography behaved on affinity gels as a mixture of active and inactive molecular species. Affinity electrophoresis has shown that the nonhemagglutinating acetylated lentil lectin and photo-oxidized or sulfenylated pea lectin retain their sugar binding properties; dissociation constants of saccharide complexes of these derivatives are similar to those of native lectins. The presence of specific immobilized sugar in the affinity gel improved the resolution of isolectins from Dolichos biflorus and Ricinus communis seeds.

Characterization of solution-phase drug-protein interactions by ultrafast affinity extraction.

PubMed

Beeram, Sandya R; Zheng, Xiwei; Suh, Kyungah; Hage, David S

2018-03-03

A number of tools based on high-performance affinity separations have been developed for studying drug-protein interactions. An example of one recent approach is ultrafast affinity extraction. This method has been employed to examine the free (or non-bound) fractions of drugs and other solutes in simple or complex samples that contain soluble binding agents. These free fractions have also been used to determine the binding constants and rate constants for the interactions of drugs with these soluble agents. This report describes the general principles of ultrafast affinity extraction and the experimental conditions under which it can be used to characterize such interactions. This method will be illustrated by utilizing data that have been obtained when using this approach to measure the binding and dissociation of various drugs with the serum transport proteins human serum albumin and alpha 1 -acid glycoprotein. A number of practical factors will be discussed that should be considered in the design and optimization of this approach for use with single-column or multi-column systems. Techniques will also be described for analyzing the resulting data for the determination of free fractions, rate constants and binding constants. In addition, the extension of this method to complex samples, such as clinical specimens, will be considered. Copyright © 2018 Elsevier Inc. All rights reserved.

Force and Stress along Simulated Dissociation Pathways of Cucurbituril-Guest Systems.

PubMed

Velez-Vega, Camilo; Gilson, Michael K

2012-03-13

The field of host-guest chemistry provides computationally tractable yet informative model systems for biomolecular recognition. We applied molecular dynamics simulations to study the forces and mechanical stresses associated with forced dissociation of aqueous cucurbituril-guest complexes with high binding affinities. First, the unbinding transitions were modeled with constant velocity pulling (steered dynamics) and a soft spring constant, to model atomic force microscopy (AFM) experiments. The computed length-force profiles yield rupture forces in good agreement with available measurements. We also used steered dynamics with high spring constants to generate paths characterized by a tight control over the specified pulling distance; these paths were then equilibrated via umbrella sampling simulations and used to compute time-averaged mechanical stresses along the dissociation pathways. The stress calculations proved to be informative regarding the key interactions determining the length-force profiles and rupture forces. In particular, the unbinding transition of one complex is found to be a stepwise process, which is initially dominated by electrostatic interactions between the guest's ammoniums and the host's carbonyl groups, and subsequently limited by the extraction of the guest's bulky bicyclooctane moiety; the latter step requires some bond stretching at the cucurbituril's extraction portal. Conversely, the dissociation of a second complex with a more slender guest is mainly driven by successive electrostatic interactions between the different guest's ammoniums and the host's carbonyl groups. The calculations also provide information on the origins of thermodynamic irreversibilities in these forced dissociation processes.

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

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.

Development and validation of an oxygen dissociation assay, a screening platform for discovering, and characterizing hemoglobin-oxygen affinity modifiers.

PubMed

Patel, Mira P; Siu, Vincent; Silva-Garcia, Abel; Xu, Qing; Li, Zhe; Oksenberg, Donna

2018-01-01

Hemoglobin (Hb) is a critical molecule necessary for all vertebrates to maintain aerobic metabolism. Hb-oxygen (O 2 ) affinity modifiers have been studied to address various diseases including sickle cell disease, hypoxemia, tumor hypoxia, and wound healing. However, drug development of exogenous Hb modifiers has been hindered by the lack of a technique to rapidly screen compounds for their ability to alter Hb-O 2 affinity. We have developed a novel screening assay based upon the spectral changes observed during Hb deoxygenation and termed it the oxygen dissociation assay (ODA). ODA allows for the quantitation of oxygenated Hb at given time points during Hb deoxygenation on a 96-well plate. This assay was validated by comparing the ability of 500 Hb modifiers to alter the Hb-O 2 affinity in the ODA vs the oxygen equilibrium curves obtained using the industry standard Hemox Analyzer instrument. A correlation ( R 2 ) of 0.7 indicated that the ODA has the potential to screen and identify potent exogenous Hb modifiers. In addition, it allows for concurrent comparison of compounds, concentrations, buffers, or pHs on the level of Hb oxygenation. With a cost-effective, simple, rapid, and highly adaptable assay, the ODA will allow researchers to rapidly characterize Hb-O 2 affinity modifiers.

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

DOE Office of Scientific and Technical Information (OSTI.GOV)

Osborne, David; Lawson, Patrick; Adams, Nigel, E-mail: ngadams@uga.edu

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,more » 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.« less

A Precise Method for Processing Data to Determine the Dissociation Constants of Polyhydroxy Carboxylic Acids via Potentiometric Titration.

PubMed

Huang, Kaixuan; Xu, Yong; Lu, Wen; Yu, Shiyuan

2017-12-01

The thermodynamic dissociation constants of xylonic acid and gluconic acid were studied via potentiometric methods, and the results were verified using lactic acid, which has a known pKa value, as a model compound. Solutions of xylonic acid and gluconic acid were titrated with a standard solution of sodium hydroxide. The determined pKa data were processed via the method of derivative plots using computer software, and the accuracy was validated using the Gran method. The dissociation constants associated with the carboxylic acid group of xylonic and gluconic acids were determined to be pKa 1  = 3.56 ± 0.07 and pKa 1  = 3.74 ± 0.06, respectively. Further, the experimental data showed that the second deprotonation constants associated with a hydroxyl group of each of the two acids were pKa 2  = 8.58 ± 0.12 and pKa 2  = 7.06 ± 0.08, respectively. The deprotonation behavior of polyhydroxy carboxylic acids was altered using various ratios with Cu(II) to form complexes in solution, and this led to proposing a hypothesis for further study.

Chromatographic studies of drug interactions with alpha1-acid glycoprotein by ultrafast affinity extraction and peak profiling.

PubMed

Beeram, Sandya; Bi, Cong; Zheng, Xiwei; Hage, David S

2017-05-12

Interactions with serum proteins such as alpha 1 -acid glycoprotein (AGP) can have a significant effect on the behavior and pharmacokinetics of drugs. Ultrafast affinity extraction and peak profiling were used with AGP microcolumns to examine these processes for several model drugs (i.e., chlorpromazine, disopyramide, imipramine, lidocaine, propranolol and verapamil). The association equilibrium constants measured for these drugs with soluble AGP by ultrafast affinity extraction were in the general range of 10 4 -10 6 M -1 at pH 7.4 and 37°C and gave good agreement with literature values. Some of these values were dependent on the relative drug and protein concentrations that were present when using a single-site binding model; these results suggested a more complex mixed-mode interaction was actually present, which was also then used to analyze the data. The apparent dissociation rate constants that were obtained by ultrafast affinity extraction when using a single-site model varied from 0.14 to 7.0s -1 and were dependent on the relative drug and protein concentrations. Lower apparent dissociation rate constants were obtained by this approach as the relative amount of drug versus protein was decreased, with the results approaching those measured by peak profiling at low drug concentrations. This information should be useful in better understanding how these and other drugs interact with AGP in the circulation. In addition, the chromatographic approaches that were optimized and used in this report to examine these systems can be adapted for the analysis of other solute-protein interactions of biomedical interest. Copyright © 2017 Elsevier B.V. All rights reserved.

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%.

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). Copyright © 2015 Elsevier B.V. All rights reserved.

Ligand-receptor binding affinities from saturation transfer difference (STD) NMR spectroscopy: the binding isotherm of STD initial growth rates.

PubMed

Angulo, Jesús; Enríquez-Navas, Pedro M; Nieto, Pedro M

2010-07-12

The direct evaluation of dissociation constants (K(D)) from the variation of saturation transfer difference (STD) NMR spectroscopy values with the receptor-ligand ratio is not feasible due to the complex dependence of STD intensities on the spectral properties of the observed signals. Indirect evaluation, by competition experiments, allows the determination of K(D), as long as a ligand of known affinity is available for the protein under study. Herein, we present a novel protocol based on STD NMR spectroscopy for the direct measurements of receptor-ligand dissociation constants (K(D)) from single-ligand titration experiments. The influence of several experimental factors on STD values has been studied in detail, confirming the marked impact on standard determinations of protein-ligand affinities by STD NMR spectroscopy. These factors, namely, STD saturation time, ligand residence time in the complex, and the intensity of the signal, affect the accumulation of saturation in the free ligand by processes closely related to fast protein-ligand rebinding and longitudinal relaxation of the ligand signals. The proposed method avoids the dependence of the magnitudes of ligand STD signals at a given saturation time on spurious factors by constructing the binding isotherms using the initial growth rates of the STD amplification factors, in a similar way to the use of NOE growing rates to estimate cross relaxation rates for distance evaluations. Herein, it is demonstrated that the effects of these factors are cancelled out by analyzing the protein-ligand association curve using STD values at the limit of zero saturation time, when virtually no ligand rebinding or relaxation takes place. The approach is validated for two well-studied protein-ligand systems: the binding of the saccharides GlcNAc and GlcNAcbeta1,4GlcNAc (chitobiose) to the wheat germ agglutinin (WGA) lectin, and the interaction of the amino acid L-tryptophan to bovine serum albumin (BSA). In all cases, the

Simulations of dissociation constants in low pressure supercritical water

NASA Astrophysics Data System (ADS)

Halstead, S. J.; An, P.; Zhang, S.

2014-09-01

This article reports molecular dynamics simulations of the dissociation of hydrochloric acid and sodium hydroxide in water from ambient to supercritical temperatures at a fixed pressure of 250 atm. Corrosion of reaction vessels is known to be a serious problem of supercritical water, and acid/base dissociation can be a significant contributing factor to this. The SPC/e model was used in conjunction with solute models determined from density functional calculations and OPLSAA Lennard-Jones parameters. Radial distribution functions were calculated, and these show a significant increase in solute-solvent ordering upon forming the product ions at all temperatures. For both dissociations, rapidly decreasing entropy of reaction was found to be the controlling thermodynamic factor, and this is thought to arise due to the ions produced from dissociation maintaining a relatively high density and ordered solvation shell compared to the reactants. The change in entropy of reaction reaches a minimum at the critical temperature. The values of pKa and pKb were calculated and both increased with temperature, in qualitative agreement with other work, until a maximum value at 748 K, after which there was a slight decrease.

Effects of alkyl chain length and solvents on thermodynamic dissociation constants of the ionic liquids with one carboxyl group in the alkyl chain of imidazolium cations.

PubMed

Chen, Yuehua; Wang, Huiyong; Wang, Jianji

2014-05-01

Thermodynamic dissociation constants of the Brønsted acidic ionic liquids (ILs) are important for their catalytic and separation applications. In this work, a series of imidazolium bromides with one carboxylic acid substitute group in their alkyl chain ([{(CH2)nCOOH}mim]Br, n = 1,3,5,7) have been synthesized, and their dissociation constants (pKa) at different ionic strengths have been determined in aqueous and aqueous organic solvents at 0.1 mole fraction (x) of ethanol, glycol, iso-propanol, and dimethyl sulfoxide by potentiometric titrations at 298.2 K. The standard thermodynamic dissociation constants (pKa(T)) of the ILs in these solvents were calculated from the extended Debye-Hückel equation. It was found that the pKa values increased with the increase of ionic strength of the media and of the addition of organic solvent in water. The pKa(T) values also increased with the increase of the alkyl chain length of cations of the ILs. In addition, the effect of solvent nature on pKa(T) values is interpreted from solvation of the dissociation components and their Gibbs energy of transfer from water to aqueous organic solutions.

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.

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;…

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

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

DOE Office of Scientific and Technical Information (OSTI.GOV)

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

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 ofmore » 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.« less

Evaluation of galectin binding by frontal affinity chromatography (FAC).

PubMed

Iwaki, Jun; Hirabayashi, Jun

2015-01-01

Frontal affinity chromatography (FAC) is a simple and versatile procedure enabling quantitative determination of diverse biological interactions in terms of dissociation constants (K d), even though these interactions are relatively weak. The method is best applied to glycans and their binding proteins, with the analytical system operating on the basis of highly reproducible isocratic elution by liquid chromatography. Its application to galectins has been successfully developed to characterize their binding specificities in detail. As a result, their minimal requirements for recognition of disaccharides, i.e., β-galactosides, as well as characteristic features of individual galectins, have been elucidated. In this chapter, we describe standard procedures to determine the K d's for interactions between a series of standard glycans and various galectins.

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

PubMed

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

2015-09-01

Artificial peptides designed for molecular recognition of a bacterial toxin have been developed. Vacuolating cytotoxin A protein (VacA) is a major virulence factor of Helicobacter pylori, a gram-negative microaerophilic bacterium inhabiting the upper gastrointestinal tract, particularly the stomach. This study attempted to identify specific peptide sequences with high affinity for VacA using systematic directed evolution in vitro, a cDNA display method. A surface plasmon resonance-based biosensor and fluorescence correlation spectroscopy to examine binding of peptides with VacA identified a peptide (GRVNQRL) with high affinity. Cyclization of the peptide by attaching cysteine residues to both termini improved its binding affinity to VacA, with a dissociation constant (Kd ) of 58 nm. This study describes a new strategy for the development of artificial functional peptides, which are promising materials in biochemical analyses and medical applications. Copyright © 2015 European Peptide Society and John Wiley & Sons, Ltd.

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

Interplay between binding affinity and kinetics in protein-protein interactions.

PubMed

Cao, Huaiqing; Huang, Yongqi; Liu, Zhirong

2016-07-01

To clarify the interplay between the binding affinity and kinetics of protein-protein interactions, and the possible role of intrinsically disordered proteins in such interactions, molecular simulations were carried out on 20 protein complexes. With bias potential and reweighting techniques, the free energy profiles were obtained under physiological affinities, which showed that the bound-state valley is deep with a barrier height of 12 - 33 RT. From the dependence of the affinity on interface interactions, the entropic contribution to the binding affinity is approximated to be proportional to the interface area. The extracted dissociation rates based on the Arrhenius law correlate reasonably well with the experimental values (Pearson correlation coefficient R = 0.79). For each protein complex, a linear free energy relationship between binding affinity and the dissociation rate was confirmed, but the distribution of the slopes for intrinsically disordered proteins showed no essential difference with that observed for ordered proteins. A comparison with protein folding was also performed. Proteins 2016; 84:920-933. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Measurement of the affinity and phosphorylation constants governing irreversible inhibition of cholinesterases by di-isopropyl phosphorofluoridate

PubMed Central

Main, A. R.; Iverson, F.

1966-01-01

1. A procedure is described for determining the affinity constant Ka and the phosphorylation constant kp for the inhibition by di-isopropyl phosphorofluoridate of erythrocyte acetylcholinesterase and serum cholinesterase. The procedure depends on the use of a specially designed reaction vessel with which incubation times as short as 1·2sec. could be obtained at any convenient temperature. 2. The Ka of acetylcholinesterase decreased from 1·58 (±0·22)×10−3m at 5° to 1·17 (±0·10)×10−3m at 25° and the associated change in enthalpy was 2980 cal. 3. The kp of acetylcholinesterase increased from 11·9 (±0·7)min.−1 at 5° to 40·7 (±1·4)min.−1 at 25°, indicating an activational energy of 9600 cal. The change in entropy associated with Ka was 23·5 cal. degree−1 at 25°. 4. At 5°, the Ka and kp of serum cholinesterase were 9·95 (±1·10)×10−6m and 11·2 (±0·63)min.−1 respectively. 5. The 150-fold difference in the inhibitory power of di-isopropyl phosphorofluoridate for the two cholinesterases was attributed entirely to differences in affinity. PMID:5968549

Highly affine and selective aptamers against cholera toxin as capture elements in magnetic bead-based sandwich ELAA.

PubMed

Frohnmeyer, Esther; Frisch, Farina; Falke, Sven; Betzel, Christian; Fischer, Markus

2018-03-10

Aptamers are single-stranded DNA or RNA oligonucleotides, which have been emerging as recognition elements in disease diagnostics and food control, including the detection of bacterial toxins. In this study, we employed the semi-automated just in time-selection to identify aptamers that bind to cholera toxin (CT) with high affinity and specificity. CT is the main virulence factor of Vibrio cholerae and the causative agent of the eponymous disease. For the selected aptamers, dissociation constants in the low nanomolar range (23-56 nM) were determined by fluorescence-based affinity chromatography and cross-reactivity against related proteins was evaluated by direct enzyme-linked aptamer assay (ELAA). Aptamer CT916 has a dissociation constant of 48.5 ± 0.5 nM and shows negligible binding to Shiga-like toxin 1B, protein A and BSA. This aptamer was chosen to develop a sandwich ELAA for the detection of CT from binding buffer and local tap water. Amine-C6- or biotin-modified CT916 was coupled to magnetic beads to serve as the capture element. Using an anti-CT polyclonal antibody as the reporter, detection limits of 2.1 ng/ml in buffer and 2.4 ng/ml in tap water, with a wide log-linear dynamic range from 1 ng/ml to 1000 ng/ml and 500 ng/ml, respectively, were achieved. Copyright © 2018 Elsevier B.V. All rights reserved.

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.

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

The Universal Statistical Distributions of the Affinity, Equilibrium Constants, Kinetics and Specificity in Biomolecular Recognition

PubMed Central

Zheng, Xiliang; Wang, Jin

2015-01-01

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. PMID:25885453

Relationship of nonreturn rates of dairy bulls to binding affinity of heparin to sperm

DOE Office of Scientific and Technical Information (OSTI.GOV)

Marks, J.L.; Ax, R.L.

1985-08-01

The binding of the glycosaminoglycan (3H) heparin to bull spermatozoa was compared with nonreturn rates of dairy bulls. Semen samples from five bulls above and five below an average 71% nonreturn rate were used. Samples consisted of first and second ejaculates on a single day collected 1 d/wk for up to 5 consecutive wk. Saturation binding assays using (TH) heparin were performed to quantitate the binding characteristics of each sample. Scatchard plot analyses indicated a significant difference in the binding affinity for (TH) heparin between bulls of high and low fertility. Dissociation constants were 69.0 and 119.3 pmol for bullsmore » of high and low fertility, respectively. In contrast, the number of binding sites for (TH) heparin did not differ significantly among bulls. Differences in binding affinity of (TH) heparin to bull sperm might be used to predict relative fertility of dairy bulls.« less

PLASS: Protein-ligand affinity statistical score a knowledge-based force-field model of interaction derived from the PDB

NASA Astrophysics Data System (ADS)

Ozrin, V. D.; Subbotin, M. V.; Nikitin, S. M.

2004-04-01

We have developed PLASS (Protein-Ligand Affinity Statistical Score), a pair-wise potential of mean-force for rapid estimation of the binding affinity of a ligand molecule to a protein active site. This scoring function is derived from the frequency of occurrence of atom-type pairs in crystallographic complexes taken from the Protein Data Bank (PDB). Statistical distributions are converted into distance-dependent contributions to the Gibbs free interaction energy for 10 atomic types using the Boltzmann hypothesis, with only one adjustable parameter. For a representative set of 72 protein-ligand structures, PLASS scores correlate well with the experimentally measured dissociation constants: a correlation coefficient R of 0.82 and RMS error of 2.0 kcal/mol. Such high accuracy results from our novel treatment of the volume correction term, which takes into account the inhomogeneous properties of the protein-ligand complexes. PLASS is able to rank reliably the affinity of complexes which have as much diversity as in the PDB.

Red cell 2,3-diphosphoglycerate and oxygen affinity.

PubMed

MacDonald, R

1977-06-01

The ease with which haemoglobin releases oxygen to the tissues is controlled by erythrocytic 2,3-diphosphoglycerate (2,3-DPG) such that an increase in the concentration of 2,3-DPG decreases oxygen affinity and vice versa. This review article describes the synthesis and breakdown of 2,3-DPG in the Embden-Meyerof pathway in red cells and briefly explains the molecular basis for its effect on oxygen affinity. Interaction of the effects of pH, Pco2, temperature and 2,3-DPG on the oxyhaemoglobin dissociation curve are discussed. The role of 2,3-DPG in the intraerythrocytic adaptation to various types of hypoxaemia is described. The increased oxygen affinity of blood stored in acid-citrate-dextrose (ACD) solution has been shown to be due to the decrease in the concentration of 2,3-DPG which occurs during storage. Methods of maintaining the concentration of 2,3-DPG in stored blood are described. The clinical implication of transfusion of elderly people, anaemic or pregnant patients with ACD stored blood to anaesthetically and surgically acceptable haemoglobin concentrations are discussed. Hypophosphataemia in association with parenteral feeding reduces 2,3-DPG concentration and so increases oxygen affinity. Since post-operative use of intravenous fluids such as dextrose or dextrose/saline also lead to hypophosphataemia, the addition of inorganic phosphorus to routine post-operative intravenous fluid may be advisable. Disorders of acid-base balance effect oxygen affinity not only by the direct effect of pH on the oxyhaemoglobin dissociation curve but by its control of 2,3-DPG metabolism. Management of acid-base disorders and pre-operative aklalinization of patients with sickle cell disease whould take account of this. It is known that anaesthesia alters the position of the oxyhaemoglobin dissociation curve, but it is thought that this is independent of any effects which anaesthetic agents may have on 2,3-DPG concentration. In vitro manipulation of 2,3-DPG concentration

Selection is more intelligent than design: improving the affinity of a bivalent ligand through directed evolution.

PubMed

Ahmad, Kareem M; Xiao, Yi; Soh, H Tom

2012-12-01

Multivalent molecular interactions can be exploited to dramatically enhance the performance of an affinity reagent. The enhancement in affinity and specificity achieved with a multivalent construct depends critically on the effectiveness of the scaffold that joins the ligands, as this determines their positions and orientations with respect to the target molecule. Currently, no generalizable design rules exist for construction of an optimal multivalent ligand for targets with known structures, and the design challenge remains an insurmountable obstacle for the large number of proteins whose structures are not known. As an alternative to such design-based strategies, we report here a directed evolution-based method for generating optimal bivalent aptamers. To demonstrate this approach, we fused two thrombin aptamers with a randomized DNA sequence and used a microfluidic in vitro selection strategy to isolate scaffolds with exceptionally high affinities. Within five rounds of selection, we generated a bivalent aptamer that binds thrombin with an apparent dissociation constant (K(d)) <10 pM, representing a ∼200-fold improvement in binding affinity over the monomeric aptamers and a ∼15-fold improvement over the best designed bivalent construct. The process described here can be used to produce high-affinity multivalent aptamers and could potentially be adapted to other classes of biomolecules.

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

The neonatal Fc receptor (FcRn) binds independently to both sites of the IgG homodimer with identical affinity.

PubMed

Abdiche, Yasmina Noubia; Yeung, Yik Andy; Chaparro-Riggers, Javier; Barman, Ishita; Strop, Pavel; Chin, Sherman Michael; Pham, Amber; Bolton, Gary; McDonough, Dan; Lindquist, Kevin; Pons, Jaume; Rajpal, Arvind

2015-01-01

The neonatal Fc receptor (FcRn) is expressed by cells of epithelial, endothelial and myeloid lineages and performs multiple roles in adaptive immunity. Characterizing the FcRn/IgG interaction is fundamental to designing therapeutic antibodies because IgGs with moderately increased binding affinities for FcRn exhibit superior serum half-lives and efficacy. It has been hypothesized that 2 FcRn molecules bind an IgG homodimer with disparate affinities, yet their affinity constants are inconsistent across the literature. Using surface plasmon resonance biosensor assays that eliminated confounding experimental artifacts, we present data supporting an alternate hypothesis: 2 FcRn molecules saturate an IgG homodimer with identical affinities at independent sites, consistent with the symmetrical arrangement of the FcRn/Fc complex observed in the crystal structure published by Burmeister et al. in 1994. We find that human FcRn binds human IgG1 with an equilibrium dissociation constant (KD) of 760 ± 60 nM (N = 14) at 25°C and pH 5.8, and shows less than 25% variation across the other human subtypes. Human IgG1 binds cynomolgus monkey FcRn with a 2-fold higher affinity than human FcRn, and binds both mouse and rat FcRn with a 10-fold higher affinity than human FcRn. FcRn/IgG interactions from multiple species show less than a 2-fold weaker affinity at 37°C than at 25°C and appear independent of an IgG's variable region. Our in vivo data in mouse and rat models demonstrate that both affinity and avidity influence an IgG's serum half-life, which should be considered when choosing animals, especially transgenic systems, as surrogates.

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

PubMed Central

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

2017-01-01

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. PMID:28054999

Dissociation constants, neutralization enthalpies and reactions of 3-styryl-2-mercaptopropenoic and 3-(1-naphthyl)-2-mercaptopropenoic acids.

PubMed

Izquierdo, A; Bosch, E; Beltran, J L

1984-06-01

Dissociation constants (pK(a1) and pK(a2) in water-ethanol medium for 3-styryl-2-mercaptopropenoic and 3-(1-naphthyl)-2-mercaptopropenoic acid have been determined potentiometrically, and pK(a2) for both in aqueous medium, spectrophotometrically. Neutralization enthalpies in water-ethanol medium have been determined by thermometric titration. The reactions with metal ions have been studied, and the main reactions are described. The most sensitive reactions are with titanium(IV) (pD = 7.00) and nickel(II) (pD = 6.50).

Regulatory mechanisms of hemoglobin oxygen affinity in acidosis and alkalosis

PubMed Central

Bellingham, A. J.; Detter, J. C.; Lenfant, C.

1971-01-01

The recent reports of the effect of 2,3-diphosphoglycerate (2,3-DPG) on hemoglobin affinity for oxygen suggested that this substance may play a role in man's adaptation to acidosis and alkalosis. A study of the effect of induced acidosis and alkalosis on the oxyhemoglobin dissociation curve of normal man was therefore carried out, and the mechanisms involved in the physiological regulation of hemoglobin oxygen affinity examined. In acute changes of plasma pH there was no alteration in red cell 2,3-DPG content. However, there were changes in hemoglobin oxygen affinity and these correlated with changes in mean corpuscular hemoglobin concentration (MCHC). With maintained acidosis and alkalosis, red cell 2,3-DPG content was altered and correlated with the changes in hemoglobin oxygen affinity. Both of these mechanisms shift the hemoglobin oxygen dissociation curve opposite to the direct pH (Bohr) effect, and providing the rate of pH change is neither too rapid nor too large, they counteract the direct pH effect and the in vivo hemoglobin oxygen affinity remains unchanged. It is also shown that approximately 35% of the change in hemoglobin oxygen affinity resulting from an alteration in red cell 2,3-DPG, is explained by effect of 2,3-DPG on the red cell pH. PMID:5545127

Human llamas: adaptation to altitude in subjects with high hemoglobin oxygen affinity.

PubMed Central

Hebbel, R P; Eaton, J W; Kronenberg, R S; Zanjani, E D; Moore, L G; Berger, E M

1978-01-01

To assess the adaptive value of the right-shift of the oxyhemoglobin dissociation curve (decreased affinity for oxygen) observed in humans upon altitude exposure, the short-term physiologic responses to altitude-induced hypoxia were evaluated in two subjects with a high oxygen affinity hemoglobin (Hb Andrew-Minneapolis) and in two of their normal siblings. In striking contrast to normal subjects, at moderately high altitude (3,100 m) the high affinity subjects manifested: (a) lesser increments in resting heart rate; (b) minimal increases in plasma and urinary erythropoietin; (c) no decrement in maximal oxygen consumption; and (d) no thrombocytopenia. There was no difference between subject pairs in 2,3-diphosphoglycerate response to altitude exposure. These results tend to contradict the belief that a decrease in hemoglobin oxygen affinity is of adaptive value to humans at moderate altitudes. Rather, they support the hypothesis that, despite disadvantages at low altitude, a left-shifted oxyhemoglobin dissociation curve may confer a degree of preadaptation to altitude. PMID:29054

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. Copyright © 2010 Elsevier B.V. All rights reserved.

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.

High-affinity receptors for bombesin-like peptides in normal guinea pig lung membranes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lach, E.; Trifilieff, A.; Landry, Y.

1991-01-01

The binding of the radiolabeled bombesin analogue ({sup 125}I-Tyr{sup 4})bombesin to guinea-pig lung membranes was investigated. Binding of ({sup 125}I-Tyr{sup 4})bombesin was specific, saturable, reversible and linearly related to the protein concentration. Scatchard analysis of equilibrium binding data at 25C indicated the presence of a single class of non-interacting binding sites for bombesin (B{sub max} = 7.7 fmol/mg protein). The value of the equilibrium dissociation constant (K{sub D} = 90 pM) agrees with a high-affinity binding site. Bombesin and structurally related peptides such as ({sup 125}I-Tyr{sup 4})bombesin, neuromedin B and neuromedin C inhibited the binding of ({sup 125}I-Tyr{sup 4})bombesin inmore » an order of potencies as follows: ({sup 125}I-Tyr{sup 4})bombesin {gt} bombesin {ge} neuromedin C {much gt} neuromedin B. These results indicate that guinea-pig lung membranes possess a single class of bombesin receptors with a high affinity for bombesin and a lower one for neuromedin B.« less

Simultaneous estimation of intracellular free Mg2+ and pH by use of a new pH-dependent dissociation constant of MgATP.

PubMed

Nakayama, Shinsuke; Nomura, Hideki; Smith, Lorraine M; Clark, Joseph F

2002-06-01

In the present technical note for 31P-NMR, we used a new pH-dependent dissociation constant of MgATP, and re-estimated changes in the intracellular free Mg2+ concentration and pH from the chemical shifts of beta- and gamma-ATP during Na+-removal in smooth muscle. We confirmed the role of Na+ - Mg2+ exchange.

Modulators of actin-myosin dissociation: basis for muscle type functional differences during fatigue

PubMed Central

Karatzaferi, Christina; Adamek, Nancy

2017-01-01

The muscle types present with variable fatigue tolerance, in part due to the myosin isoform expressed. However, the critical steps that define “fatigability” in vivo of fast vs. slow myosin isoforms, at the molecular level, are not yet fully understood. We examined the modulation of the ATP-induced myosin subfragment 1 (S1) dissociation from pyrene-actin by inorganic phosphate (Pi), pH, and temperature using a specially modified stopped-flow system that allowed fast kinetics measurements at physiological temperature. We contrasted the properties of rabbit psoas (fast) and bovine masseter (slow) myosins (obtained from samples collected from New Zealand rabbits and from a licensed abattoir, respectively, according to institutional and national ethics permits). To identify ATP cycling biochemical intermediates, we assessed ATP binding to a preequilibrated mixture of actomyosin and variable [ADP], pH (pH 7 vs. pH 6.2), and Pi (zero, 15, or 30 added mM Pi) in a range of temperatures (5 to 45°C). Temperature and pH variations had little, if any, effect on the ADP dissociation constant (KADP) for fast S1, but for slow S1, KADP was weakened with increasing temperature or low pH. In the absence of ADP, the dissociation constant for phosphate (KPi) was weakened with increasing temperature for fast S1. In the presence of ADP, myosin type differences were revealed at the apparent phosphate affinity, depending on pH and temperature. Overall, the newly revealed kinetic differences between myosin types could help explain the in vivo observed muscle type functional differences at rest and during fatigue. PMID:28931538

Determination of the Acid Dissociation Constant of a Phenolic Acid by High Performance Liquid Chromatography: An Experiment for the Upper Level Analytical Chemistry Laboratory

ERIC Educational Resources Information Center

Raboh, Ghada

2018-01-01

A high performance liquid chromatography (HPLC) experiment for the upper level analytical chemistry laboratory is described. The students consider the effect of mobile-phase composition and pH on the retention times of ionizable compounds in order to determine the acid dissociation constant, K[subscript a], of a phenolic acid. Results are analyzed…

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 K d value for the wild type ODC protein-AZ heterodimer ([ODC_WT]-AZ) is approximately 0.22 μM, while the K d 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 K d was 1.4 and 1.2 μM, respectively. These affinities are 6-fold higher than the WT_ODC K d, which suggests that residues 119 and 137 play a role in AZ binding. PMID:22073206

Into the theory of the partial-filling affinity capillary electrophoresis and the determination of apparent stability constants of analyte-ligand complexes.

PubMed

Ansorge, Martin; Dubský, Pavel; Ušelová, Kateřina

2018-03-01

The partial-filling affinity capillary electrophoresis (pf-ACE) works with a ligand present in a background electrolyte that forms a weak complex with an analyte. In contrast to a more popular mobility-shift affinity capillary electrophoresis, only a short plug of the ligand is introduced into a capillary in the pf-ACE. Both methods can serve for determining apparent stability constants of the formed complexes but this task is hindered in the pf-ACE by the fact that the analyte spends only a part of its migration time in a contact with the ligand. In 1998, Amini and Westerlund published a linearization strategy that allows for extracting an effective mobility of an analyte in the presence of a neutral ligand out of the pf-ACE data. The main purpose of this paper is to show that the original formula is only approximate. We derive a new formula and demonstrate its applicability by means of computer simulations. We further inspect several strategies of data processing in the pf-ACE regarding a risk of an error propagation. This establishes a good practice of determining apparent stability constants of analyte-ligand complexes by means of the pf-ACE. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Thiacarbocyanine as ligand in dye-affinity chromatography for protein purification. II. Dynamic binding capacity using lysozyme as a model.

PubMed

Boto, R E F; Anyanwu, U; Sousa, F; Almeida, P; Queiroz, J A

2009-09-01

A constant development of dye-affinity chromatography to replace more traditional techniques is verified, with the aim of increasing specificity in the purification of biomolecules. The establishment of a new dye-affinity chromatographic support imposes their complete characterization, namely with relation to the binding capacity for proteins, in order to evaluate its applicability on global purification processes. Following previous studies, the adsorption of lysozyme onto a thiacarbocyanine dye immobilized on beaded cellulose was investigated. The effect of different parameters, such as temperature, ionic strength, pH, protein concentration and flow rate, on the dynamic binding capacity of the support to retain lysozyme was also studied. Increasing the temperature and the lysozyme concentration had a positive effect on the dynamic binding capacity (DBC), whereas increasing the ionic strength and the flow rate resulted in the opposite. It was also discovered that the pH used had an important impact on the lysozyme binding onto the immobilized dye. The maximum DBC value obtained for lysozyme was 8.6 mg/mL, which was achieved at 30 degrees C and pH 9 with a protein concentration of 0.5 mg/mL and a flow rate of 0.05 mL/min. The dissociation constant (K(d)) obtained was 2.61 +/- 0.36 x 10(-5 )m, proving the affinity interaction between the thiacarbocyanine dye ligand and the lysozyme. Copyright (c) 2009 John Wiley & Sons, Ltd.

Distinct dissociation kinetics between ion pairs: Solvent-coordinate free-energy landscape analysis.

PubMed

Yonetani, Yoshiteru

2015-07-28

Different ion pairs exhibit different dissociation kinetics; however, while the nature of this process is vital for understanding various molecular systems, the underlying mechanism remains unclear. In this study, to examine the origin of different kinetic rate constants for this process, molecular dynamics simulations were conducted for LiCl, NaCl, KCl, and CsCl in water. The results showed substantial differences in dissociation rate constant, following the trend kLiCl < kNaCl < kKCl < kCsCl. Analysis of the free-energy landscape with a solvent reaction coordinate and subsequent rate component analysis showed that the differences in these rate constants arose predominantly from the variation in solvent-state distribution between the ion pairs. The formation of a water-bridging configuration, in which the water molecule binds to an anion and a cation simultaneously, was identified as a key step in this process: water-bridge formation lowers the related dissociation free-energy barrier, thereby increasing the probability of ion-pair dissociation. Consequently, a higher probability of water-bridge formation leads to a higher ion-pair dissociation rate.

Choline Uptake in Agrobacterium tumefaciens by the High-Affinity ChoXWV Transporter▿

PubMed Central

Aktas, Meriyem; Jost, Kathinka A.; Fritz, Christiane; Narberhaus, Franz

2011-01-01

Agrobacterium tumefaciens is a facultative phytopathogen that causes crown gall disease. For successful plant transformation A. tumefaciens requires the membrane lipid phosphatidylcholine (PC), which is produced via the methylation and the PC synthase (Pcs) pathways. The latter route is dependent on choline. Although choline uptake has been demonstrated in A. tumefaciens, the responsible transporter(s) remained elusive. In this study, we identified the first choline transport system in A. tumefaciens. The ABC-type choline transporter is encoded by the chromosomally located choXWV operon (ChoX, binding protein; ChoW, permease; and ChoV, ATPase). The Cho system is not critical for growth and PC synthesis. However, [14C]choline uptake is severely reduced in A. tumefaciens choX mutants. Recombinant ChoX is able to bind choline with high affinity (equilibrium dissociation constant [KD] of ≈2 μM). Since other quaternary amines are bound by ChoX with much lower affinities (acetylcholine, KD of ≈80 μM; betaine, KD of ≈470 μM), the ChoXWV system functions as a high-affinity transporter with a preference for choline. Two tryptophan residues (W40 and W87) located in the predicted ligand-binding pocket are essential for choline binding. The structural model of ChoX built on Sinorhizobium meliloti ChoX resembles the typical structure of substrate binding proteins with a so-called “Venus flytrap mechanism” of substrate binding. PMID:21803998

Affine group formulation of the Standard Model coupled to gravity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chou, Ching-Yi, E-mail: l2897107@mail.ncku.edu.tw; Ita, Eyo, E-mail: ita@usna.edu; Soo, Chopin, E-mail: cpsoo@mail.ncku.edu.tw

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 themore » 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.« less

Structural Basis of Low-Affinity Nickel Binding to the Nickel-Responsive Transcription Factor NikR from Escherichia coli†‡

PubMed Central

2010-01-01

Escherichia coli NikR regulates cellular nickel uptake by binding to the nik operon in the presence of nickel and blocking transcription of genes encoding the nickel uptake transporter. NikR has two binding affinities for the nik operon: a nanomolar dissociation constant with stoichiometric nickel and a picomolar dissociation constant with excess nickel [Bloom, S. L., and Zamble, D. B. (2004) Biochemistry 43, 10029−10038; Chivers, P. T., and Sauer, R. T. (2002) Chem. Biol. 9, 1141−1148]. While it is known that the stoichiometric nickel ions bind at the NikR tetrameric interface [Schreiter, E. R., et al. (2003) Nat. Struct. Biol. 10, 794−799; Schreiter, E. R., et al. (2006) Proc. Natl. Acad. Sci. U.S.A. 103, 13676−13681], the binding sites for excess nickel ions have not been fully described. Here we have determined the crystal structure of NikR in the presence of excess nickel to 2.6 Å resolution and have obtained nickel anomalous data (1.4845 Å) in the presence of excess nickel for both NikR alone and NikR cocrystallized with a 30-nucleotide piece of double-stranded DNA containing the nik operon. These anomalous data show that excess nickel ions do not bind to a single location on NikR but instead reveal a total of 22 possible low-affinity nickel sites on the NikR tetramer. These sites, for which there are six different types, are all on the surface of NikR, and most are found in both the NikR alone and NikR−DNA structures. Using a combination of crystallographic data and molecular dynamics simulations, the nickel sites can be described as preferring octahedral geometry, utilizing one to three protein ligands (typically histidine) and at least two water molecules. PMID:20704276

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.

Fragment screening of cyclin G-associated kinase by weak affinity chromatography.

PubMed

Meiby, Elinor; Knapp, Stefan; Elkins, Jonathan M; Ohlson, Sten

2012-11-01

Fragment-based drug discovery (FBDD) has become a new strategy for drug discovery where lead compounds are evolved from small molecules. These fragments form low affinity interactions (dissociation constant (K(D)) = mM - μM) with protein targets, which require fragment screening methods of sufficient sensitivity. Weak affinity chromatography (WAC) is a promising new technology for fragment screening based on selective retention of fragments by a drug target. Kinases are a major pharmaceutical target, and FBDD has been successfully applied to several of these targets. In this work, we have demonstrated the potential to use WAC in combination with mass spectrometry (MS) detection for fragment screening of a kinase target-cyclin G-associated kinase (GAK). One hundred seventy fragments were selected for WAC screening by virtual screening of a commercial fragment library against the ATP-binding site of five different proteins. GAK protein was immobilized on a capillary HPLC column, and compound binding was characterized by frontal affinity chromatography. Compounds were screened in sets of 13 or 14, in combination with MS detection for enhanced throughput. Seventy-eight fragments (46 %) with K(D) < 200 μM were detected, including a few highly efficient GAK binders (K(D) of 2 μM; ligand efficiency = 0.51). Of special interest is that chiral screening by WAC may be possible, as two stereoisomeric fragments, which both contained one chiral center, demonstrated twin peaks. This ability, in combination with the robustness, sensitivity, and simplicity of WAC makes it a new method for fragment screening of considerable potential.

Affinity purification of seminalplasmin and characterization of its interaction with calmodulin.

PubMed Central

Comte, M; Malnoë, A; Cox, J A

1986-01-01

Bull seminalplasmin antagonizes with high potency and selectivity the activating effect of calmodulin on target enzymes [Gietzen & Galla (1985) Biochem. J. 230, 277-280]. In the present paper we establish that seminalplasmin forms a 1:1, Ca2+-dependent and urea-resistant complex with calmodulin. The dissociation constant equals 1.6 nM. In the absence of Ca2+ a low-affinity complex is formed that is disrupted by 4 M-urea. On the basis of these properties, a fast affinity purification of seminalplasmin was developed. The high specificity of seminalplasmin as a calmodulin antagonist was demonstrated for the multipathway-regulated adenylate cyclase of bovine cerebellum. Far-u.v. c.d. properties are consistent with a random form of seminalplasmin in aqueous solution; 23% alpha-helix is induced on interaction with calmodulin. The fluorescence properties of the single tryptophan residue of seminalplasmin are markedly changed on formation of the complex. These studies allowed us to locate tentatively the peptide segment that interacts with calmodulin, and to ascertain the structural homology between seminalplasmin and other calmodulin-binding peptides. Additional material, showing the inhibition of calmodulin-mediated activation of bovine brain phosphodiesterase by melittin and seminalplasmin and also the near-u.v. spectrum of affinity-purified seminalplasmin, has been deposited as supplement SUP 50135 (4 pages) at the British Library Lending Division, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies may be obtained on the terms indicated in Biochem. J. (1986) 233, 5. Images Fig. 2. PMID:3814096

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

Isolation and characterization of high affinity aptamers against DNA polymerase iota.

PubMed

Lakhin, Andrei V; Kazakov, Andrei A; Makarova, Alena V; Pavlov, Yuri I; Efremova, Anna S; Shram, Stanislav I; Tarantul, Viacheslav Z; Gening, Leonid V

2012-02-01

Human DNA-polymerase iota (Pol ι) is an extremely error-prone enzyme and the fidelity depends on the sequence context of the template. Using the in vitro systematic evolution of ligands by exponential enrichment (SELEX) procedure, we obtained an oligoribonucleotide with a high affinity to human Pol ι, named aptamer IKL5. We determined its dissociation constant with homogenous preparation of Pol ι and predicted its putative secondary structure. The aptamer IKL5 specifically inhibits DNA-polymerase activity of the purified enzyme Pol ι, but did not inhibit the DNA-polymerase activities of human DNA polymerases beta and kappa. IKL5 suppressed the error-prone DNA-polymerase activity of Pol ι also in cellular extracts of the tumor cell line SKOV-3. The aptamer IKL5 is useful for studies of the biological role of Pol ι and as a potential drug to suppress the increase of the activity of this enzyme in malignant cells.

Immobilized metal ion affinity electrophoresis. A study with several model proteins containing histidine.

PubMed

Goubran-Botros, H; Nanak, E; Abdul Nour, J; Birkenmeir, G; Vijayalakshmi, M A

1992-04-24

Immobilized metal ion affinity electrophoresis (IMA-Elec) is one among the many methods derived from the immobilized metal ion affinity chromatography. Two approaches for incorporating the metal ligand, were studied. One was in the form of insoluble particulate material based on Sepharose 6B and the other in the form of soluble polymer based on polyethylene glycol (PEG) 5000. Both the polymers coupled with iminodiacetate and metallized with copper or zinc were used as ligands, incorporated into soluble agarose as the electrophoretic gel. Several histidine-containing model proteins were studied with both the systems and their metal binding strengths were determined as the dissociation constants, Kd. The results clearly demonstrated that the mechanism of protein recognition by immobilized copper or zinc via the accessible histidyl residues was maintained in the IMA-Elec system. Proteins with increasing numbers of histidine residues showed increasing binding strength (lower Kd values). While this basic mechanism was conserved, the supporting polymers (Sepharose 6B and the PEG 5000) showed significant differences in the metal binding to the protein. The polysaccharide Sepharose 6B enhanced the binding strength compared with PEG 5000. The optimum electrophoretic parameters were determined to be current intensities up to 20 mA and pH ca. 7.0. At pH greater than 8.0, a significant decrease in the affinity was observed, this decrease being greater with PEG 5000 than Sepharose 6B as supporting material.

Method for accurate determination of dissociation constants of optical ratiometric systems: chemical probes, genetically encoded sensors, and interacting molecules.

PubMed

Pomorski, Adam; Kochańczyk, Tomasz; Miłoch, Anna; Krężel, Artur

2013-12-03

Ratiometric chemical probes and genetically encoded sensors are of high interest for both analytical chemists and molecular biologists. Their high sensitivity toward the target ligand and ability to obtain quantitative results without a known sensor concentration have made them a very useful tool in both in vitro and in vivo assays. Although ratiometric sensors are widely used in many applications, their successful and accurate usage depends on how they are characterized in terms of sensing target molecules. The most important feature of probes and sensors besides their optical parameters is an affinity constant toward analyzed molecules. The literature shows that different analytical approaches are used to determine the stability constants, with the ratio approach being most popular. However, oversimplification and lack of attention to detail results in inaccurate determination of stability constants, which in turn affects the results obtained using these sensors. Here, we present a new method where ratio signal is calibrated for borderline values of intensities of both wavelengths, instead of borderline ratio values that generate errors in many studies. At the same time, the equation takes into account the cooperativity factor or fluorescence artifacts and therefore can be used to characterize systems with various stoichiometries and experimental conditions. Accurate determination of stability constants is demonstrated utilizing four known optical ratiometric probes and sensors, together with a discussion regarding other, currently used methods.

Affine theory of gravitation

NASA Astrophysics Data System (ADS)

Popławski, Nikodem

2014-01-01

We propose a theory of gravitation, in which the affine connection is the only dynamical variable describing the gravitational field. We construct a simple dynamical Lagrangian density that is entirely composed from the connection, via its curvature and torsion, and is a polynomial function of its derivatives. It is given by the contraction of the Ricci tensor with a tensor which is inverse to the symmetric, contracted square of the torsion tensor, . We vary the total action for the gravitational field and matter with respect to the affine connection, assuming that the matter fields couple to the connection only through . We derive the resulting field equations and show that they are identical with the Einstein equations of general relativity with a nonzero cosmological constant if the tensor is regarded as proportional to the metric tensor. The cosmological constant is simply a constant of proportionality between the two tensors, which together with and provides a natural system of units in gravitational physics. This theory therefore provides a physical construction of the metric as a polynomial function of the connection, and explains dark energy as an intrinsic property of spacetime.

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.

Density Functional Study of Structures and Electron Affinities of BrO4F/BrO4F−

PubMed Central

Gong, Liangfa; Xiong, Jieming; Wu, Xinmin; Qi, Chuansong; Li, Wei; Guo, Wenli

2009-01-01

The structures, electron affinities and bond dissociation energies of BrO4F/BrO4F− species have been investigated with five density functional theory (DFT) methods with DZP++ basis sets. The planar F-Br…O2…O2 complexes possess 3A′ electronic state for neutral molecule and 4A′ state for the corresponding anion. Three types of the neutral-anion energy separations are the adiabatic electron affinity (EAad), the vertical electron affinity (EAvert), and the vertical detachment energy (VDE). The EAad value predicted by B3LYP method is 4.52 eV. The bond dissociation energies De (BrO4F → BrO4-mF + Om) (m = 1–4) and De− (BrO4F− → BrO4-mF− + Om and BrO4F− → BrO4-mF + Om−) are predicted. The adiabatic electron affinities (EAad) were predicted to be 4.52 eV for F-Br…O2…O2 (3A′←4A′) (B3LYP method). PMID:19742128

Binding affinities of anti-acetylcholine receptor autoantibodies in myasthenia gravis

DOE Office of Scientific and Technical Information (OSTI.GOV)

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 rangemore » 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.« less

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

DOE Office of Scientific and Technical Information (OSTI.GOV)

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,more » 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.« less

A rigorous multiple independent binding site model for determining cell-based equilibrium dissociation constants.

PubMed

Drake, Andrew W; Klakamp, Scott L

2007-01-10

A new 4-parameter nonlinear equation based on the standard multiple independent binding site model (MIBS) is presented for fitting cell-based ligand titration data in order to calculate the ligand/cell receptor equilibrium dissociation constant and the number of receptors/cell. The most commonly used linear (Scatchard Plot) or nonlinear 2-parameter model (a single binding site model found in commercial programs like Prism(R)) used for analysis of ligand/receptor binding data assumes only the K(D) influences the shape of the titration curve. We demonstrate using simulated data sets that, depending upon the cell surface receptor expression level, the number of cells titrated, and the magnitude of the K(D) being measured, this assumption of always being under K(D)-controlled conditions can be erroneous and can lead to unreliable estimates for the binding parameters. We also compare and contrast the fitting of simulated data sets to the commonly used cell-based binding equation versus our more rigorous 4-parameter nonlinear MIBS model. It is shown through these simulations that the new 4-parameter MIBS model, when used for cell-based titrations under optimal conditions, yields highly accurate estimates of all binding parameters and hence should be the preferred model to fit cell-based experimental nonlinear titration data.

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.

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

Rate constants for proteins binding to substrates with multiple binding sites using a generalized forward flux sampling expression

NASA Astrophysics Data System (ADS)

Vijaykumar, Adithya; ten Wolde, Pieter Rein; Bolhuis, Peter G.

2018-03-01

To predict the response of a biochemical system, knowledge of the intrinsic and effective rate constants of proteins is crucial. The experimentally accessible effective rate constant for association can be decomposed in a diffusion-limited rate at which proteins come into contact and an intrinsic association rate at which the proteins in contact truly bind. Reversely, when dissociating, bound proteins first separate into a contact pair with an intrinsic dissociation rate, before moving away by diffusion. While microscopic expressions exist that enable the calculation of the intrinsic and effective rate constants by conducting a single rare event simulation of the protein dissociation reaction, these expressions are only valid when the substrate has just one binding site. If the substrate has multiple binding sites, a bound enzyme can, besides dissociating into the bulk, also hop to another binding site. Calculating transition rate constants between multiple states with forward flux sampling requires a generalized rate expression. We present this expression here and use it to derive explicit expressions for all intrinsic and effective rate constants involving binding to multiple states, including rebinding. We illustrate our approach by computing the intrinsic and effective association, dissociation, and hopping rate constants for a system in which a patchy particle model enzyme binds to a substrate with two binding sites. We find that these rate constants increase as a function of the rotational diffusion constant of the particles. The hopping rate constant decreases as a function of the distance between the binding sites. Finally, we find that blocking one of the binding sites enhances both association and dissociation rate constants. Our approach and results are important for understanding and modeling association reactions in enzyme-substrate systems and other patchy particle systems and open the way for large multiscale simulations of such systems.

Identification of Elf-1 and B61 as high affinity ligands for the receptor tyrosine kinase MDK1.

PubMed

Ciossek, T; Ullrich, A

1997-01-09

Mouse Developmental Kinase 1 (MDK1) is a receptor tyrosine kinase of the eck/eph subfamily expressed in a variety of tissues during early mouse embryogenesis. To obtain further insight into the function of MDK1, we determined identity and localisation of its physiological ligand(s). Staining whole embryos with fusion proteins between the extracellular domain of MDK1 and human secreted alkaline phosphatase revealed areas of high receptor binding in the caudal mesencephalon, the frontal neocortex and the limb buds. This staining was sensitive to treatment with phosphatidylinositol-specific phospholipase C. Using Scatchard analysis, high affinity binding of Elf-1 (1.7 x 10(-10) M) and B61 (2.2 x 10(-10) M) towards MDK1 could be demonstrated. However, the transmembrane ligand Lerk2 displayed no measurable affinity for MDK1. Elf-1 and B61 bind to the three full-length MDK1 isoforms with similar dissociation constants. Slightly lower affinities were observed for the two truncated receptors MDK1-Tl and MDK1-T2. The activation of MDK1 with Elf-1 or B61 leads to the rapid autophosphorylation of MDK1 as well as tyrosine phosphorylation of an unknown 62 kDa phosphoprotein in Rat1 cells. These findings implicate MDK1 in patterning processes during early mouse embryogenesis and suggest MDK1 involvement in early organogenesis and midbrain development.

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

PubMed Central

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

2016-01-01

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. PMID:27609516

Improving the affinity of an antibody for its antigen via long-range electrostatic interactions.

PubMed

Fukunaga, Atsushi; Tsumoto, Kouhei

2013-12-01

To address how long-range electrostatic force can affect antibody-antigen binding, we focused on the interactions between human cardiac troponin I and its specific single-chain antibodies (scFvs). We first isolated two scFvs against two linear epitopes with distinct isoelectric points. For the scFv against the acidic epitope (A1scFv), we mutated five residues of framework region 3 of the light chain to Lys or Arg, designated as the K- or R-mutant, respectively. For the scFv against the basic epitope (A2scFv), we mutated four or three residues in framework region 3 of the light or heavy chain to Asp, to generate the VL- and VH-mutant, respectively. Surface plasmon resonance analyses showed that the kon values of all of the mutants were greater than that of wild type, even though framework region 3 does not make direct contact with the epitope. The affinity of the K-mutant was pM range, and that of the R-mutant improved further by more than two orders of magnitude due to a decrease in the dissociation rate constant. For the A2scFv mutants, the affinity of the VL-mutant for its target improved through an increase in the kon value without a decrease in the koff value. The stability slightly decreased in all mutants. These results suggest that introducing electrostatic interaction can improve the affinity of an antibody for its target, even if the mutation reduces stability of the antibody.

Impaired binding affinity of electronegative low-density lipoprotein (LDL) to the LDL receptor is related to nonesterified fatty acids and lysophosphatidylcholine content.

PubMed

Benítez, Sonia; Villegas, Virtudes; Bancells, Cristina; Jorba, Oscar; González-Sastre, Francesc; Ordóñez-Llanos, Jordi; Sánchez-Quesada, José Luis

2004-12-21

The binding characteristics of electropositive [LDL(+)] and electronegative LDL [LDL(-)] subfractions to the LDL receptor (LDLr) were studied. Saturation kinetic studies in cultured human fibroblasts demonstrated that LDL(-) from normolipemic (NL) and familial hypercholesterolemic (FH) subjects had lower binding affinity than their respective LDL(+) fractions (P < 0.05), as indicated by higher dissociation constant (K(D)) values. FH-LDL(+) also showed lower binding affinity (P < 0.05) than NL-LDL(+) (K(D), sorted from lower to higher affinity: NL-LDL(-), 33.0 +/- 24.4 nM; FH-LDL(-), 24.4 +/- 7.1 nM; FH-LDL(+), 16.6 +/- 7.0 nM; NL-LDL(+), 10.9 +/- 5.7 nM). These results were confirmed by binding displacement studies. The impaired affinity binding of LDL(-) could be attributed to altered secondary and tertiary structure of apolipoprotein B, but circular dichroism (CD) and tryptophan fluorescence (TrpF) studies revealed no structural differences between LDL(+) and LDL(-). To ascertain the role of increased nonesterified fatty acids (NEFA) and lysophosphatidylcholine (LPC) content in LDL(-), LDL(+) was enriched in NEFA or hydrolyzed with secretory phospholipase A(2). Modification of LDL gradually decreased the affinity to LDLr in parallel to the increasing content of NEFA and/or LPC. Modified LDLs with a NEFA content similar to that of LDL(-) displayed similar affinity. ApoB structure studies of modified LDLs by CD and TrpF showed no difference compared to LDL(+) or LDL(-). Our results indicate that NEFA loading or phospholipase A(2) lipolysis of LDL leads to changes that affect the affinity of LDL to LDLr with no major effect on apoB structure. Impaired affinity to the LDLr shown by LDL(-) is related to NEFA and/or LPC content rather than to structural differences in apolipoprotein B.

Predicting both passive intestinal absorption and the dissociation constant toward albumin using the PAMPA technique.

PubMed

Bujard, Alban; Sol, Marine; Carrupt, Pierre-Alain; Martel, Sophie

2014-10-15

The parallel artificial membrane permeability assay (PAMPA) is a high-throughput screening (HTS) method that is widely used to predict in vivo passive permeability through biological barriers, such as the skin, the blood brain barrier (BBB) and the gastrointestinal tract (GIT). The PAMPA technique has also been used to predict the dissociation constant (Kd) between a compound and human serum albumin (HSA) while disregarding passive permeability. Furthermore, the assay is based on the use of two separate 5-point kinetic experiments, which increases the analysis time. In the present study, we adapted the hexadecane membrane (HDM)-PAMPA assay to both predict passive gastrointestinal absorption via the permeability coefficient logPe value and determine the Kd. Two assays were performed: one in the presence and one in the absence of HSA in the acceptor compartment. In the absence of HSA, logPe values were determined after a 4-h incubation time, as originally described, but the dimethylsulfoxide (DMSO) percentage and pH were altered to be compatible with the protein. In parallel, a second PAMPA assay was performed in the presence of HSA during a 16-h incubation period. By adding HSA, a variation in the amount of compound crossing the membrane was observed compared to the permeability measured in the absence of HSA. The concentration of compound reaching the acceptor compartment in each case was used to determine both parameters (logPe and logKd) using numerical simulations, which highlighted the originality of this method because these calculations required only two endpoint measurements instead of a complete kinetic study. It should be noted that the amount of compound that reaches the acceptor compartment in the presence of HSA is modulated by complex dissociation in the receptor compartment. Only compounds that are moderately bound to albumin (-3

Tandem mass spectrometry of large biomolecule ions by blackbody infrared radiative dissociation.

PubMed

Price, W D; Schnier, P D; Williams, E R

1996-03-01

A new method for the dissociation of large ions formed by electrospray ionization is demonstrated. Ions trapped in a Fourier transform mass spectrometer at pressures below 10(-)(8) Torr are dissociated by elevating the vacuum chamber to temperatures up to 215 °C. Rate constants for dissociation are measured and found to be independent of pressure below 10(-)(7) Torr. This indicates that the ions are activated by absorption of blackbody radiation emitted from the chamber walls. Dissociation efficiencies as high as 100% are obtained. There is no apparent mass limit to this method; ions as large as ubiquitin (8.6 kDa) are readily dissociated. Thermally stable ions, such as melittin 3+ (2.8 kDa), did not dissociate at temperatures up to 200 °C. This method is highly selective for low-energy fragmentation, from which limited sequence information can be obtained. From the temperature dependence of the dissociation rate constants, Arrhenius activation energies in the low-pressure limit are obtained. The lowest energy dissociation processes for the singly and doubly protonated ions of bradykinin are loss of NH(3) and formation of the b(2)/y(7) complementary pair, with activation energies of 1.3 and 0.8 eV, respectively. No loss of NH(3) is observed for the doubly protonated ion; some loss of H(2)O occurs. These results show that charge-charge interactions not only lower the activation energy for dissociation but also can dramatically change the fragmentation, most likely through changes in the gas-phase conformation of the ion. Dissociation of ubiquitin ions produces fragmentation similar to that obtained by IRMPD and SORI-CAD. Higher charge state ions dissociate to produce y and b ions; the primary fragmentation process for low charge state ions is loss of H(2)O.

Hit-Validation Methodologies for Ligands Isolated from DNA-Encoded Chemical Libraries.

PubMed

Zimmermann, Gunther; Li, Yizhou; Rieder, Ulrike; Mattarella, Martin; Neri, Dario; Scheuermann, Jörg

2017-05-04

DNA-encoded chemical libraries (DECLs) are large collections of compounds linked to DNA fragments, serving as amplifiable barcodes, which can be screened on target proteins of interest. In typical DECL selections, preferential binders are identified by high-throughput DNA sequencing, by comparing their frequency before and after the affinity capture step. Hits identified in this procedure need to be confirmed, by resynthesis and by performing affinity measurements. In this article we present new methods based on hybridization of oligonucleotide conjugates with fluorescently labeled complementary oligonucleotides; these facilitate the determination of affinity constants and kinetic dissociation constants. The experimental procedures were demonstrated with acetazolamide, a binder to carbonic anhydrase IX with a dissociation constant in the nanomolar range. The detection of binding events was compatible not only with fluorescence polarization methodologies, but also with Alphascreen technology and with microscale thermophoresis. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

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.

Theoretical dissociation energies for the alkali and alkaline-earth monofluorides and monochlorides

NASA Technical Reports Server (NTRS)

Langhoff, S. R.; Bauschlicher, C. W., Jr.; Partridge, H.

1986-01-01

Spectroscopic parameters are accurately determined for the alkali and alkaline-earth monofluorides and monochlorides by means of ab initio self-consistent field and correlated wave function calculations. Numerical Hartree-Fock calculations are performed on selected systems to ensure that the extended Slater basis sets employed are near the Hartree-Fock limit. Since the bonding is predominantly electrostatic in origin, a strong correlation exists between the dissociation energy (to ions) and the spectroscopic parameter r(e). By dissociating to the ionic limits, most of the differential correlation effects can be embedded in the accurate experimental electron affinities and ionization potentials.

Swirling flow of a dissociated gas

NASA Technical Reports Server (NTRS)

Wolfram, W. R., Jr.; Walker, W. F.

1975-01-01

Most physical applications of the swirling flow, defined as a vortex superimposed on an axial flow in the nozzle, involve high temperatures and the possibility of real gas effects. The generalized one-dimensional swirling flow in a converging-diverging nozzle is analyzed for equilibrium and frozen dissociation using the ideal dissociating gas model. Numerical results are provided to illustrate the major effects and to compare with results obtained for a perfect gas with constant ratio of specific heats. It is found that, even in the case of real gases, perfect gas calculations can give a good estimate of the reduction in mass flow due to swirl.

Temperature Dependence of Dissociative Electron Attachment to Halogenated Hydrocarbons

NASA Astrophysics Data System (ADS)

Wang, Yicheng; Christophorou, Loucas G.

1996-10-01

Most of the gas mixtures currently in use for plasma processing of semiconductors involve halogenated hydrocarbons such as the strongly electronegative gases CCl4 and CFCl_3, the weakly electronegative gas CF_2Cl2 and the very weakly electronegative gases CHF3 and CF_4. Many dissociation processes are known to occur for these molecules. One of these dissociation reactions which is particularly effective for the strongly electronegative hydrocarbons is dissociative electron attachment. Even for weakly electron attaching gases, molecular dissociation via dissociative electron attachment at low energies can be an efficient dissociation process if the gas temperature is higher than ambient. Dissociative electron attachment is known to increase with increasing temperature above room temperature for many such compounds. In this paper, we report our measurements on the increases of the total electron attachment rate constant for CF_2Cl2 with increasing gas temperature from room temperature to about 600 K. -Research sponsored in part by the U.S. Air Force Wright Laboratory under contract F33615-96-C-2600 with the University of Tennessee. Also, Department of Physics, The University of Tennessee, Knoxville, TN.

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

Influence of Sulfolane on ESI-MS Measurements of Protein-Ligand Affinities

NASA Astrophysics Data System (ADS)

Yao, Yuyu; Richards, Michele R.; Kitova, Elena N.; Klassen, John S.

2016-03-01

The results of an investigation into the influence of sulfolane, a commonly used supercharging agent, on electrospray ionization mass spectrometry (ESI-MS) measurements of protein-ligand affinities are described. Binding measurements carried out on four protein-carbohydrate complexes, lysozyme with β- d-GlcNAc-(1→4)-β- d-GlcNAc-(1→4)-β- d-GlcNAc-(1→4)- d-GlcNAc, a single chain variable fragment and α- d-Gal-(1→2)-[α- d-Abe-(1→3)]-α- d-Man-OCH3, cholera toxin B subunit homopentamer with β- d-Gal-(1→3)-β- d-GalNAc-(1→4)[α- d-Neu5Ac-(2→3)]-β- d-Gal-(1→4)-β- d-Glc, and a fragment of galectin 3 and α- l-Fuc-(1→2)-β- d-Gal-(1→3)-β- d-GlcNAc-(1→3)-β- d-Gal-(1→4)-β- d-Glc, revealed that sulfolane generally reduces the apparent (as measured by ESI-MS) protein-ligand affinities. To establish the origin of this effect, a detailed study was undertaken using the lysozyme-tetrasaccharide interaction as a model system. Measurements carried out using isothermal titration calorimetry (ITC), circular dichroism, and nuclear magnetic resonance spectroscopies reveal that sulfolane reduces the binding affinity in solution but does not cause any significant change in the higher order structure of lysozyme or to the intermolecular interactions. These observations confirm that changes to the structure of lysozyme in bulk solution are not responsible for the supercharging effect induced by sulfolane. Moreover, the agreement between the ESI-MS and ITC-derived affinities indicates that there is no dissociation of the complex during ESI or in the gas phase (i.e., in-source dissociation). This finding suggests that supercharging of lysozyme by sulfolane is not related to protein unfolding during the ESI process. Binding measurements performed using liquid sample desorption ESI-MS revealed that protein supercharging with sulfolane can be achieved without a reduction in affinity.

Influence of Sulfolane on ESI-MS Measurements of Protein-Ligand Affinities.

PubMed

Yao, Yuyu; Richards, Michele R; Kitova, Elena N; Klassen, John S

2016-03-01

The results of an investigation into the influence of sulfolane, a commonly used supercharging agent, on electrospray ionization mass spectrometry (ESI-MS) measurements of protein-ligand affinities are described. Binding measurements carried out on four protein-carbohydrate complexes, lysozyme with β-D-GlcNAc-(1→4)-β-D-GlcNAc-(1→4)-β-D-GlcNAc-(1→4)-D-GlcNAc, a single chain variable fragment and α-D-Gal-(1→2)-[α-D-Abe-(1→3)]-α-D-Man-OCH3, cholera toxin B subunit homopentamer with β-D-Gal-(1→3)-β-D-GalNAc-(1→4)[α-D-Neu5Ac-(2→3)]-β-D-Gal-(1→4)-β-D-Glc, and a fragment of galectin 3 and α-L-Fuc-(1→2)-β-D-Gal-(1→3)-β-D-GlcNAc-(1→3)-β-D-Gal-(1→4)-β-D-Glc, revealed that sulfolane generally reduces the apparent (as measured by ESI-MS) protein-ligand affinities. To establish the origin of this effect, a detailed study was undertaken using the lysozyme-tetrasaccharide interaction as a model system. Measurements carried out using isothermal titration calorimetry (ITC), circular dichroism, and nuclear magnetic resonance spectroscopies reveal that sulfolane reduces the binding affinity in solution but does not cause any significant change in the higher order structure of lysozyme or to the intermolecular interactions. These observations confirm that changes to the structure of lysozyme in bulk solution are not responsible for the supercharging effect induced by sulfolane. Moreover, the agreement between the ESI-MS and ITC-derived affinities indicates that there is no dissociation of the complex during ESI or in the gas phase (i.e., in-source dissociation). This finding suggests that supercharging of lysozyme by sulfolane is not related to protein unfolding during the ESI process. Binding measurements performed using liquid sample desorption ESI-MS revealed that protein supercharging with sulfolane can be achieved without a reduction in affinity.

Fractal binding and dissociation kinetics of lecithin cholesterol acyl transferase (LCAT), a heart-related compound, on biosensor surfaces

NASA Astrophysics Data System (ADS)

Doke, Atul M.; Sadana, Ajit

2006-05-01

A fractal analysis is presented for the binding and dissociation of different heart-related compounds in solution to receptors immobilized on biosensor surfaces. The data analyzed include LCAT (lecithin cholesterol acyl transferase) concentrations in solution to egg-white apoA-I rHDL immobilized on a biosensor chip surface.1 Single- and dual- fractal models were employed to fit the data. Values of the binding and the dissociation rate coefficient(s), affinity values, and the fractal dimensions were obtained from the regression analysis provided by Corel Quattro Pro 8.0 (Corel Corporation Limited).2 The binding rate coefficients are quite sensitive to the degree of heterogeneity on the sensor chip surface. Predictive equations are developed for the binding rate coefficient as a function of the degree of heterogeneity present on the sensor chip surface and on the LCAT concentration in solution, and for the affinity as a function of the ratio of fractal dimensions present in the binding and the dissociation phases. The analysis presented provided physical insights into these analyte-receptor reactions occurring on different biosensor surfaces.

Sulfated Metabolites of Polychlorinated Biphenyls Are High-Affinity Ligands for the Thyroid Hormone Transport Protein Transthyretin

PubMed Central

Grimm, Fabian A.; Lehmler, Hans-Joachim; He, Xianran; Robertson, Larry W.

2013-01-01

Background: The displacement of l-thyroxine (T4) from binding sites on transthyretin (TTR) is considered a significant contributing mechanism in polychlorinated biphenyl (PCB)-induced thyroid disruption. Previous research has discovered hydroxylated PCB metabolites (OH-PCBs) as high-affinity ligands for TTR, but the binding potential of conjugated PCB metabolites such as PCB sulfates has not been explored. Objectives: We evaluated the binding of five lower-chlorinated PCB sulfates to human TTR and compared their binding characteristics to those determined for their OH-PCB precursors and for T4. Methods: We used fluorescence probe displacement studies and molecular docking simulations to characterize the binding of PCB sulfates to TTR. The stability of PCB sulfates and the reversibility of these interactions were characterized by HPLC analysis of PCB sulfates after their binding to TTR. The ability of OH-PCBs to serve as substrates for human cytosolic sulfotransferase 1A1 (hSULT1A1) was assessed by OH-PCB–dependent formation of adenosine-3´,5´-diphosphate, an end product of the sulfation reaction. Results: All five PCB sulfates were able to bind to the high-affinity binding site of TTR with equilibrium dissociation constants (Kd values) in the low nanomolar range (4.8–16.8 nM), similar to that observed for T4 (4.7 nM). Docking simulations provided corroborating evidence for these binding interactions and indicated multiple high-affinity modes of binding. All OH-PCB precursors for these sulfates were found to be substrates for hSULT1A1. Conclusions: Our findings show that PCB sulfates are high-affinity ligands for human TTR and therefore indicate, for the first time, a potential relevance for these metabolites in PCB-induced thyroid disruption. PMID:23584369

Statistical modeling of competitive threshold collision-induced dissociation

NASA Astrophysics Data System (ADS)

Rodgers, M. T.; Armentrout, P. B.

1998-08-01

Collision-induced dissociation of (R1OH)Li+(R2OH) with xenon is studied using guided ion beam mass spectrometry. R1OH and R2OH include the following molecules: water, methanol, ethanol, 1-propanol, 2-propanol, and 1-butanol. In all cases, the primary products formed correspond to endothermic loss of one of the neutral alcohols, with minor products that include those formed by ligand exchange and loss of both ligands. The cross-section thresholds are interpreted to yield 0 and 298 K bond energies for (R1OH)Li+-R2OH and relative Li+ binding affinities of the R1OH and R2OH ligands after accounting for the effects of multiple ion-molecule collisions, internal energy of the reactant ions, and dissociation lifetimes. We introduce a means to simultaneously analyze the cross sections for these competitive dissociations using statistical theories to predict the energy dependent branching ratio. Thermochemistry in good agreement with previous work is obtained in all cases. In essence, this statistical approach provides a detailed means of correcting for the "competitive shift" inherent in multichannel processes.

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.

A nonaqueous potentiometric titration study of the dissociation of t-butyl methacrylate-methacrylic acid copolymers.

PubMed

Nakatani, Kiyoharu; Yamashita, Jun; Sekine, Tomomi; Toriumi, Minoru; Itani, Toshiro

2003-05-01

The dissociation of t-butyl methacrylate-methacrylic acid copolymers in dimethyl sulfoxide was analyzed by a nonaqueous potentiometric titration technique. The negative logarithm of the dissociation constant of the monomer unit of a methacrylic acid (MAA) monotonously increased with the increasing degree of dissociation corresponding to the titrant/MAA amount ratio, and was highly influenced by the copolymerization ratio. The results are discussed in terms of the suppression of the dissociation of MAA by a neighboring charged methacrylate anion unit.

Quantification of transcription factor-DNA binding affinity in a living cell

PubMed Central

Belikov, Sergey; Berg, Otto G.; Wrange, Örjan

2016-01-01

The apparent dissociation constant (Kd) for specific binding of glucocorticoid receptor (GR) and androgen receptor (AR) to DNA was determined in vivo in Xenopus oocytes. The total nuclear receptor concentration was quantified as specifically retained [3H]-hormone in manually isolated oocyte nuclei. DNA was introduced by nuclear microinjection of single stranded phagemid DNA, chromatin is then formed during second strand synthesis. The fraction of DNA sites occupied by the expressed receptor was determined by dimethylsulphate in vivo footprinting and used for calculation of the receptor-DNA binding affinity. The forkhead transcription factor FoxA1 enhanced the DNA binding by GR with an apparent Kd of ∼1 μM and dramatically stimulated DNA binding by AR with an apparent Kd of ∼0.13 μM at a composite androgen responsive DNA element containing one FoxA1 binding site and one palindromic hormone receptor binding site known to bind one receptor homodimer. FoxA1 exerted a weak constitutive- and strongly cooperative DNA binding together with AR but had a less prominent effect with GR, the difference reflecting the licensing function of FoxA1 at this androgen responsive DNA element. PMID:26657626

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

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.

Determination of thermodynamic affinities of various polar olefins as hydride, hydrogen atom, and electron acceptors in acetonitrile.

PubMed

Cao, Ying; Zhang, Song-Chen; Zhang, Min; Shen, Guang-Bin; Zhu, Xiao-Qing

2013-07-19

A series of 69 polar olefins with various typical structures (X) were synthesized and the thermodynamic affinities (defined in terms of the molar enthalpy changes or the standard redox potentials in this work) of the polar olefins obtaining hydride anions, hydrogen atoms, and electrons, the thermodynamic affinities of the radical anions of the polar olefins (X(•-)) obtaining protons and hydrogen atoms, and the thermodynamic affinities of the hydrogen adducts of the polar olefins (XH(•)) obtaining electrons in acetonitrile were determined using titration calorimetry and electrochemical methods. The pure C═C π-bond heterolytic and homolytic dissociation energies of the polar olefins (X) in acetonitrile and the pure C═C π-bond homolytic dissociation energies of the radical anions of the polar olefins (X(•-)) in acetonitrile were estimated. The remote substituent effects on the six thermodynamic affinities of the polar olefins and their related reaction intermediates were examined using the Hammett linear free-energy relationships; the results show that the Hammett linear free-energy relationships all hold in the six chemical and electrochemical processes. The information disclosed in this work could not only supply a gap of the chemical thermodynamics of olefins as one class of very important organic unsaturated compounds but also strongly promote the fast development of the chemistry and applications of olefins.

Subunit dissociation in fish hemoglobins.

PubMed

Edelstein, S J; McEwen, B; Gibson, Q H

1976-12-10

solutions containing guanidine hydrochloride indicated that there are changes in both the absolute rates and the proportions of the fast and slow components, which along with other factors complicated the analysis of the data in terms of dissociation constants. Measurements were also made in solutions containing urea to promote dissociation, but with this agent very high concentrations (about 6 M) were required to give measureable dissociation and the fish hemoglobins were unstable under these conditions, with appreciable loss of absorbance spectra in both the sedimentation and kinetic experiments.

Dissociation of Heme–Globin Complexes by Blackbody Infrared Radiative Dissociation: Molecular Specificity in the Gas Phase?

PubMed Central

Gross, Deborah S.; Zhao, Yuexing; Williams, Evan R.

2005-01-01

The temperature dependence of the unimolecular kinetics for dissociation of the heme group from holo-myoglobin (Mb) and holo-hemoglobin α-chain (Hb-α) was investigated with blackbody infrared radiative dissociation (BIRD). The rate constant for dissociation of the 9 + charge state of Mb formed by electrospray ionization from a “pseudo-native” solution is 60% lower than that of Hb-α at each of the temperatures investigated. In solutions of pH 5.5–8.0, the thermal dissociation rate for Mb is also lower than that of HB-α (Hargrove, M. S. et al. J. Biol. Chem. 1994, 269, 4207–4214). Thus, Mb is thermally more stable with respect to heme loss than Hb-α both in the gas phase and in solution. The Arrhenius activation parameters for both dissociation processes are indistinguishable within the current experimental error (activation energy 0.9 eV and pre-exponential factor of 108–10 s−1). The 9+ to 12+ charge states of Mb have similar Arrhenius parameters when these ions are formed from pseudo-native solutions. In contrast, the activation energies and pre-exponential factors decrease from 0.8 to 0.3 eV and 107 to 102 s−1, respectively, for the 9 + to 12 + charge states formed from acidified solutions in which at least 50% of the secondary structure is lost. These results demonstrate that gas-phase Mb ions retain clear memory of the composition of the solution from which they are formed and that these differences can be probed by BIRD. PMID:16479269

Dissociation of heme-globin complexes by blackbody infrared radiative dissociation: molecular specificity in the gas phase?

PubMed

Gross, D S; Zhao, Y; Williams, E R

1997-05-01

The temperature dependence of the unimolecular kinetics for dissociation of the heme group from holo-myoglobin (Mb) and holo-hemoglobin alpha-chain (Hb-alpha) was investigated with blackbody infrared radiative dissociation (BIRD). The rate constant for dissociation of the 9 + charge state of Mb formed by electrospray ionization from a "pseudo-native" solution is 60% lower than that of Hb-alpha at each of the temperatures investigated. In solutions of pH 5.5-8.0, the thermal dissociation rate for Mb is also lower than that of HB-alpha (Hargrove, M. S. et al. J. Biol. Chem.1994, 269, 4207-4214). Thus, Mb is thermally more stable with respect to heme loss than Hb-alpha both in the gas phase and in solution. The Arrhenius activation parameters for both dissociation processes are indistinguishable within the current experimental error (activation energy 0.9 eV and pre-exponential factor of 10(8-10) s(-1)). The 9+ to 12+ charge states of Mb have similar Arrhenius parameters when these ions are formed from pseudo-native solutions. In contrast, the activation energies and pre-exponential factors decrease from 0.8 to 0.3 eV and 10(7) to 10(2) s(-1), respectively, for the 9 + to 12 + charge states formed from acidified solutions in which at least 50% of the secondary structure is lost. These results demonstrate that gas-phase Mb ions retain clear memory of the composition of the solution from which they are formed and that these differences can be probed by BIRD.

Nitrocellulose-bound antigen repeatedly used for the affinity purification of specific polyclonal antibodies for screening DNA expression libraries.

PubMed

Robinson, P A; Anderton, B H; Loviny, T L

1988-04-06

We present a simple, efficient and rapid method for affinity-purifying antibodies from a relatively crude antiserum in quantities large enough to screen a DNA expression library. The method presents a very convenient way to remove crossreacting or contaminating antibody specificities. The affinity matrix, antigen non-covalently bound to nitrocellulose, is prepared by the electrophoretic separation of antigen by sodium dodecyl sulphate-polyacrylamide gel electrophoresis, followed by the transfer of antigen to nitrocellulose. The matrix can be used repeatedly. A brief wash with 6 M guanidine hydrochloride is included between steps to remove residual antibodies which bind with high affinity to nitrocellulose-bound antigen. Various buffer solutions were assessed as antibody/antigen-dissociating agents. Glycine/HCl buffer, pH 2.5, appeared to be the most efficient in our hands, although a number of other less efficient dissociating reagents, including 4.5 M magnesium chloride, pH 7.5, 6 M urea, pH 7, and 0.05 M diethylamine, pH 11.5, also could be used; these may be the elution conditions of choice for other antibody/antigen combinations. The use of affinity-purified antibody solutions instead of the corresponding antisera gave increased signal-to-noise ratios with the detection systems that are commonly used to identify positive signals in screening expression libraries. Protein A- and goat anti-rabbit-alkaline phosphatase conjugates gave the most sensitive signals.

Surface Plasmon Resonance Biosensor Method for Palytoxin Detection Based on Na+,K+-ATPase Affinity

PubMed Central

Alfonso, Amparo; Pazos, María-José; Fernández-Araujo, Andrea; Tobio, Araceli; Alfonso, Carmen; Vieytes, Mercedes R.; Botana, Luis M.

2013-01-01

Palytoxin (PLTX), produced by dinoflagellates from the genus Ostreopsis was first discovered, isolated, and purified from zoanthids belonging to the genus Palythoa. The detection of this toxin in contaminated shellfish is essential for human health preservation. A broad range of studies indicate that mammalian Na+,K+-ATPase is a high affinity cellular receptor for PLTX. The toxin converts the pump into an open channel that stimulates sodium influx and potassium efflux. In this work we develop a detection method for PLTX based on its binding to the Na+,K+-ATPase. The method was developed by using the phenomenon of surface plasmon resonance (SPR) to monitor biomolecular reactions. This technique does not require any labeling of components. The interaction of PLTX over immobilized Na+,K+-ATPase is quantified by injecting different concentrations of toxin in the biosensor and checking the binding rate constant (kobs). From the representation of kobs versus PLTX concentration, the kinetic equilibrium dissociation constant (KD) for the PLTX-Na+,K+-ATPase association can be calculated. The value of this constant is KD = 6.38 × 10−7 ± 6.67 × 10−8 M PLTX. In this way the PLTX-Na+,K+-ATPase association was used as a suitable method for determination of the toxin concentration in a sample. This method represents a new and useful approach to easily detect the presence of PLTX-like compounds in marine products using the mechanism of action of these toxins and in this way reduce the use of other more expensive and animal based methods. PMID:24379088

Surface plasmon resonance biosensor method for palytoxin detection based on Na+,K+-ATPase affinity.

PubMed

Alfonso, Amparo; Pazos, María-José; Fernández-Araujo, Andrea; Tobio, Araceli; Alfonso, Carmen; Vieytes, Mercedes R; Botana, Luis M

2013-12-27

Palytoxin (PLTX), produced by dinoflagellates from the genus Ostreopsis was first discovered, isolated, and purified from zoanthids belonging to the genus Palythoa. The detection of this toxin in contaminated shellfish is essential for human health preservation. A broad range of studies indicate that mammalian Na+,K+-ATPase is a high affinity cellular receptor for PLTX. The toxin converts the pump into an open channel that stimulates sodium influx and potassium efflux. In this work we develop a detection method for PLTX based on its binding to the Na+,K+-ATPase. The method was developed by using the phenomenon of surface plasmon resonance (SPR) to monitor biomolecular reactions. This technique does not require any labeling of components. The interaction of PLTX over immobilized Na+,K+-ATPase is quantified by injecting different concentrations of toxin in the biosensor and checking the binding rate constant (Kobs). From the representation of Kobs versus PLTX concentration, the kinetic equilibrium dissociation constant (K(D)) for the PLTX-Na+,K+-ATPase association can be calculated. The value of this constant is K(D) = 6.38 × 10-7 ± 6.67 × 10-8 M PLTX. In this way the PLTX-Na+,K+-ATPase association was used as a suitable method for determination of the toxin concentration in a sample. This method represents a new and useful approach to easily detect the presence of PLTX-like compounds in marine products using the mechanism of action of these toxins and in this way reduce the use of other more expensive and animal based methods.

Synthesis, anticancer activity, and iron affinity of the Actinoplanes metabolite 7,8-dihydroxy-1-methylnaphtho[2,3-c]furan-4,9-dione.

PubMed

Breyer, Sandra; Effenberger-Neidnicht, Katharina; Knauer, Sebastian; Schobert, Rainer

2011-02-01

The first synthesis of 7,8-dihydroxy-1-methylnaphtho[2,3-c]furan-4,9-dione (1), an isofuranonaphthoquinone produced by an Actinoplanes strain is described. Lactone ring opening of 6-methylfuro[3,4-c]furan-1(3H)-one (4) with ortho-lithiated veratrole (3), oxidation of product alcohol 5, and Friedel-Crafts acylation of the resulting aroylcarboxylic acid 7 afforded the mono methyl ether 2 of the target compound. The latter was obtained by demethylation of 2 with BBr(3) in 14% overall yield. While mono ether 2 was distinctly more cytotoxic than catechol 1 against a panel of five cancer cell lines, only the latter showed a siderophore-like binding affinity for Fe(III) with a complex dissociation constant K(D) of approximately 10(-29) M(3) (pM = 25.9). Copyright © 2010 Elsevier Ltd. All rights reserved.

Partial filling affinity capillary electrophoresis as a useful tool for fragment-based drug discovery: A proof of concept on thrombin.

PubMed

Farcaş, E; Bouckaert, C; Servais, A-C; Hanson, J; Pochet, L; Fillet, M

2017-09-01

With the emergence of more challenging targets, a relatively new approach, fragment-based drug discovery (FBDD), proved its efficacy and gained increasing importance in the pharmaceutical industry. FBDD identifies low molecular-weight (MW) ligands (fragments) that bind to biologically important macromolecules, then a structure-guided fragment growing or merging approach is performed, contributing to the quality of the lead. However, to select the appropriate fragment to be evolved, sensitive analytical screening methods must be used to measure the affinity in the μM or even mM range. In this particular context, we developed a robust and selective partial filling affinity CE (ACE) method for the direct binding screening of a small fragment library in order to identify new thrombin inhibitors. To demonstrate the accuracy of our assay, the complex dissociation constants of three known thrombin inhibitors, namely benzamidine, p-aminobenzamidine and nafamostat were determined and found to be in good concordance with the previously reported values. Finally, the screening of a small library was performed and demonstrated the high discriminatory power of our method towards weak binders compared to classical spectrophotometric activity assay, proving the interest of our method in the context of FBDD. Copyright © 2017 Elsevier B.V. All rights reserved.

The 6300 A O/1-D/ airglow and dissociative recombination

NASA Technical Reports Server (NTRS)

Wickwar, V. B.; Cogger, L. L.; Carlson, H. C.

1974-01-01

Measurements of night-time 6300 A airglow intensities at the Arecibo Observatory have been compared with dissociative recombination calculations based on electron densities derived from simultaneous incoherent backscatter measurements. The agreement indicates that the nightglow can be fully accounted for by dissociative recombination. The comparisons are examined to determine the importance of quenching, heavy ions, ionization above the F-layer peak, and the temperature parameter of the model atmosphere. Comparable fits between the observed and calculated intensities are found for several available model atmospheres. The least-squares fitting process, used to make the comparisons, produces comparable fits over a wide range of combinations of neutral densities and of reaction constants. Yet, the fitting places constraints upon the possible combinations; these constraints indicate that the latest laboratory chemical constants and densities extrapolated to a base altitude are mutually consistent.

Psychiatric symptoms and dissociation in conversion, somatization and dissociative disorders.

PubMed

Espirito-Santo, Helena; Pio-Abreu, Jose Luis

2009-03-01

Conversion, dissociation and somatization are historically related in the long established concept of hysteria. Somewhere along the way they were separated due to the Cartesian dualistic view. The aim of the present study was to compare these pathologies and investigate whether symptoms of these pathologies overlap in their clinical appearance in a Portuguese sample. Twenty-six patients with conversion disorder, 38 with dissociative disorders, 40 with somatization disorder, and a comparison group of 46 patients having other psychiatric disorders answered questions about dissociation (Dissociative Experiences Scale), somatoform dissociation (Somatoform Dissociation Questionnaire), and psychopathological symptoms (Brief Symptom Inventory). Dissociative and somatoform symptoms were significantly more frequent in dissociative and conversion disorder than in somatization disorder and controls. There were no significant differences between dissociative and conversion patients. Conversion disorder is closely related to dissociative disorders. These results support the ICD-10 categorization of conversion disorder among dissociative disorders and the hypothesis of analogous psychopathological processes in conversion and dissociative disorders versus somatization disorder.

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.

Development and validation of an affinity chromatography step using a peptide ligand for cGMP production of factor VIII.

PubMed

Kelley, Brian D; Tannatt, Molly; Magnusson, Robert; Hagelberg, Sigrid; Booth, James

2004-08-05

An affinity chromatography step was developed for purification of recombinant B-Domain Deleted Factor VIII (BDDrFVIII) using a peptide ligand selected from a phage display library. The peptide library had variegated residues, contained both within a disulfide bond-constrained ring and flanking the ring. The peptide ligand binds to BDDrFVIII with a dissociation constant of approximately 1 microM both in free solution and when immobilized on a chromatographic resin. The peptide is chemically synthesized and the affinity resin is produced by coupling the peptide to an agarose matrix preactivated with N-hydroxysuccinimide. Coupling conditions were optimized to give consistent and complete ligand incorporation and validated with a robustness study that tested various combinations of processing limits. The peptide affinity chromatographic operation employs conditions very similar to an immunoaffinity chromatography step currently in use for BDDrFVIII manufacture. The process step provides excellent recovery of BDDrFVIII from a complex feed stream and reduces host cell protein and DNA by 3-4 logs. Process validation studies established resin reuse over 26 cycles without changes in product recovery or purity. A robustness study using a factorial design was performed and showed that the step was insensitive to small changes in process conditions that represent normal variation in commercial manufacturing. A scaled-down model of the process step was qualified and used for virus removal studies. A validation package addressing the safety of the leached peptide included leaching rate measurements under process conditions, testing of peptide levels in product pools, demonstration of robust removal downstream by spiking studies, end product testing, and toxicological profiling of the ligand. The peptide ligand affinity step was scaled up for cGMP production of BDDrFVIII for clinical trials.

Quantum gravity in the Eddington purely affine picture

NASA Astrophysics Data System (ADS)

Martellini, M.

1984-06-01

It was shown by Kijowski and Tulczjew that pure gravity with a cosmological constant can be obtained by a covariant Legendre transformation of a purely affine Lagrangian "in the manner of Eddington" constructed from a symmetric linear connection. In this paper I prove by explicit calculations that the Eddington Lagrangian is equivalent, in the sense which gives the same field equations, to a polynomial effective Lagrangian which turns out to be power-counting renormalizable. Then a formal proof of the unitarity of this theory is stated in the Kugo-Ojima formalism on the basis of the existence of two local Becchi-Rouet-Stora symmetries. These supertransformations are related to the algebra of the diffeomorphisms of the space-time, as well as to that of the volume-preserving space-time transformations which are not fixed by the gauge fixing used for the diffeomorphism group itself. Furthermore, I find that in the purely affine picture quantum gravity exhibits an infrared freedom. Since now the self-coupling constant is given by the cosmological constant, such a property could explain the observed almost zero value of the cosmological term at very large distances, i.e., to very low energies.

Antigen-Dissociation from Antibody-Modified Nanotransistor Sensor Arrays as a Direct Biomarker Detection Method in Unprocessed Biosamples.

PubMed

Krivitsky, Vadim; Zverzhinetsky, Marina; Patolsky, Fernando

2016-10-12

The detection of biomolecules is critical for a wide spectrum of applications in life sciences and medical diagnosis. Nonetheless, biosamples are highly complex solutions, which contain an enormous variety of biomolecules, cells, and chemical species. Consequently, the intrinsic chemical complexity of biosamples results in a significant analytical background noise and poses an immense challenge to any analytical measurement, especially when applied without prior efficient separation and purification steps. Here, we demonstrate the application of antigen-dissociation regime, from antibody-modified Si-nanowire sensors, as a simple and effective direct sensing mechanism of biomarkers of interest in complex biosamples, such as serum and untreated blood, which does not require ex situ time-consuming biosample manipulation steps, such as centrifugation, filtering, preconcentration, and desalting, thus overcoming the detrimental Debye screening limitation of nanowire-based biosensors. We found that two key parameters control the capability to perform quantitative biomarkers analysis in biosamples: (i) the affinity strength (k off rate) of the antibody-antigen recognition pair, which dictates the time length of the high-affinity slow dissociation subregime, and (ii) the "flow rate" applied during the solution exchange dissociation step, which controls the time width of the low-affinity fast-dissociation subregime. Undoubtedly, this is the simplest and most convenient approach for the SiNW FET-based detection of antigens in complex untreated biosamples. The lack of ex situ biosample manipulation time-consuming processes enhances the portability of the sensing platform and reduces to minimum the required volume of tested sample, as it allows the direct detection of untreated biosamples (5-10 μL blood or serum), while readily reducing the detection cycle duration to less than 5 min, factors of great importance in near-future point-of-care medical applications. We believe

Label-free Fab and Fc affinity/avidity profiling of the antibody complex half-life for polyclonal and monoclonal efficacy screening.

PubMed

Read, Thomas; Olkhov, Rouslan V; Williamson, E Diane; Shaw, Andrew M

2015-09-01

A unified approach to affinity screening for Fab and Fc interactions of an antibody for its antigen and FcγR receptor has been developed. An antigen array is used for the Fab affinity and cross-reactivity screening and protein A/G proxy is the FcγR receptor. The affinities are derived using a simple 1:1 binding model with a consistent error analysis. The association and dissociation kinetics are measured over optimised times for accurate determination. The Fab/Fc affinities are derived for ten antibodies: mAb-actin (mouse), pAb-BSA (sheep), pAb-collagen V (rabbit), pAb-CRP (goat), mAb-F1 (mouse), mAbs (mouse) 7.3, 12.3, 29.3, 36.3 and 46.3 raised against LcrV in Yersinia pestis. The rate of the dissociation of antigen-antibody complexes relates directly to their immunological function as does the Fc-FcγR complex and a new half-life plot has been defined with a Fab/Fc half-life range of 17-470 min. The upper half-life value points to surface avidity. Two antibodies that are protective as an immunotherapy define a Fab half-life >250 min and an Fc half-life >50 min as characteristics of ideal interactions which can form the basis of an antibody screen for immunotherapy.

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.

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

PubMed Central

Bao, Junwei Lucas; Zhang, Xin

2016-01-01

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. PMID:27834727

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 (C 2 F 4 ), 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.

Molecular dynamics simulation of low dielectric constant polymer electrolytes

NASA Astrophysics Data System (ADS)

Wheatle, Bill; Lynd, Nathaniel; Ganesan, Venkat

Recent experimental studies measured the ionic conductivities of a series of poly(glycidyl ether)s with varying neat dielectric constants (ɛ), viscosities (η), and glass transition temperatures (Tg), as hosts for lithium bistrifluoromethanesulfonimide (LiTFSI) salt. In such a context, it was demonstrated that the ionic conductivity of these polymer electrolytes was a function of ɛ rather than Tg or η, suggesting that there may exist regimes in which ionic conductivity is not limited by slow segmental dynamics but rather by low ionic dissociation. Motivated by such results, we used atomistic molecular dynamics to study the structure and transport characteristics of the same set of host polymers. We found that the coordination number of TFSI- about Li+ in the first solvation shell and the total fraction of free ions increased as a function of ɛ, implying the polymer hosts enhanced ion dissociation. In addition, we found that increasing the dielectric constant of the host polymer enhanced self-correlated ion transport, as evidenced by an increase in the diffusion coefficients of each ion species. Overall, we confirmed that limited ion dissociation in low- ɛ polymer electrolyte hosts hampers ionic conductivity. We would like to thank the National Science Foundation Graduate Research Fellowship Program for funding this research endeavor.

G-protein signaling leverages subunit-dependent membrane affinity to differentially control βγ translocation to intracellular membranes.

PubMed

O'Neill, Patrick R; Karunarathne, W K Ajith; Kalyanaraman, Vani; Silvius, John R; Gautam, N

2012-12-18

Activation of G-protein heterotrimers by receptors at the plasma membrane stimulates βγ-complex dissociation from the α-subunit and translocation to internal membranes. This intermembrane movement of lipid-modified proteins is a fundamental but poorly understood feature of cell signaling. The differential translocation of G-protein βγ-subunit types provides a valuable experimental model to examine the movement of signaling proteins between membranes in a living cell. We used live cell imaging, mathematical modeling, and in vitro measurements of lipidated fluorescent peptide dissociation from vesicles to determine the mechanistic basis of the intermembrane movement and identify the interactions responsible for differential translocation kinetics in this family of evolutionarily conserved proteins. We found that the reversible translocation is mediated by the limited affinity of the βγ-subunits for membranes. The differential kinetics of the βγ-subunit types are determined by variations among a set of basic and hydrophobic residues in the γ-subunit types. G-protein signaling thus leverages the wide variation in membrane dissociation rates among different γ-subunit types to differentially control βγ-translocation kinetics in response to receptor activation. The conservation of primary structures of γ-subunits across mammalian species suggests that there can be evolutionary selection for primary structures that confer specific membrane-binding affinities and consequent rates of intermembrane movement.

Quantitative analysis of the interactions between prenyl Rab9, GDP dissociation inhibitor-alpha, and guanine nucleotides.

PubMed

Shapiro, A D; Pfeffer, S R

1995-05-12

Rab9 is a Ras-like GTPase required for the transport of mannose 6-phosphate receptors between late endosomes and the trans Golgi network. Rab9 occurs in the cytosol as a complex with GDP dissociation inhibitor (GDI), which we have shown delivers prenyl Rab9 to late endosomes in a functional form. We report here basal rate constants for guanine nucleotide dissociation and GTP hydrolysis for prenyl Rab9. Both rate constants were influenced in part by the hydrophobic environment of the prenyl group. Guanine nucleotide dissociation and GTP hydrolysis rates were lower in the presence of lipid; detergent stimulated intrinsic nucleotide exchange. GDI-alpha inhibited GDP dissociation from prenyl Rab9 by 2.4-fold. GDI-alpha associated with prenyl Rab9 with a KD of 60 nM in 0.1% Lubrol and 23 nM in 0.02% Lubrol. In 0.1% Lubrol, GDI-alpha inhibited GDP dissociation half maximally at 72 +/- 18 nM, consistent with the KD determinations. These data suggest that GDI-alpha associates with prenyl Rab9 with a KD of < or = 23 nM under physiological conditions. Finally, a previously uncharacterized minor form of GDI-alpha inhibited GDP dissociation from prenyl Rab9 by 1.9-fold and bound prenyl Rab9 with a KD of 67 nM in 0.1% Lubrol.

Proteins feel more than they see: fine-tuning of binding affinity by properties of the non-interacting surface.

PubMed

Kastritis, Panagiotis L; Rodrigues, João P G L M; Folkers, Gert E; Boelens, Rolf; Bonvin, Alexandre M J J

2014-07-15

Protein-protein complexes orchestrate most cellular processes such as transcription, signal transduction and apoptosis. The factors governing their affinity remain elusive however, especially when it comes to describing dissociation rates (koff). Here we demonstrate that, next to direct contributions from the interface, the non-interacting surface (NIS) also plays an important role in binding affinity, especially polar and charged residues. Their percentage on the NIS is conserved over orthologous complexes indicating an evolutionary selection pressure. Their effect on binding affinity can be explained by long-range electrostatic contributions and surface-solvent interactions that are known to determine the local frustration of the protein complex surface. Including these in a simple model significantly improves the affinity prediction of protein complexes from structural models. The impact of mutations outside the interacting surface on binding affinity is supported by experimental alanine scanning mutagenesis data. These results enable the development of more sophisticated and integrated biophysical models of binding affinity and open new directions in experimental control and modulation of biomolecular interactions. Copyright © 2014. Published by Elsevier Ltd.

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. Copyright © 2014 Elsevier Ltd. All rights reserved.

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

DOE Office of Scientific and Technical Information (OSTI.GOV)

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 deglycosylatedmore » 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.« less

Single-cell measurement of red blood cell oxygen affinity.

PubMed

Di Caprio, Giuseppe; Stokes, Chris; Higgins, John M; Schonbrun, Ethan

2015-08-11

Oxygen is transported throughout the body by hemoglobin (Hb) in red blood cells (RBCs). Although the oxygen affinity of blood is well-understood and routinely assessed in patients by pulse oximetry, variability at the single-cell level has not been previously measured. In contrast, single-cell measurements of RBC volume and Hb concentration are taken millions of times per day by clinical hematology analyzers, and they are important factors in determining the health of the hematologic system. To better understand the variability and determinants of oxygen affinity on a cellular level, we have developed a system that quantifies the oxygen saturation, cell volume, and Hb concentration for individual RBCs in high throughput. We find that the variability in single-cell saturation peaks at an oxygen partial pressure of 2.9%, which corresponds to the maximum slope of the oxygen-Hb dissociation curve. In addition, single-cell oxygen affinity is positively correlated with Hb concentration but independent of osmolarity, which suggests variation in the Hb to 2,3-diphosphoglycerate (2-3 DPG) ratio on a cellular level. By quantifying the functional behavior of a cellular population, our system adds a dimension to blood cell analysis and other measurements of single-cell variability.

Single-cell measurement of red blood cell oxygen affinity

PubMed Central

Di Caprio, Giuseppe; Stokes, Chris; Higgins, John M.; Schonbrun, Ethan

2015-01-01

Oxygen is transported throughout the body by hemoglobin (Hb) in red blood cells (RBCs). Although the oxygen affinity of blood is well-understood and routinely assessed in patients by pulse oximetry, variability at the single-cell level has not been previously measured. In contrast, single-cell measurements of RBC volume and Hb concentration are taken millions of times per day by clinical hematology analyzers, and they are important factors in determining the health of the hematologic system. To better understand the variability and determinants of oxygen affinity on a cellular level, we have developed a system that quantifies the oxygen saturation, cell volume, and Hb concentration for individual RBCs in high throughput. We find that the variability in single-cell saturation peaks at an oxygen partial pressure of 2.9%, which corresponds to the maximum slope of the oxygen–Hb dissociation curve. In addition, single-cell oxygen affinity is positively correlated with Hb concentration but independent of osmolarity, which suggests variation in the Hb to 2,3-diphosphoglycerate (2–3 DPG) ratio on a cellular level. By quantifying the functional behavior of a cellular population, our system adds a dimension to blood cell analysis and other measurements of single-cell variability. PMID:26216973

Label-Free Determination of Protein Binding in Aqueous Solution using Overlayer Enhanced Attenuated Total Reflectance Fourier Transform Infrared Spectroscopy (OE-ATR-FTIR)

NASA Astrophysics Data System (ADS)

Ruthenburg, Travis; Aweda, Tolulope; Park, Simon; Meares, Claude; Land, Donald

2009-03-01

Protein binding/affinity studies are often performed using Surface Plasmon Resonance techniques that don't produce much spectral information. Measurement of protein binding affinity using FTIR is traditionally performed using high protein concentration or deuterated solvent. By immobilizing a protein near the surface of a gold-coated germanium internal reflection element interactions can be measured between an immobilized protein and free proteins or small molecules in aqueous solution. By monitoring the on and off rates of these interactions, the dissociation constant for the system can be determined. The dissociation constant for the molecule Yttrium-DOTA binding to the antibody 2D12.5 system was determined to be 100nM. Results will also be presented from our measurements of Bovine Serum Albumin (BSA) binding to anti-BSA.

Dissociative experiences and dissociative minds: Exploring a nomological network of dissociative functioning.

PubMed

Schimmenti, Adriano

2016-01-01

In this study, the psychometric properties of the Dissociative Experiences Scale-II (DES-II) were tested in a sample of Italian adults, and a nomological network of dissociative functioning based on current psychodynamic research was examined. A total of 794 participants (55% females) ranging in age from 18 to 64 completed the DES-II and other measures of theory of mind, alexithymia, attachment style, and empathy. The Italian translation of the DES-II showed high internal consistency, adequate item-to-scale homogeneity, and good split-half reliability. A single-factor solution including the 8 items of pathological dissociation (DES-T) adequately fit the data. Participants who reported higher levels of dissociative experiences showed significantly lower scores on theory of mind and empathy than other participants. They also showed significantly higher scores on alexithymia, preoccupied attachment, and fearful attachment. Results of the study support the view that people who suffer from severe dissociative experiences may also have difficulties mentalizing and regulating affects and that they may feel uncomfortable in close relationships because they have a negative view of the self. This can inform clinical work with dissociative individuals, who could benefit from therapies that consider their potential problems with mentalization, empathy, affect regulation, and attachment.

Ab Initio Calculations of the N-N Bond Dissociation for the Gas-phase RDX and HMX.

PubMed

Liu, Lin-Lin; Liu, Pei-Jin; Hu, Song-Qi; He, Guo-Qiang

2017-01-17

NO 2 fission is a vital factor for 1,3,5-Trinitroperhydro-1,3,5-triazine (RDX) and octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine (HMX) decomposition. In this study, the geometry of the gas-phase RDX and HMX molecules was optimized, and the bond order and the bond dissociation energy of the N-N bonds were examined. Moreover, the rate constants of the gas-phase RDX and HMX conformers, concerning the N-N bond dissociation, were evaluated using the microcanonical variational transition state theory (μVT). The calculation results have shown that HMX is more stable than RDX in terms of the N-N bond dissociation, and the conformers stability parameters were as follows: RDXaaa < RDXaae < HMX I < HMX II. In addition, for the RDX conformers, the N-N bond of the pseudo-equatorial positioning of the nitro group was more stable than the N-N bond of the axial positioning of the nitro group, while the results were opposite in the case of the HMX conformers. Moreover, it has been shown that the dissociation rate constant of the N-N bond is influenced by the temperature significantly, thus the rate constants were much lower (<10 -10  s -1 ) when the temperature was less than 1000 K.

Ab Initio Calculations of the N-N Bond Dissociation for the Gas-phase RDX and HMX

PubMed Central

Liu, Lin-lin; Liu, Pei-jin; Hu, Song-qi; He, Guo-qiang

2017-01-01

NO2 fission is a vital factor for 1,3,5-Trinitroperhydro-1,3,5-triazine (RDX) and octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine (HMX) decomposition. In this study, the geometry of the gas-phase RDX and HMX molecules was optimized, and the bond order and the bond dissociation energy of the N-N bonds were examined. Moreover, the rate constants of the gas-phase RDX and HMX conformers, concerning the N-N bond dissociation, were evaluated using the microcanonical variational transition state theory (μVT). The calculation results have shown that HMX is more stable than RDX in terms of the N-N bond dissociation, and the conformers stability parameters were as follows: RDXaaa < RDXaae < HMX I < HMX II. In addition, for the RDX conformers, the N-N bond of the pseudo-equatorial positioning of the nitro group was more stable than the N-N bond of the axial positioning of the nitro group, while the results were opposite in the case of the HMX conformers. Moreover, it has been shown that the dissociation rate constant of the N-N bond is influenced by the temperature significantly, thus the rate constants were much lower (<10−10 s−1) when the temperature was less than 1000 K. PMID:28094774

Ab Initio Calculations of the N-N Bond Dissociation for the Gas-phase RDX and HMX

NASA Astrophysics Data System (ADS)

Liu, Lin-Lin; Liu, Pei-Jin; Hu, Song-Qi; He, Guo-Qiang

2017-01-01

NO2 fission is a vital factor for 1,3,5-Trinitroperhydro-1,3,5-triazine (RDX) and octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine (HMX) decomposition. In this study, the geometry of the gas-phase RDX and HMX molecules was optimized, and the bond order and the bond dissociation energy of the N-N bonds were examined. Moreover, the rate constants of the gas-phase RDX and HMX conformers, concerning the N-N bond dissociation, were evaluated using the microcanonical variational transition state theory (μVT). The calculation results have shown that HMX is more stable than RDX in terms of the N-N bond dissociation, and the conformers stability parameters were as follows: RDXaaa < RDXaae < HMX I < HMX II. In addition, for the RDX conformers, the N-N bond of the pseudo-equatorial positioning of the nitro group was more stable than the N-N bond of the axial positioning of the nitro group, while the results were opposite in the case of the HMX conformers. Moreover, it has been shown that the dissociation rate constant of the N-N bond is influenced by the temperature significantly, thus the rate constants were much lower (<10-10 s-1) when the temperature was less than 1000 K.

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.

Blackbody infrared radiative dissociation of protonated oligosaccharides.

PubMed

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 (Cel(6)), which is composed of β-(1→4)-linked glucopyranose rings, and five malto-oligosaccharides (Mal(x), 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 (B(1) and B(2)) also occurs. Rate constants for dissociation of the protonated oligosaccharides and the corresponding Arrhenius activation parameters (E(a) and A) were determined. The E(a) and A-factors measured for protonated Mal(x) (x>4) are indistinguishable within error (~19 kcal mol(-1), 10(10) s(-1)), which is consistent with the ions being in the rapid energy exchange limit. In contrast, the Arrhenius parameters for protonated Cel(6) (24 kcal mol(-1), 10(12) 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. © American Society for Mass Spectrometry, 2011

Equations for calculating the properties of dissociated steam

NASA Astrophysics Data System (ADS)

Aminov, R. Z.; Gudym, A. A.

2017-08-01

The equations of state for dissociated steam have been developed in the temperature and pressure ranges of 1250-2300 K and 0.01-10.00 MPa for calculating thermodynamic processes in thermal power units operating on high-temperature steam. These equations are based on the property tables for dissociated steam derived at a reference temperature of 0 K. It is assumed that the initial substance is steam, the dissociation of which—in accordance with the most likely chemical reactions—results in formation of molecules of hydrogen, oxygen, steam, hydroxyl, and atoms of oxygen and hydrogen. Differential thermodynamic correlations, considering a change in the chemical potential and the composition of the mixture, during the steam dissociation are used. A reference temperature of 0.01°C used in the calculation of parameters of nondissociated steam has been adopted to predict processes in thermal power units without matching the reference temperatures and to account for transformation of dissociated steam into its usual form for which there is the international system of equations with the water triple point of 0.01°C taken as the reference. In the investigated region, the deviation of dissociated steam properties from those of nondissociated steam, which increases with decreasing the pressure or increasing the temperature, was determined. For a pressure of 0.02 MPa and a temperature of 2200 K, these deviations are 512 kJ/kg for the enthalpy, 0.2574 kJ/(kg K) for the entropy, and 3.431 kJ/(kg K) for the heat capacity at constant pressure. The maximum deviation of the dissociated steam properties calculated by the developed equations from the handbook values that these equations are based on does not exceed 0.03-0.05%.

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

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

PubMed Central

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

2009-01-01

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

The Interaction Affinity between Vascular Cell Adhesion Molecule-1 (VCAM-1) and Very Late Antigen-4 (VLA-4) Analyzed by Quantitative FRET

PubMed Central

Wu, Shu-Han; Karmenyan, Artashes; Chiou, Arthur

2015-01-01

Very late antigen-4 (VLA-4), a member of integrin superfamily, interacts with its major counter ligand vascular cell adhesion molecule-1 (VCAM-1) and plays an important role in leukocyte adhesion to vascular endothelium and immunological synapse formation. However, irregular expressions of these proteins may also lead to several autoimmune diseases and metastasis cancer. Thus, quantifying the interaction affinity of the VCAM-1/VLA-4 interaction is of fundamental importance in further understanding the nature of this interaction and drug discovery. In this study, we report an ‘in solution’ steady state organic fluorophore based quantitative fluorescence resonance energy transfer (FRET) assay to quantify this interaction in terms of the dissociation constant (Kd). We have used, in our FRET assay, the Alexa Fluor 488-VLA-4 conjugate as the donor, and Alexa Fluor 546-VCAM-1 as the acceptor. From the FRET signal analysis, Kd of this interaction was determined to be 41.82 ± 2.36 nM. To further confirm our estimation, we have employed surface plasmon resonance (SPR) technique to obtain Kd = 39.60 ± 1.78 nM, which is in good agreement with the result obtained by FRET. This is the first reported work which applies organic fluorophore based ‘in solution’ simple quantitative FRET assay to obtain the dissociation constant of the VCAM-1/VLA-4 interaction, and is also the first quantification of this interaction. Moreover, the value of Kd can serve as an indicator of abnormal protein-protein interactions; hence, this assay can potentially be further developed into a drug screening platform of VLA-4/VCAM-1 as well as other protein-ligand interactions. PMID:25793408

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

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

Electron affinities, molecular structures, and thermochemistry of the fluorine, chlorine and bromine substituted methyl radicals

NASA Astrophysics Data System (ADS)

Li, Qian-Shu; Zhao, Jun-Fang; Xie, Yaoming; Schaefer, Henry F., III

Four independent density functional theory (DFT) methods have been employed to study the structures and electron affinities of the methyl and F-, Cl- and Br-substituted methyl radicals and their anions. The methods used have been carefully calibrated against a comprehensive tabulation of experimental electron affinities (Chemical Reviews, 2002, 102, 231). The first dissociation energies together with the vibrational frequencies of these species are also reported. The basis sets used in this work are of double- ζ plus polarization quality with additional s- and p-type diffuse functions, labelled as DZP++. Previously observed trends in the prediction of bond lengths by the DFT methods are also demonstrated for the F-, Cl- and Br-substituted methyl radicals and their anions. Generally, the Hartree-Fock/DFT hybrid methods predict shorter and more reliable bond lengths than the pure DFT methods. Neutral-anion energy differences reported in this work are the adiabatic electron affinity (EAad), the vertical electron affinity (EAvert), and the vertical detachment energy (VDE). Compared with the available experimental electron affinities, the BHLYP method predicts much lower values, while the other methods predict values (EAad, EAvert, VDE) close to each other and almost within the experimental range. For those systems without reliable experimental measurements, our best adiabatic EAs predicted by BLYP are 0.78 (CHF2), 1.23 (CHFCl), 1.44 (CHFBr), 1.61 (CHClBr), 2.24 (CF2Cl), 2.42 (CF2Br), 2.56 (CFBr2), 2.36 (CCl2Br), 2.46 (CClBr2), and 2.44 eV (CFClBr). The most striking feature of these predictions is that they display an inverse relationship between halogen electronegativity and EA. The DZP++ B3LYP method determines the vibrational frequencies in best agreement with available experimental results for this series, with an average relative error of ~2%. The value of using a variety of DFT methods is observed in that BHLYP does best for geometries, BLYP for electron

Proton affinity determinations and proton-bound dimer structure indications in C2 to C15, (alpha),(omega)-alkyldiamines

NASA Technical Reports Server (NTRS)

Karpas, Z.; Harden, C. S.; Smith, P. B. W.

1995-01-01

The 'kinetic method' was used to determine the proton affinity (PA) of a,coalkyldiamines from collision induced dissociation (CID) studies of protonated heterodimers. These PA values were consistently lower than those reported in the proton affinity scale. The apparent discrepancy was rationalized in terms of differences in the conformation of the protonated diamine monomers. The minimum energy species, formed by equilibrium proton transfer processes, have a cyclic conformation and the ion charge is shared by both amino-groups which are bridged by the proton. On the other hand, the species formed through dissociation of protonated dimers have a linear structure and the charge is localized on one of the amino-groups. Thus, the difference in the PA values obtained by both methods is a measure of the additional stability acquired by the protonated diamines through cyclization and charge delocalization. The major collision dissociation pathway of the protonated diamine monomers involved elimination of an ammonia moiety. Other reactions observed included loss of the second amino-group and several other bond cleavages. CID of the protonated dimers involved primarily formation of a protonated monomer through cleavage of the weaker hydrogen bond and subsequently loss of ammonia at higher collision energies. As observed from the CID studies, doubly charged ions were also formed from the diamines under conditions of the electrospray ionization.

Dissociative electron attachment to C{sub 2}F{sub 5} radicals

DOE Office of Scientific and Technical Information (OSTI.GOV)

Haughey, Sean A.; Field, Thomas A.; Langer, Judith

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. Themore » rate constant increases slowly with increasing temperature, but the rise observed is smaller than the experimental uncertainty of 35%.« less

Analysis of Structural Features Contributing to Weak Affinities of Ubiquitin/Protein Interactions.

PubMed

Cohen, Ariel; Rosenthal, Eran; Shifman, Julia M

2017-11-10

Ubiquitin is a small protein that enables one of the most common post-translational modifications, where the whole ubiquitin molecule is attached to various target proteins, forming mono- or polyubiquitin conjugations. As a prototypical multispecific protein, ubiquitin interacts non-covalently with a variety of proteins in the cell, including ubiquitin-modifying enzymes and ubiquitin receptors that recognize signals from ubiquitin-conjugated substrates. To enable recognition of multiple targets and to support fast dissociation from the ubiquitin modifying enzymes, ubiquitin/protein interactions are characterized with low affinities, frequently in the higher μM and lower mM range. To determine how structure encodes low binding affinity of ubiquitin/protein complexes, we analyzed structures of more than a hundred such complexes compiled in the Ubiquitin Structural Relational Database. We calculated various structure-based features of ubiquitin/protein binding interfaces and compared them to the same features of general protein-protein interactions (PPIs) with various functions and generally higher affinities. Our analysis shows that ubiquitin/protein binding interfaces on average do not differ in size and shape complementarity from interfaces of higher-affinity PPIs. However, they contain fewer favorable hydrogen bonds and more unfavorable hydrophobic/charge interactions. We further analyzed how binding interfaces change upon affinity maturation of ubiquitin toward its target proteins. We demonstrate that while different features are improved in different experiments, the majority of the evolved complexes exhibit better shape complementarity and hydrogen bond pattern compared to wild-type complexes. Our analysis helps to understand how low-affinity PPIs have evolved and how they could be converted into high-affinity PPIs. Copyright © 2017 Elsevier Ltd. All rights reserved.

Labeling by ( sup 3 H)1,3-di(2-tolyl)guanidine of two high affinity binding sites in guinea pig brain: Evidence for allosteric regulation by calcium channel antagonists and pseudoallosteric modulation by sigma ligands

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rothman, R.B.; Reid, A.; Mahboubi, A.

1991-02-01

Equilibrium binding studies with the sigma receptor ligand ({sup 3}H)1,3-di(2-tolyl)guanidine (({sup 3}H)DTG) demonstrated two high affinity binding sites in membranes prepared from guinea pig brain. The apparent Kd values of DTG for sites 1 and 2 were 11.9 and 37.6 nM, respectively. The corresponding Bmax values were 1045 and 1423 fmol/mg of protein. Site 1 had high affinity for (+)-pentazocine, haloperidol, (R)-(+)-PPP, carbepentane, and other sigma ligands, suggesting a similarity with the dextromethorphan/sigma 1 binding site described by Musacchio et al. (Life Sci. 45:1721-1732 (1989)). Site 2 had high affinity for DTG and haloperidol (Ki = 36.1 nM) and lowmore » affinity for most other sigma ligands. Kinetic experiments demonstrated that ({sup 3}H)DTG dissociated in a biphasic manner from both site 1 and site 2. DTG and haloperidol increased the dissociation rate of ({sup 3}H)DTG from site 1 and site 2, demonstrating the presence of pseudoallosteric interactions. Inorganic calcium channel blockers such as Cd2+ selectively increased the dissociation rate of ({sup 3}H)DTG from site 2, suggesting an association of this binding site with calcium channels.« less

Synthesis, crystal structure, DFT studies, acid dissociation constant, and antimicrobial activity of methyl 2-(4-chlorophenyl)-7a-((4-chlorophenyl)carbamothioyl)-1-oxo-5,5-diphenyl-3-thioxo-hexahydro-1H-pyrrolo[1,2-e]imidazole-6-carboxylate

NASA Astrophysics Data System (ADS)

Nural, Yahya; Gemili, Muge; Seferoglu, Nurgul; Sahin, Ertan; Ulger, Mahmut; Sari, Hayati

2018-05-01

A novel bicyclic thiohydantoin fused to pyrrolidine compound, methyl 2-(4-chlorophenyl)-7a-((4-chlorophenyl)carbamothioyl)-1-oxo-5,5-diphenyl-3-thioxo-hexahydro-1H-pyrrolo[1,2-e]imidazole-6-carboxylate, was synthesized by the cyclization reaction of dimethyl 5,5-diphenylpyrrolidine-2,4-dicarboxylate and 4-chlorophenyl isothiocyanate in the presence of 4-(dimethylamino)pyridine to form methyl 2-(4-chlorophenyl)-1-oxo-5,5-diphenyl-3-thioxo-hexahydro-1H-pyrrolo[1,2-e]imidazole-6-carboxylate with concomitant addition reaction of the 4-chlorophenyl isothiocyanate in 79% yield. The structural characterization was performed by NMR, FT-IR, MS and HRMS techniques, and the stereochemistry of the compound was determined by single crystal X-ray diffraction study. In addition, the molecular structure and 1H and 13C NMR chemical shifts of the compound were obtained with the density functional theory and Hartree-Fock calculations. Acid dissociation constants of the compound were determined using potentiometric titration method in 25% (v/v) dimethyl sulfoxide-water hydroorganic solvent at 25 ± 0.1 °C, at an ionic background of 0.1 mol/L of NaCl using the HYPERQUAD computer program. Four acid dissociation constants were obtained for the compound, and we suggest that these acid dissociation constants are related to the NH, for two groups of enthiols and enol groups. Antimicrobial activity study was performed against S. aureus, B. subtilis, A. hydrophila, E. coli and A. baumannii as bacterial standard strains, and against M. tuberculosis H37Rv as mycobacterial strain. The compound exhibited antibacterial activity in the range of 31.25-62.5 μg/mL, and antimycobacterial activity with a MIC value of 40 μg/mL against the indicated strains.

The effective temperature of Peptide ions dissociated by sustained off-resonance irradiation collisional activation in fourier transform mass spectrometry.

PubMed

Schnier, P D; Jurchen, J C; Williams, E R

1999-01-28

A method for determining the internal energy of biomolecule ions activated by collisions is demonstrated. The dissociation kinetics of protonated leucine enkephalin and doubly protonated bradykinin were measured using sustained off-resonance irradiation (SORI) collisionally activated dissociation (CAD) in a Fourier transform mass spectrometer. Dissociation rate constants are obtained from these kinetic data. In combination with Arrhenius parameters measured with blackbody infrared radiative dissociation, the "effective" temperatures of these ions are obtained. Effects of excitation voltage and frequency and the ion cell pressure were investigated. With typical SORI-CAD experimental conditions, the effective temperatures of these peptide ions range between 200 and 400 degrees C. Higher temperatures can be easily obtained for ions that require more internal energy to dissociate. The effective temperatures of both protonated leucine enkephalin and doubly protonated bradykinin measured with the same experimental conditions are similar. Effective temperatures for protonated leucine enkephalin can also be obtained from the branching ratio of the b(4) and (M + H - H(2)O)(+) pathways. Values obtained from this method are in good agreement with those obtained from the overall dissociation rate constants. Protonated leucine enkephalin is an excellent "thermometer" ion and should be well suited to establishing effective temperatures of ions activated by other dissociation techniques, such as infrared photodissociation, as well as ionization methods, such as matrix assisted laser desorption/ionization.

The Effective Temperature of Peptide Ions Dissociated by Sustained Off-Resonance Irradiation Collisional Activation in Fourier Transform Mass Spectrometry

PubMed Central

Schnier, Paul D.; Jurchen, John C.; Williams, Evan R.

2005-01-01

A method for determining the internal energy of biomolecule ions activated by collisions is demonstrated. The dissociation kinetics of protonated leucine enkephalin and doubly protonated bradykinin were measured using sustained off-resonance irradiation (SORI) collisionally activated dissociation (CAD) in a Fourier transform mass spectrometer. Dissociation rate constants are obtained from these kinetic data. In combination with Arrhenius parameters measured with blackbody infrared radiative dissociation, the “effective” temperatures of these ions are obtained. Effects of excitation voltage and frequency and the ion cell pressure were investigated. With typical SORI–CAD experimental conditions, the effective temperatures of these peptide ions range between 200 and 400 °C. Higher temperatures can be easily obtained for ions that require more internal energy to dissociate. The effective temperatures of both protonated leucine enkephalin and doubly protonated bradykinin measured with the same experimental conditions are similar. Effective temperatures for protonated leucine enkephalin can also be obtained from the branching ratio of the b4 and (M + H − H2O)+ pathways. Values obtained from this method are in good agreement with those obtained from the overall dissociation rate constants. Protonated leucine enkephalin is an excellent “thermometer” ion and should be well suited to establishing effective temperatures of ions activated by other dissociation techniques, such as infrared photodissociation, as well as ionization methods, such as matrix assisted laser desorption/ionization. PMID:16614752

The eΠ3g state of C2: A pathway to dissociation

NASA Astrophysics Data System (ADS)

Welsh, B. A.; Krechkivska, O.; Nauta, K.; Bacskay, G. B.; Kable, S. H.; Schmidt, T. W.

2017-07-01

The lowest 13 vibrational levels, v = 0-12, of the eΠ3g state of the C2 molecule have been measured by laser-induced fluorescence of new bands of the Fox-Herzberg system. The newly observed levels, v = 5-12, which span the eΠ3g electronic state up to and beyond the first dissociation threshold of C2, were analyzed to afford highly accurate molecular constants, including band origins, and rotational and spin-orbit constants. The spin-orbit coupling constants of the previously published lowest five levels are revised in sign and magnitude, requiring an overhaul of previously published molecular constants. The analysis is supported by high level ab initio calculations. Lifetimes of all observed levels were recorded and found to be in excellent agreement with ab initio predicted values up to v = 11. v = 12 was found to exhibit a much reduced lifetime and fluorescence quantum yield, which is attributed to the onset of predissociation. This brackets the dissociation energy of ground state XΣ+1g C2 between 6.1803 and 6.2553 eV, in agreement with the Active Thermochemical Tables.

Yeast hexokinase: substrate-induced association--dissociation reactions in the binding of glucose to hexokinase P-II.

PubMed

Hoggett, J G; Kellett, G L

1976-06-15

A method is described for the purification of native hexokinases P-I and P-II from yeast using preparative isoelectric focussing to separate the isozymes. The binding of glucose to hexokinase P-II, and the effect of this on the monomer--dimer association--dissociation reaction have been investigated quantitatively by a combination of titrations of intrinsic protein fluorescence and equilibrium ultracentrifugation. Association constants for the monomer-dimer reaction decreased with increasing pH, ionic strength and concentration of glucose. Saturating concentrations of glucose did not bring about complete dissociation of the enzyme showing that both sites were occupired in the dimer. At pH 8.0 and high ionic strength, where the enzyme existed as monomer, the dissociation constant of the enzyme-glucose complex was 3 X 10(-4) mol 1(-1) and was independent of the concentration of enzyme. Binding to the dimeric form at low pH and ionic strength (I=0.02 mol 1(-1), pH less than 7.5) was also independent of enzyme concentration (in the range 10-1000 mug ml-1) but was much weaker. The process could be described by a single dissociation constant, showing that the two available sites on the dimer were equivalent and non-cooperative; values of the intrinsic dissociation constant varied from 2.5 X 10(-3) mol 1(-1) at pH 7.0 to 6 X 10(-3) at pH 6.5. Under intermediate conditions (pH 7.0, ionic strength=0.15 mol 1(-1)), where monomer and dimer coexisted, the binding of glucose showed weak positive cooperatively (Hill coefficient 1.2); in addition, the binding was dependent upon the concentration of enzyme in the direction of stronger binding at lower concentrations. The results show that the phenomenon of half-sites reactivity observed in the binding of glucose to crystalline hexokinase P-II does not occur in solution; the simplest explanation of our finding the two sites to be equivalent is that the dimer results from the homologous association of two identical subunits.

Probing the electrostatics and pharmacologic 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 Central

Munde, Manoj; Poon, Gregory M. K.; Wilson, W. David

2013-01-01

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 (≤105 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 (>107 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. PMID:23416556

Targeting protein-protein interactions with trimeric ligands: high affinity inhibitors of the MAGUK protein family.

PubMed

Nissen, Klaus B; Haugaard-Kedström, Linda M; Wilbek, Theis S; Nielsen, Line S; Åberg, Emma; Kristensen, Anders S; Bach, Anders; Jemth, Per; Strømgaard, Kristian

2015-01-01

PDZ domains in general, and those of PSD-95 in particular, are emerging as promising drug targets for diseases such as ischemic stroke. We have previously shown that dimeric ligands that simultaneously target PDZ1 and PDZ2 of PSD-95 are highly potent inhibitors of PSD-95. However, PSD-95 and the related MAGUK proteins contain three consecutive PDZ domains, hence we envisioned that targeting all three PDZ domains simultaneously would lead to more potent and potentially more specific interactions with the MAGUK proteins. Here we describe the design, synthesis and characterization of a series of trimeric ligands targeting all three PDZ domains of PSD-95 and the related MAGUK proteins, PSD-93, SAP-97 and SAP-102. Using our dimeric ligands targeting the PDZ1-2 tandem as starting point, we designed novel trimeric ligands by introducing a PDZ3-binding peptide moiety via a cysteine-derivatized NPEG linker. The trimeric ligands generally displayed increased affinities compared to the dimeric ligands in fluorescence polarization binding experiments and optimized trimeric ligands showed low nanomolar inhibition towards the four MAGUK proteins, thus being the most potent inhibitors described. Kinetic experiments using stopped-flow spectrometry showed that the increase in affinity is caused by a decrease in the dissociation rate of the trimeric ligand as compared to the dimeric ligands, likely reflecting the lower probability of simultaneous dissociation of all three PDZ ligands. Thus, we have provided novel inhibitors of the MAGUK proteins with exceptionally high affinity, which can be used to further elucidate the therapeutic potential of these proteins.

Fluorogen-Activating-Proteins as Universal Affinity Biosensors for Immunodetection

PubMed Central

Gallo, Eugenio; Vasilev, Kalin V.; Jarvik, Jonathan

2014-01-01

Fluorogen-activating-proteins (FAPs) are a novel platform of fluorescence biosensors utilized for protein discovery. The technology currently demands molecular manipulation methods that limit its application and adaptability. Here, we highlight an alternative approach based on universal affinity reagents for protein detection. The affinity reagents were engineered as bi-partite fusion proteins, where the specificity moiety is derived from IgG-binding proteins –Protein-A or Protein-G – and the signaling element is a FAP. In this manner, primary antibodies provide the antigenic selectivity against a desired protein in biological samples, while FAP affinity reagents target the constant region (Fc) of antibodies and provide the biosensor component of detection. Fluorescence results using various techniques indicate minimal background and high target specificity for exogenous and endogenous proteins in mammalian cells. Additionally, FAP-based affinity reagents provide enhanced properties of detection previously absent using conventional affinity systems. Distinct features explored in this report include: (1) unfixed signal wavelengths (excitation and emission) determined by the particular fluorogen chosen, (2) real-time user controlled fluorescence on-set and off-set, (3) signal wavelength substitution while performing live analysis, and (4) enhanced resistance to photobleaching. PMID:24122476

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.

Substrate-Induced Facilitated Dissociation of the Competitive Inhibitor from the Active Site of O-Acetyl Serine Sulfhydrylase Reveals a Competitive-Allostery Mechanism.

PubMed

Singh, Appu Kumar; Ekka, Mary Krishna; Kaushik, Abhishek; Pandya, Vaibhav; Singh, Ravi P; Banerjee, Shrijita; Mittal, Monica; Singh, Vijay; Kumaran, S

2017-09-19

By classical competitive antagonism, a substrate and competitive inhibitor must bind mutually exclusively to the active site. The competitive inhibition of O-acetyl serine sulfhydrylase (OASS) by the C-terminus of serine acetyltransferase (SAT) presents a paradox, because the C-terminus of SAT binds to the active site of OASS with an affinity that is 4-6 log-fold (10 4 -10 6 ) greater than that of the substrate. Therefore, we employed multiple approaches to understand how the substrate gains access to the OASS active site under physiological conditions. Single-molecule and ensemble approaches showed that the active site-bound high-affinity competitive inhibitor is actively dissociated by the substrate, which is not consistent with classical views of competitive antagonism. We employed fast-flow kinetic approaches to demonstrate that substrate-mediated dissociation of full length SAT-OASS (cysteine regulatory complex) follows a noncanonical "facilitated dissociation" mechanism. To understand the mechanism by which the substrate induces inhibitor dissociation, we resolved the crystal structures of enzyme·inhibitor·substrate ternary complexes. Crystal structures reveal a competitive allosteric binding mechanism in which the substrate intrudes into the inhibitor-bound active site and disengages the inhibitor before occupying the site vacated by the inhibitor. In summary, here we reveal a new type of competitive allosteric binding mechanism by which one of the competitive antagonists facilitates the dissociation of the other. Together, our results indicate that "competitive allostery" is the general feature of noncanonical "facilitated/accelerated dissociation" mechanisms. Further understanding of the mechanistic framework of "competitive allosteric" mechanism may allow us to design a new family of "competitive allosteric drugs/small molecules" that will have improved selectivity and specificity as compared to their competitive and allosteric counterparts.

Weak acid-concentration Atot and dissociation constant Ka of plasma proteins in racehorses.

PubMed

Stampfli, H R; Misiaszek, S; Lumsden, J H; Carlson, G P; Heigenhauser, G J

1999-07-01

The plasma proteins are a significant contributor to the total weak acid concentration as a net anionic charge. Due to potential species difference, species-specific values must be confirmed for the weak acid anionic concentrations of proteins (Atot) and the effective dissociation constant for plasma weak acids (Ka). We studied the net anion load Atot of equine plasma protein in 10 clinically healthy mature Standardbred horses. A multi-step titration procedure, using a tonometer covering a titration range of PCO2 from 25 to 145 mmHg at 37 degrees C, was applied on the plasma of these 10 horses. Blood gases (pH, PCO2) and electrolytes required to calculate the strong ion difference ([SID] = [(Na(+) + K(+) + Ca(2+) + Mg(2+))-(Cl(-) + Lac(-) + PO4(2-))]) were simultaneously measured over a physiological pH range from 6.90-7.55. A nonlinear regression iteration to determine Atot and Ka was performed using polygonal regression curve fitting applied to the electrical neutrality equation of the physico-chemical system. The average anion-load Atot for plasma protein of 10 Standardbred horses was 14.89 +/- 0.8 mEq/l plasma and Ka was 2.11 +/- 0.50 x 10(-7) Eq/l (pKa = 6.67). The derived conversion factor (iterated Atot concentration/average plasma protein concentration) for calculation of Atot in plasma is 0.21 mEq/g protein (protein-unit: g/l). This value compares closely with the 0.24 mEq/g protein determined by titration of Van Slyke et al. (1928) and 0.22 mEq/g protein recently published by Constable (1997) for horse plasma. The Ka value compares closely with the value experimentally determined by Constable in 1997 (2.22 x 10(7) Eq/l). Linear regression of a set of experimental data from 5 Thoroughbred horses on a treadmill exercise test, showed excellent correlation with the regression lines not different from identity for the calculated and measured variables pH, HCO3 and SID. Knowledge of Atot and Ka for the horse is useful especially in exercise studies and in

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.

The stoichiometric dissociation constants of carbonic acid in seawater brines from 298 to 267 K

NASA Astrophysics Data System (ADS)

Papadimitriou, Stathys; Loucaides, Socratis; Rérolle, Victoire M. C.; Kennedy, Paul; Achterberg, Eric P.; Dickson, Andrew G.; Mowlem, Matthew; Kennedy, Hilary

2018-01-01

The stoichiometric dissociation constants of carbonic acid (K1C∗ and K2C∗) were determined by measurement of all four measurable parameters of the carbonate system (total alkalinity, total dissolved inorganic carbon, pH on the total proton scale, and CO2 fugacity) in natural seawater and seawater-derived brines, with a major ion composition equivalent to that of Reference Seawater, to practical salinity (SP) 100 and from 25 °C to the freezing point of these solutions and -6 °C temperature minimum. These values, reported in the total proton scale, provide the first such determinations at below-zero temperatures and for SP > 50. The temperature (T, in Kelvin) and SP dependence of the current pK1C∗ and pK2C∗ (as negative common logarithms) within the salinity and temperature ranges of this study (33 ≤ SP ≤ 100, -6 °C ≤ t ≤ 25 °C) is described by the following best-fit equations: pK1C∗ = -176.48 + 6.14528 SP0.5 - 0.127714 SP + 7.396 × 10-5SP2 + (9914.37 - 622.886 SP0.5 + 29.714 SP) T-1 + (26.05129 - 0.666812 SP0.5) lnT (σ = 0.011, n = 62), and pK2C∗ = -323.52692 + 27.557655 SP0.5 + 0.154922 SP - 2.48396 × 10-4 SP2 + (14763.287 - 1014.819 SP0.5 - 14.35223 SP) T-1 + (50.385807 - 4.4630415 SP0.5) lnT (σ = 0.020, n = 62). These functions are suitable for application to investigations of the carbonate system of internal sea ice brines with a conservative major ion composition relative to that of Reference Seawater and within the temperature and salinity ranges of this study.

Inter-residue coupling contributes to high-affinity subtype-selective binding of α-bungarotoxin to nicotinic receptors

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sine, Steven M.; Huang, Sun; Li, Shu-Xing

2013-09-01

The crystal structure of a pentameric α7 ligand-binding domain chimaera with bound α-btx (α-bungarotoxin) showed that of the five conserved aromatic residues in α7, only Tyr 184 in loop C of the ligand-binding site was required for high-affinity binding. To determine whether the contribution of Tyr 184 depends on local residues, we generated mutations in an α7/5HT 3A (5-hydroxytryptamine type 3A) receptor chimaera, individually and in pairs, and measured 125I-labelled α-btx binding. The results show that mutations of individual residues near Tyr 184 do not affect α-btx affinity, but pairwise mutations decrease affinity in an energetically coupled manner. Kinetic measurementsmore » show that the affinity decreases arise through increases in the α-btx dissociation rate with little change in the association rate. Replacing loop C in α7 with loop C from the α-btx-insensitive α2 or α3 subunits abolishes high-affinity α-btx binding, but preserves acetylcholine-elicited single channel currents. However, in both the α2 and α3 construct, mutating either residue that flanks Tyr 184 to its α7 counterpart restores high-affinity α-btx binding. Analogously, in α7, mutating both residues that flank Tyr 184 to the α2 or α3 counterparts abolishes high-affinity α-btx binding. Thus interaction between Tyr 184 and local residues contributes to high-affinity subtype-selective α-btx binding.« less

Dissociative recombination of N2/+/ in the ionosphere

NASA Technical Reports Server (NTRS)

Torr, M. R.; Torr, D. G.

1979-01-01

N2(+) ion measurements are examined which were made with the Atmospheric Explorer-C satellite during a phase of solar activity in 1978 that was significantly higher than near the earlier minimum. It is found that the major source of N2(+) is photoionization, rather than charge exchange with O(+) (2D), and that the major loss process above 300 km is dissociative recombination with electrons. A data sample for which the electron temperature (T sub e) covered the range from 1000 to 3400 K is used to evaluate the rate constant, alpha, for the dissociative-recombination process. The results show good agreement with laboratory measurements given by the expression: alpha = 1.8 x 10 to the -7th (T sub e/300) to the -0.39th cu cm/sec.

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…

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. Copyright © 2015 Elsevier Inc. All rights reserved.

The effect of 2,3-diphosphoglycerate on the oxygen dissociation curve of human haemoglobin.

PubMed Central

Goodford, P J; Norrington, F E; Paterson, R A; Wootton, R

1977-01-01

1. Oxygen dissociation curves for concentrated human haemoglobin solutions (1.6 mmol dm-3 in haem) have been measured by mixing known quantities of oxy- and deoxyhaemoglobin solutions and measuring the resulting partial pressure of oxygen with an oxygen electrode. 2. Observations in the presence of 2,3-diphosphoglycerate support previous conclusions derived from experiments at low haemoglobin concentrations, the validity of which has been questioned. 3. The two affinity state model of Monod, Wyman & Changeux (1965) does not fully describe the actions of 2,3-diphosphoglycerate and a model in which this allosteric effector not only binds preferentially to the T state but also lowers the oxygen affinity of this state gives an improved fit to the data. PMID:604451

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. Copyright © 2012 Elsevier Inc. All rights reserved.

Dissociation and serenity induction.

PubMed

Zoellner, Lori A; Sacks, Matthew B; Foa, Edna B

2007-09-01

Dissociation is a common experience during or immediately after a traumatic event; yet, most of the current knowledge regarding dissociation is retrospective in nature. The aim of the present study investigated a non-pharmacological method of dissociative induction with a clinical sample. Participants with PTSD and non-trauma exposed participants were randomly assigned to receive either a dissociative induction, or a serenity induction, based on modified Velten mood induction procedures. Participants receiving the dissociative induction reported higher state-dissociation than those receiving the serenity induction. The PTSD group reported greater state dissociation than the non-trauma exposed group, regardless of induction. State dissociation was related to trait dissociation, PTSD severity, and depression. The present results provide an initial demonstration of the viability for inducing state dissociation in the laboratory with a PTSD sample.

[Role of hemoglobin affinity to oxygen in adaptation to hypoxemia].

PubMed

Kwasiborski, Przemysław Jerzy; Kowalczyk, Paweł; Zieliński, Jakub; Przybylski, Jacek; Cwetsch, Andrzej

2010-04-01

One of the basic mechanisms of adapting to hypoxemia is a decrease in the affinity of hemoglobin for oxygen. This process occurs mainly due to the increased synthesis of 2,3-diphosphoglycerate (2,3-DPG) in the erythrocytes, as well as through the Bohr effect. Hemoglobin with decreased affinity for oxygen increases the oxygenation of tissues, because it gives up oxygen more easily during microcirculation. In foetal circulation, however, at a partial oxygen pressure (pO2) of 25 mmHg in the umbilical vein, the oxygen carrier is type F hemoglobin which has a high oxygen affinity. The commonly accepted role for hemoglobin F is limited to facilitating diffusion through the placenta. Is fetal life the only moment when haemoglobin F is useful? THE AIM OF STUDY was to create a mathematical model, which would answer the question at what conditions an increase, rather than a decrease, in haemoglobin oxygen affinity is of benefit to the body. Using the kinetics of dissociation of oxygen from hemoglobin described by the Hill equation as the basis for further discussion, we created a mathematical model describing the pO2 value in the microcirculatory system and its dependence on arterial blood pO2. The calculations were performed for hemoglobin with low oxygen affinity (adult type) and high-affinity hemoglobin (fetal type). The modelling took into account both physiological and pathological ranges of acid-base equilibrium and tissue oxygen extraction parameters. It was shown that for the physiological range of acid-base equilibrium and the resting level of tissue oxygen extraction parameters, with an arterial blood pO2 of 26.8 mmHg, the higher-affinity hemoglobin becomes the more effective oxygen carrier. It was also demonstrated that the arterial blood pO2, below which the high-affinity hemoglobin becomes the more effective carrier, is dependent on blood pH and the difference between the arterial and venous oxygen saturation levels. Simulations performed for the pathological

High Affinity Binding of Indium and Ruthenium Ions by Gastrins

PubMed Central

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. PMID:26457677

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

PubMed

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

2013-11-01

We previously demonstrated an important role of the constant region in the pathogenicity of anti-DNA antibodies. To determine the mechanisms by which the constant region affects autoantibody binding, a panel of isotype-switch variants (IgG1, IgG2a, IgG2b) was generated from the murine PL9-11 IgG3 autoantibody. The affinity of the PL9-11 antibody panel for histone was measured by surface plasmon resonance (SPR). Tryptophan fluorescence was used to determine wavelength shifts of the antibody panel upon binding to DNA and histone. Finally, circular dichroism spectroscopy was used to measure changes in secondary structure. SPR analysis revealed significant differences in histone binding affinity between members of the PL9-11 panel. The wavelength shifts of tryptophan fluorescence emission were found to be dependent on the antibody isotype, while circular dichroism analysis determined that changes in antibody secondary structure content differed between isotypes upon antigen binding. Thus, the antigen binding affinity is dependent on the particular constant region expressed. Moreover, the effects of antibody binding to antigen were also constant region dependent. Alteration of secondary structures influenced by constant regions may explain differences in fine specificity of anti-DNA antibodies between antibodies with similar variable regions, as well as cross-reactivity of anti-DNA antibodies with non-DNA antigens. Copyright © 2013 Elsevier Ltd. All rights reserved.

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.

Tracking scientific interest in the dissociative disorders: a study of scientific publication output 1984-2003.

PubMed

Pope, Harrison G; Barry, Steven; Bodkin, Alexander; Hudson, James I

2006-01-01

We attempted to track scientific interest in dissociative amnesia and dissociative identity disorder by estimating the annual output of publications regarding these entities over the last 20 years. Using a standard medical index, PsycINFO, we counted the number of indexed publications involving dissociative amnesia and dissociative identity disorder listed for each year. We then compared these rates with those of well-established diagnoses such as anorexia nervosa, alcohol abuse, and schizophrenia. We also systematically reviewed all publications involving dissociative amnesia that appeared in 2003. Annual publications involving dissociative amnesia and dissociative identity disorder rose from low levels in the 1980's to a sharp peak in the mid 1990's, followed by an equally sharp decline to only about one quarter of their peak levels by 2002 and 2003. In contrast, all of the 25 comparison diagnoses in our survey showed constant or steadily rising publication rates; none showed the 'bubble' pattern of the dissociative disorders. Of the 34 papers involving dissociative amnesia identified by PsycINFO for 2003, 10 (32%) appeared skeptical of the validity of dissociative amnesia and/or recovered-memory therapy. Despite a detailed search using multiple medical indices and search terms, we could find only 13 explicit cases of individuals with dissociative amnesia worldwide in the 2003 literature. Dissociative amnesia and dissociative identity disorder have not generated consistent scientific interest over the years, but instead apparently enjoyed a brief period of fashion that now has waned. Overall, our observations suggest that these diagnostic entities presently do not command widespread scientific acceptance. Copyright 2006 S. Karger AG, Basel.

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.

Rate Constants and Mechanisms of Protein–Ligand Binding

PubMed Central

Pang, Xiaodong; Zhou, Huan-Xiang

2017-01-01

Whereas protein–ligand binding affinities have long-established prominence, binding rate constants and binding mechanisms have gained increasing attention in recent years. Both new computational methods and new experimental techniques have been developed to characterize the latter properties. It is now realized that binding mechanisms, like binding rate constants, can and should be quantitatively determined. In this review, we summarize studies and synthesize ideas on several topics in the hope of providing a coherent picture of and physical insight into binding kinetics. The topics include microscopic formulation of the kinetic problem and its reduction to simple rate equations; computation of binding rate constants; quantitative determination of binding mechanisms; and elucidation of physical factors that control binding rate constants and mechanisms. PMID:28375732

Hydride affinity scale of various substituted arylcarbeniums in acetonitrile.

PubMed

Zhu, Xiao-Qing; Wang, Chun-Hua

2010-12-23

certainly serve as one of the most effective ways to estimate the new E values from ΔG(H(-)A)(R(+)) or vice versa. Combining the hydride affinities and the reduction potentials of the arylcarbeniums, we obtained the bond homolytic dissociation Gibbs free energy changes of the C-H bonds in the corresponding hydride adducts in acetonitrile, ΔG(HD)(RH), and found that the effects of the substituent on ΔG(HD)(RH) are very small. Simple thermodynamic analytic platforms for the three C-H cleavage modes were constructed. It is evident that the present work would be helpful in understanding the nature of the stabilities of the carbeniums and mechanisms of the hydride transfers between carbeniums and other hydride donors.

Simultaneous Glycan-Peptide Characterization Using Hydrophilic Interaction Chromatography and Parallel Fragmentation by CID, Higher Energy Collisional Dissociation, and Electron Transfer Dissociation MS Applied to the N-Linked Glycoproteome of Campylobacter jejuni*

PubMed Central

Scott, Nichollas E.; Parker, Benjamin L.; Connolly, Angela M.; Paulech, Jana; Edwards, Alistair V. G.; Crossett, Ben; Falconer, Linda; Kolarich, Daniel; Djordjevic, Steven P.; Højrup, Peter; Packer, Nicolle H.; Larsen, Martin R.; Cordwell, Stuart J.

2011-01-01

Campylobacter jejuni is a gastrointestinal pathogen that is able to modify membrane and periplasmic proteins by the N-linked addition of a 7-residue glycan at the strict attachment motif (D/E)XNX(S/T). Strategies for a comprehensive analysis of the targets of glycosylation, however, are hampered by the resistance of the glycan-peptide bond to enzymatic digestion or β-elimination and have previously concentrated on soluble glycoproteins compatible with lectin affinity and gel-based approaches. We developed strategies for enriching C. jejuni HB93-13 glycopeptides using zwitterionic hydrophilic interaction chromatography and examined novel fragmentation, including collision-induced dissociation (CID) and higher energy collisional (C-trap) dissociation (HCD) as well as CID/electron transfer dissociation (ETD) mass spectrometry. CID/HCD enabled the identification of glycan structure and peptide backbone, allowing glycopeptide identification, whereas CID/ETD enabled the elucidation of glycosylation sites by maintaining the glycan-peptide linkage. A total of 130 glycopeptides, representing 75 glycosylation sites, were identified from LC-MS/MS using zwitterionic hydrophilic interaction chromatography coupled to CID/HCD and CID/ETD. CID/HCD provided the majority of the identifications (73 sites) compared with ETD (26 sites). We also examined soluble glycoproteins by soybean agglutinin affinity and two-dimensional electrophoresis and identified a further six glycosylation sites. This study more than doubles the number of confirmed N-linked glycosylation sites in C. jejuni and is the first to utilize HCD fragmentation for glycopeptide identification with intact glycan. We also show that hydrophobic integral membrane proteins are significant targets of glycosylation in this organism. Our data demonstrate that peptide-centric approaches coupled to novel mass spectrometric fragmentation techniques may be suitable for application to eukaryotic glycoproteins for simultaneous

Simultaneous glycan-peptide characterization using hydrophilic interaction chromatography and parallel fragmentation by CID, higher energy collisional dissociation, and electron transfer dissociation MS applied to the N-linked glycoproteome of Campylobacter jejuni.

PubMed

Scott, Nichollas E; Parker, Benjamin L; Connolly, Angela M; Paulech, Jana; Edwards, Alistair V G; Crossett, Ben; Falconer, Linda; Kolarich, Daniel; Djordjevic, Steven P; Højrup, Peter; Packer, Nicolle H; Larsen, Martin R; Cordwell, Stuart J

2011-02-01

Campylobacter jejuni is a gastrointestinal pathogen that is able to modify membrane and periplasmic proteins by the N-linked addition of a 7-residue glycan at the strict attachment motif (D/E)XNX(S/T). Strategies for a comprehensive analysis of the targets of glycosylation, however, are hampered by the resistance of the glycan-peptide bond to enzymatic digestion or β-elimination and have previously concentrated on soluble glycoproteins compatible with lectin affinity and gel-based approaches. We developed strategies for enriching C. jejuni HB93-13 glycopeptides using zwitterionic hydrophilic interaction chromatography and examined novel fragmentation, including collision-induced dissociation (CID) and higher energy collisional (C-trap) dissociation (HCD) as well as CID/electron transfer dissociation (ETD) mass spectrometry. CID/HCD enabled the identification of glycan structure and peptide backbone, allowing glycopeptide identification, whereas CID/ETD enabled the elucidation of glycosylation sites by maintaining the glycan-peptide linkage. A total of 130 glycopeptides, representing 75 glycosylation sites, were identified from LC-MS/MS using zwitterionic hydrophilic interaction chromatography coupled to CID/HCD and CID/ETD. CID/HCD provided the majority of the identifications (73 sites) compared with ETD (26 sites). We also examined soluble glycoproteins by soybean agglutinin affinity and two-dimensional electrophoresis and identified a further six glycosylation sites. This study more than doubles the number of confirmed N-linked glycosylation sites in C. jejuni and is the first to utilize HCD fragmentation for glycopeptide identification with intact glycan. We also show that hydrophobic integral membrane proteins are significant targets of glycosylation in this organism. Our data demonstrate that peptide-centric approaches coupled to novel mass spectrometric fragmentation techniques may be suitable for application to eukaryotic glycoproteins for simultaneous

( sup 3 H)-DOB(4-bromo-2,5-dimethoxyphenylisopropylamine) and ( sup 3 H) ketanserin label two affinity states of the cloned human 5-hydroxytryptamine2 receptor

DOE Office of Scientific and Technical Information (OSTI.GOV)

Branchek, T.; Adham, N.; Macchi, M.

1990-11-01

The binding properties of the 5-hydroxytryptamine2 (5-HT2) receptor have been the subject of much interest and debate in recent years. The hallucinogenic amphetamine derivative 4-bromo-2,5-dimethoxyphenylisopropylamine (DOB) has been shown to bind to a small number of binding sites with properties very similar to (3H)ketanserin-labeled 5-HT2 receptors, but with much higher agonist affinities. Some researchers have interpreted this as evidence for the existence of a new subtype of 5-HT2 receptor (termed 5-HT2A), whereas others have interpreted these data as indicative of agonist high affinity and agonist low affinity states for the 5-HT2 receptor. In this investigation, a cDNA clone encoding themore » serotonin 5-HT2 receptor was transiently transfected into monkey kidney Cos-7 cells and stably transfected into mouse fibroblast L-M(TK-) cells. In both systems, expression of this single serotonin receptor cDNA led to the appearance of both (3H)DOB and (3H)ketanserin binding sites with properties that matched their binding characteristics in mammalian brain homogenates. Addition of guanosine 5'-(beta, gamma-imido) triphosphate (Gpp(NH)p) to this system caused a rightward shift and steepening of agonist competition curves for (3H) ketanserin binding, converting a two-site binding curve to a single low affinity binding state. Gpp(NH)p addition also caused a 50% decrease in the number of high affinity (3H)DOB binding sites, with no change in the dissociation constant of the remaining high affinity states. These data on a single human 5-HT2 receptor cDNA expressed in two different transfection host cells indicate that (3H)DOB and (3H)ketanserin binding reside on the same gene product, apparently interacting with agonist and antagonist conformations of a single human 5-HT2 receptor protein.« less

Dissociation in patients with dissociative seizures: relationships with trauma and seizure symptoms.

PubMed

Pick, S; Mellers, J D C; Goldstein, L H

2017-05-01

This study aimed to extend the current understanding of dissociative symptoms experienced by patients with dissociative (psychogenic, non-epileptic) seizures (DS), including psychological and somatoform types of symptomatology. An additional aim was to assess possible relationships between dissociation, traumatic experiences, post-traumatic symptoms and seizure manifestations in this group. A total of 40 patients with DS were compared with a healthy control group (n = 43), matched on relevant demographic characteristics. Participants completed several self-report questionnaires, including the Multiscale Dissociation Inventory (MDI), Somatoform Dissociation Questionnaire-20, Traumatic Experiences Checklist and the Post-Traumatic Diagnostic Scale. Measures of seizure symptoms and current emotional distress (Hospital Anxiety and Depression Scale) were also administered. The clinical group reported significantly more psychological and somatoform dissociative symptoms, trauma, perceived impact of trauma, and post-traumatic symptoms than controls. Some dissociative symptoms (i.e. MDI disengagement, MDI depersonalization, MDI derealization, MDI memory disturbance, and somatoform dissociation scores) were elevated even after controlling for emotional distress; MDI depersonalization scores correlated positively with trauma scores while seizure symptoms correlated with MDI depersonalization, derealization and identity dissociation scores. Exploratory analyses indicated that somatoform dissociation specifically mediated the relationship between reported sexual abuse and DS diagnosis, along with depressive symptoms. A range of psychological and somatoform dissociative symptoms, traumatic experiences and post-traumatic symptoms are elevated in patients with DS relative to healthy controls, and seem related to seizure manifestations. Further studies are needed to explore peri-ictal dissociative experiences in more detail.

Temperature Dependence of Inorganic Nitrogen Uptake: Reduced Affinity for Nitrate at Suboptimal Temperatures in Both Algae and Bacteria

PubMed Central

Reay, David S.; Nedwell, David B.; Priddle, Julian; Ellis-Evans, J. Cynan

1999-01-01

Nitrate utilization and ammonium utilization were studied by using three algal isolates, six bacterial isolates, and a range of temperatures in chemostat and batch cultures. We quantified affinities for both substrates by determining specific affinities (specific affinity = maximum growth rate/half-saturation constant) based on estimates of kinetic parameters obtained from chemostat experiments. At suboptimal temperatures, the residual concentrations of nitrate in batch cultures and the steady-state concentrations of nitrate in chemostat cultures both increased. The specific affinity for nitrate was strongly dependent on temperature (Q10 ≈ 3, where Q10 is the proportional change with a 10°C temperature increase) and consistently decreased at temperatures below the optimum temperature. In contrast, the steady-state concentrations of ammonium remained relatively constant over the same temperature range, and the specific affinity for ammonium exhibited no clear temperature dependence. This is the first time that a consistent effect of low temperature on affinity for nitrate has been identified for psychrophilic, mesophilic, and thermophilic bacteria and algae. The different responses of nitrate uptake and ammonium uptake to temperature imply that there is increasing dependence on ammonium as an inorganic nitrogen source at low temperatures. PMID:10347046

Iodination of (Tyr11)somatostatin yields a super high affinity ligand for somatostatin receptors in GH4C1 pituitary cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

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 reachmore » 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%).« less

Evidence that Ribulose 1,5-Bisphosphate (RuBP) Binds to Inactive Sites of RuBP Carboxylase in Vivo and an Estimate of the Rate Constant for Dissociation 1

PubMed Central

Cardon, Zoe G.; Mott, Keith A.

1989-01-01

The binding of ribulose 1,5-bisphosphate (RuBP) to inactive (noncarbamylated) sites of the enzyme RuBP carboxylase in vivo was investigated in Spinacia oleracea and Helianthus annuus. The concentrations of RuBP and inactive sites were determined in leaf tissue as a function of time after a change to darkness. RuBP concentrations fell rapidly after the change to darkness and were approximately equal to the concentration of inactive sites after 60 s. Variations in the concentration of inactive sites, which were induced by differences in the light intensity before the light-dark transition, correlated with the concentration of RuBP between 60 and 120 s after the change to darkness. These data are discussed as evidence that RuBP binds to inactive sites of RuBP carboxylase in vivo. After the concentration of RuBP fell below that of inactive sites (at times longer than 60 s of darkness), the decline in RuBP was logarithmic with time. This would be expected if the dissociation of RuBP from inactive sites controlled the decline in RuBP concentration. These data were used to estimate the rate constant for dissociation of RuBP from inactive sites in vivo. PMID:16666692

Activation of Peptide ions by blackbody radiation: factors that lead to dissociation kinetics in the rapid energy exchange limit.

PubMed

Price, W D; Williams, E R

1997-11-20

Unimolecular rate constants for blackbody infrared radiative dissociation (BIRD) were calculated for the model protonated peptide (AlaGly)(n) (n = 2-32) using a variety of dissociation parameters. Combinations of dissociation threshold energies ranging from 0.8 to 1.7 eV and transition entropies corresponding to Arrhenius preexponential factors ranging from very "tight" (A(infinity) = 10(9.9) s(-1)) to "loose" (A(infinity) = 10(16.8) s(-1)) were selected to represent dissociation parameters within the experimental temperature range (300-520 K) and kinetic window (k(uni) = 0.001-0.20 s(-1)) typically used in the BIRD experiment. Arrhenius parameters were determined from the temperature dependence of these values and compared to those in the rapid energy exchange (REX) limit. In this limit, the internal energy of a population of ions is given by a Boltzmann distribution, and kinetics are the same as those in the traditional high-pressure limit. For a dissociation process to be in this limit, the rate of photon exchange between an ion and the vacuum chamber walls must be significantly greater than the dissociation rate. Kinetics rapidly approach the REX limit either as the molecular size or threshold dissociation energy increases or as the transition-state entropy or experimental temperature decreases. Under typical experimental conditions, peptide ions larger than 1.6 kDa should be in the REX limit. Smaller ions may also be in the REX limit depending on the value of the threshold dissociation energy and transition-state entropy. Either modeling or information about the dissociation mechanism must be known in order to confirm REX limit kinetics for these smaller ions. Three principal factors that lead to the size dependence of REX limit kinetics are identified. With increasing molecular size, rates of radiative absorption and emission increase, internal energy distributions become relatively narrower, and the microcanonical dissociation rate constants increase more

Activation of Peptide Ions by Blackbody Radiation: Factors That Lead to Dissociation Kinetics in the Rapid Energy Exchange Limit

PubMed Central

Price, William D.

2005-01-01

Unimolecular rate constants for blackbody infrared radiative dissociation (BIRD) were calculated for the model protonated peptide (AlaGly)n (n = 2–32) using a variety of dissociation parameters. Combinations of dissociation threshold energies ranging from 0.8 to 1.7 eV and transition entropies corresponding to Arrhenius preexponential factors ranging from very “tight” (A∞ = 109.9 s−1) to “loose” (A∞ = 1016.8 s−1) were selected to represent dissociation parameters within the experimental temperature range (300–520 K) and kinetic window (kuni = 0.001–0.20 s−1) typically used in the BIRD experiment. Arrhenius parameters were determined from the temperature dependence of these values and compared to those in the rapid energy exchange (REX) limit. In this limit, the internal energy of a population of ions is given by a Boltzmann distribution, and kinetics are the same as those in the traditional high-pressure limit. For a dissociation process to be in this limit, the rate of photon exchange between an ion and the vacuum chamber walls must be significantly greater than the dissociation rate. Kinetics rapidly approach the REX limit either as the molecular size or threshold dissociation energy increases or as the transition-state entropy or experimental temperature decreases. Under typical experimental conditions, peptide ions larger than 1.6 kDa should be in the REX limit. Smaller ions may also be in the REX limit depending on the value of the threshold dissociation energy and transition-state entropy. Either modeling or information about the dissociation mechanism must be known in order to confirm REX limit kinetics for these smaller ions. Three principal factors that lead to the size dependence of REX limit kinetics are identified. With increasing molecular size, rates of radiative absorption and emission increase, internal energy distributions become relatively narrower, and the microcanonical dissociation rate constants increase more slowly

Dissociation Energetics and Mechanisms of Leucine Enkephalin (M + H)+ and (2M + X)+ Ions (X = H, Li, Na, K, and Rb) Measured by Blackbody Infrared Radiative Dissociation

PubMed Central

Schnier, Paul D.; Price, William D.; Strittmatter, Eric F.; Williams, Evan R.

2005-01-01

The dissociation kinetics of protonated leucine enkephalin and its proton and alkali metal bound dimers were investigated by blackbody infrared radiative dissociation in a Fourier-transform mass spectrometer. From the temperature dependence of the unimolecular dissociation rate constants, Arrhenius activation parameters in the zero-pressure limit are obtained. Protonated leucine enkephalin dissociates to form b4 and (M−H2O)+ ions with an average activation energy (Ea) of 1.1 eV and an A factor of 1010.5 s−1. The value of the A factor indicates that these dissociation processes are rearrangements. The b4 ions subsequently dissociate to form a4 ions via a process with a relatively high activation energy (1.3 eV), but one that is entropically favored. For the cationized dimers, the thermal stability decreases with increasing cation size, consistent with a simple electrostatic interaction in these noncovalent ion–molecule complexes. The Ea and A factors are indistinguishable within experimental error with values of ~1.5 eV and 1017 s−1, respectively. Although not conclusive, results from master equation modeling indicate that all these BIRD processes, except for b4 → a4, are in the rapid energy exchange limit. In this limit, the internal energy of the precursor ion population is given by a Boltzmann distribution and information about the energetics and dynamics of the reaction are obtained directly from the measured Arrhenius parameters. PMID:16554908

Dissociation energetics and mechanisms of leucine enkephalin (M + H)+ and (2M + X)+ ions (X = H, Li, Na, K, and Rb) measured by blackbody infrared radiative dissociation.

PubMed

Schnier, P D; Price, W D; Strittmatter, E F; Williams, E R

1997-08-01

The dissociation kinetics of protonated leucine enkephalin and its proton and alkali metal bound dimers were investigated by blackbody infrared radiative dissociation in a Fourier-transform mass spectrometer. From the temperature dependence of the unimolecular dissociation rate constants, Arrhenius activation parameters in the zero-pressure limit are obtained. Protonated leucine enkephalin dissociates to form b(4) and (M-H(2)O)(+) ions with an average activation energy (E(a)) of 1.1 eV and an A factor of 10(10.5) s(-1). The value of the A factor indicates that these dissociation processes are rearrangements. The b(4) ions subsequently dissociate to form a(4) ions via a process with a relatively high activation energy (1.3 eV), but one that is entropically favored. For the cationized dimers, the thermal stability decreases with increasing cation size, consistent with a simple electrostatic interaction in these noncovalent ion-molecule complexes. The E(a) and A factors are indistinguishable within experimental error with values of approximately 1.5 eV and 10(17) s(-1), respectively. Although not conclusive, results from master equation modeling indicate that all these BIRD processes, except for b(4) --> a(4), are in the rapid energy exchange limit. In this limit, the internal energy of the precursor ion population is given by a Boltzmann distribution and information about the energetics and dynamics of the reaction are obtained directly from the measured Arrhenius parameters.

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

Affinity ranking of antibodies using flow cytometry: application in antibody phage display-based target discovery.

PubMed

Geuijen, Cecilia A W; Clijsters-van der Horst, Marieke; Cox, Freek; Rood, Pauline M L; Throsby, Mark; Jongeneelen, Mandy A C; Backus, Harold H J; van Deventer, Els; Kruisbeek, Ada M; Goudsmit, Jaap; de Kruif, John

2005-07-01

Application of antibody phage display to the identification of cell surface antigens with restricted expression patterns is often complicated by the inability to demonstrate specific binding to a certain cell type. The specificity of an antibody can only be properly assessed when the antibody is of sufficient high affinity to detect low-density antigens on cell surfaces. Therefore, a robust and simple assay for the prediction of relative antibody affinities was developed and compared to data obtained using surface plasmon resonance (SPR) technology. A panel of eight anti-CD46 antibody fragments with different affinities was selected from phage display libraries and reformatted into complete human IgG1 molecules. SPR was used to determine K(D) values for these antibodies. The association and dissociation of the antibodies for binding to CD46 expressed on cell surfaces were analysed using FACS-based assays. We show that ranking of the antibodies based on FACS data correlates well with ranking based on K(D) values as measured by SPR and can therefore be used to discriminate between high- and low-affinity antibodies. Finally, we show that a low-affinity antibody may only detect high expression levels of a surface marker while failing to detect lower expression levels of this molecule, which may lead to a false interpretation of antibody specificity.

Adenosine transport systems on dissociated brain cells from mouse, guinea-pig, and rat

DOE Office of Scientific and Technical Information (OSTI.GOV)

Johnston, M.E.; Geiger, J.D.

1990-09-01

The kinetics and sodium dependence of adenosine transport were determined using an inhibitor-stop method on dissociated cell body preparations obtained from mouse, guinea-pig and rat brain. Transport affinity (KT) values for the high affinity adenosine transport systems KT(H) were significantly different between these three species; mean +/- SEM values were 0.34 +/- 0.1 in mouse, 0.9 +/- 0.2 in rat, and 1.5 +/- 0.5 microM in guinea-pig. The KT values for the low affinity transport system KT(L) were not different between the three species. Brain cells from rat displayed a significantly greater maximal capacity to accumulate (3H)adenosine (Vmax) than didmore » mouse or guinea-pig for the high affinity system, or than did mouse for the low affinity system. When sodium chloride was replaced in the transport medium with choline chloride, the KT(H) values for guinea-pig and rat were both increased by approximately 100%; only in rat did the change reach statistical significance. The sodium-dependence of adenosine transport in mouse brain was clearly absent. The differences between KT(H) values in mouse and those in guinea-pig or rat were accentuated in the absence of sodium. The differences in kinetic values, ionic requirements, and pharmacological characteristics between adenosine transporters in CNS tissues of mouse, guinea-pig and rat may help account for some of the variability noted among species in terms of their physiological responses to adenosine.« less

A possible molecular mechanism of the action of digitalis: ouabain action on calcium binding to sites associated with a purified sodium-potassium-activated adenosine triphosphatase from kidney.

PubMed

Gervais, A; Lane, L K; Anner, B M; Lindenmayer, G E; Schwartz, A

1977-01-01

Calcium binding at 0 degrees C to a purified sheep kidney Na+,K+-ATPase was described by linear Scatchard plots. Binding at saturating free calcium was 65-80 nmol/mg of protein, or 30-40 mol of calcium/mol of enzyme. Aqueous emulsions of lipids extracted from Na+,K+-ATPase yielded dissociation constants and maximum calcium-binding values that were similar to those for native Na+,K+-ATPase. Phospholipase A treatment markedly reduced calcium binding. Pretreatment of native Na+,K+-ATPase with ouabain increased the dissociation constant for calcium binding from 131 +/- 7 to 192 +/- 7 muM without altering maximum calcium binding. Ouabain pretreatment did not affect calcium binding to extracted phospholipids, ouabain-insensitive ATPases, or heat denatured Na+,K+-ATPase, Na+ and K+ (5-20 mM) increased the dissociation constants for calcium, which suggests competition between the monovalent cations and calcium for the binding sites. At higher concentrations of monovalent cations, ouabain increased the apparent affinity of binding sites for calcium. Extrapolation to physiological cation concentrations revealed that the ouabain-induced increase in apparent affinity for calcium may be as much as 2- to 3-fold. These results suggest: (1) calcium binds to phospholipids associated with Na+,K+-ATPase; (2) ouabain interaction with Na+,K+-ATPase induces a perturbation that is transmitted to adjacent phospholipids, altering their affinity for calcium; and (3) at physiological concentrations of Na+ or K+, or both, ouabain interaction with Na+,K+-ATPase may lead to an increased pool of membrane-bound calcium.

Binding Interactions of Dopamine and Apomorphine in D2High and D2Low States of Human Dopamine D2 Receptor Using Computational and Experimental Techniques.

PubMed

Durdagi, Serdar; Salmas, Ramin Ekhteiari; Stein, Matthias; Yurtsever, Mine; Seeman, Philip

2016-02-17

We have recently reported G-protein coupled receptor (GPCR) model structures for the active and inactive states of the human dopamine D2 receptor (D2R) using adrenergic crystal structures as templates. Since the therapeutic concentrations of dopamine agonists that suppress the release of prolactin are the same as those that act at the high-affinity state of the D2 receptor (D2High), D2High in the anterior pituitary gland is considered to be the functional state of the receptor. In addition, the therapeutic concentrations of anti-Parkinson drugs are also related to the dissociation constants in the D2High form of the receptor. The discrimination between the high- and low-affinity (D2Low) components of the D2R is not obvious and requires advanced computer-assisted structural biology investigations. Therefore, in this work, the derived D2High and D2Low receptor models (GPCR monomer and dimer three-dimensional structures) are used as drug-binding targets to investigate binding interactions of dopamine and apomorphine. The study reveals a match between the experimental dissociation constants of dopamine and apomorphine at their high- and low-affinity sites of the D2 receptor in monomer and dimer and their calculated dissociation constants. The allosteric receptor-receptor interaction for dopamine D2R dimer is associated with the accessibility of adjacent residues of transmembrane region 4. The measured negative cooperativity between agonist ligand at dopamine D2 receptor is also correctly predicted using the D2R homodimerization model.

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).

Peroxidative oxidation of halides catalysed by myeloperoxidase. Effect of fluoride on halide oxidation.

PubMed

Zgliczyński, J M; Stelmaszyńska, T; Olszowska, E; Krawczyk, A; Kwasnowska, E; Wróbel, J T

1983-01-01

It was found that all halides can compete with cyanide for binding with myeloperoxidase. The lower is the pH, the higher is the affinity of halides. The apparent dissociation constants (Kd) of myeloperoxidase-cyanide complex were determined in the presence of F-, Cl-, Br- and I- in the pH range of 4 to 7. In slightly acidic pH (4 - 6) fluoride and chloride exhibit a higher affinity towards the enzyme than bromide and iodide. Taking into account competition between cyanide and halides for binding with myeloperoxidase the dissociation constants of halide-myeloperoxidase complexes were calculated. All halides except fluoride can be oxidized by H2O2 in the presence of myeloperoxidase. However, since fluoride can bind with myeloperoxidase, it can competitively inhibit the oxidation of other halides. Fluoride was a competitive inhibitor with respect to other halides as well as to H2O2. Inhibition constants (Ki) for fluoride as a competitive inhibitor with respect to H2O2 increased from iodide oxidation through bromide to chloride oxidation.

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.

Affinity interactions between natural pigments and human whole saliva.

PubMed

Yao, Jiang-Wu; Lin, Feng; Tao, Tao; Lin, Chang-Jian

2011-03-01

The aim of the present study was to assess the null hypothesis that there are no differences of affinity between pigments and human whole saliva (WS), and the affinity is not influenced by the functional groups of pigments, temperatures, pH values, and salt concentrations. The affinity constants of interactions between WS and theaflavin (TF)/curcumin (Cur)/cyanidin (Cy) were determined by surface plasmon resonance (SPR) and fluorescence quenching. Mass-uptake at various temperatures, pH values, and salt concentrations was also carried out. The order of affinity of the pigments binding to WS is TF>Cur>Cy. A large number of complexes and precipitations of pigments/proteins were formed through a quick, strong, and almost irreversible binding process. The mass-uptake of pigments was affected not only by the functional groups, but also by molecular weight of pigments, temperatures, pH values, and salt concentrations. The complex of pigments may easily and rapidly deposit onto the WS film, and are difficult to remove from the WS surface. However, the complex of pigments can be reduced by properly regulating the physicochemical conditions, such as temperatures, pH values, and salt concentrations. Copyright © 2010 Elsevier Ltd. All rights reserved.

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

Determination of stability constants of aminoglycoside antibiotics with their metal complexes

NASA Astrophysics Data System (ADS)

Tiwow, Vanny M. A.

2014-03-01

One group of aminoglycoside antibiotics contains aminosugars. The aminosugar neomycin B with its derivate product neamine (2-Deoxy-4-0-(2,6-diamino-2,6-dideoxy-α-D-glucopyranosyl)-D-Streptamine) was identified as a free ligands and metal complexes. In particular, the stability constants of metal complexes by potentiometric titration techniques were investigated. Our previous study had determined the acid dissociation constants of these aminosugars with few metal complexes in fair depth. In this work, the complexation of two pyridine-containing amino alcohols and an amino sugar (neamine) have been measured potentiometrically. For instance, the stability constant of copper(II) complexation were determine and the model system generated an excellent fit. Stability constants with several metals have been determined and will be reported.

Influence of N-ethylmaleimide on cholinoceptors and responses in longitudinal muscles from guinea-pig ileum.

PubMed Central

Aronstam, R. S.; Carrier, G. O.

1982-01-01

1 The binding of carbamylcholine to membranes prepared from the longitudinal muscle of guinea-pig ileum was determined from its inhibition of the binding of [3H]-3-quinuclidinyl benzilate. Carbamylcholine binding was resolved into high and low affinity components with apparent dissociation constants of 0.11 +/- 0.02 and 11 +/- 1 microM; 42% of the receptors displayed high affinity carbamylcholine binding. 2 Alkylation of longitudinal muscle membranes with N-ethylmaleimide increased muscarinic receptor affinity for carbamylcholine in a manner consistent with a conversion of low affinity to high affinity receptors. After exposure the muscle membrane fragments to 1 mM N-ethylmaleimide for 20 min at 35 degrees C, carbamylcholine binding was resolved into two components with apparent dissociation constants of 0.11 +/- 0.01 and 9 +/- 2 microM, with 74% of the receptors displaying the higher affinity. 3 Exposure of longitudinal membranes mounted in an organ chamber to 1 mM N-ethylmaleimide for 30s depressed isometric contractions in response to acetylcholine by 80%, while contractions induced by K+ and Ba2+ were reduced by less than 20% and 10%, respectively. Acetylcholine dose-response curves were shifted to the right while Ba2+ curves were unaffected. 4 It is suggested that N-ethylmaleimide has a selective effect on muscarinic responses in the longitudinal muscle by disrupting processes occurring after receptor occupancy but before the induction of phospholipid turnover or calcium influx in the postsynaptic membrane. PMID:7126999

Direct dynamics simulations of the unimolecular dissociation of dioxetane: Probing the non-RRKM dynamics

NASA Astrophysics Data System (ADS)

Malpathak, Shreyas; Ma, Xinyou; Hase, William L.

2018-04-01

In a previous UB3LYP/6-31G* direct dynamics simulation, non-Rice-Ramsperger-Kassel-Marcus (RRKM) unimolecular dynamics was found for vibrationally excited 1,2-dioxetane (DO); [R. Sun et al., J. Chem. Phys. 137, 044305 (2012)]. In the work reported here, these dynamics are studied in more detail using the same direct dynamics method. Vibrational modes of DO were divided into 4 groups, based on their characteristic motions, and each group excited with the same energy. To compare with the dynamics of these groups, an additional group of trajectories comprising a microcanonical ensemble was also simulated. The results of these simulations are consistent with the previous study. The dissociation probability, N(t)/N(0), for these excitation groups were all different. Groups A, B, and C, without initial excitation in the O-O stretch reaction coordinate, had a time lag to of 0.25-1.0 ps for the first dissociation to occur. Somewhat surprisingly, the C-H stretch Group A and out-of-plane motion Group C excitations had exponential dissociation probabilities after to, with a rate constant ˜2 times smaller than the anharmonic RRKM value. Groups B and D, with excitation of the H-C-H bend and wag, and ring bend and stretch modes, respectively, had bi-exponential dissociation probabilities. For Group D, with excitation localized in the reaction coordinate, the initial rate constant is ˜7 times larger than the anharmonic RRKM value, substantial apparent non-RRKM dynamics. N(t)/N(0) for the random excitation trajectories was non-exponential, indicating intrinsic non-RRKM dynamics. For the trajectory integration time of 13.5 ps, 9% of these trajectories did not dissociate in comparison to the RRKM prediction of 0.3%. Classical power spectra for these trajectories indicate they have regular intramolecular dynamics. The N(t)/N(0) for the excitation groups are well described by a two-state coupled phase space model. From the intercept of N(t)/N(0) with random excitation, the

Increasing functional avidity of TCR-redirected T cells by removing defined N-glycosylation sites in the TCR constant domain

PubMed Central

Hauptrock, Beate; Malina, Victoria; Antunes, Edite; Voss, Ralf-Holger; Wolfl, Matthias; Strong, Roland; Theobald, Matthias; Greenberg, Philip D.

2009-01-01

Adoptive transfer of T lymphocytes transduced with a T cell receptor (TCR) to impart tumor reactivity has been reported as a potential strategy to redirect immune responses to target cancer cells (Schumacher, T.N. 2002. Nat. Rev. Immunol. 2:512–519). However, the affinity of most TCRs specific for shared tumor antigens that can be isolated is usually low. Thus, strategies to increase the affinity of TCRs or the functional avidity of TCR-transduced T cells might be therapeutically beneficial. Because glycosylation affects the flexibility, movement, and interactions of surface molecules, we tested if selectively removing conserved N-glycoslyation sites in the constant regions of TCR α or β chains could increase the functional avidity of T cells transduced with such modified TCRs. We observed enhanced functional avidity and improved recognition of tumor cells by T cells harboring TCR chains with reduced N-glycosylation (ΔTCR) as compared with T cells with wild-type (WT) TCR chains. T cells transduced with WT or ΔTCR chains bound tetramer equivalently at 4°C, but tetramer binding was enhanced at 37°C, predominantly as a result of reduced tetramer dissociation. This suggested a temperature-dependent mechanism such as TCR movement in the cell surface or structural changes of the TCR allowing improved multimerization. This strategy was effective with mouse and human TCRs specific for different antigens and, thus, should be readily translated to TCRs with any specificity. PMID:19171765

Photoaffinity labelling of the cardiac calcium channel. (-)-[3H]azidopine labels a 165 kDa polypeptide, and evidence against a [3H]-1,4-dihydropyridine-isothiocyanate being a calcium-channel-specific affinity ligand.

PubMed

Ferry, D R; Goll, A; Glossmann, H

1987-04-01

The arylazide 1,4-dihydropyridine (-)-[3H]azidopine binds to a saturable population of sites in guinea-pig heart membranes with a dissociation constant (KD) of 30 +/- 7 pM and a density (Bmax.) of 670 +/- 97 fmol/mg of protein. This high-affinity binding site is assumed to reside on voltage-operated calcium channels because reversible binding is blocked stereoselectively by 1,4-dihydropyridine channel blockers and by the enantiomers of Bay K 8644. A low-affinity (KD 25 +/- 7 nM) high-capacity (Bmax. 21.6 +/- 9 pmol/mg of protein) site does not bind (-)- or (+)-Bay K 8644, but is blocked by high concentrations (greater than 500 nM) of dihydro-2,6-dimethyl-4-(2-isothiocyanatophenyl)-3,5-pyridinedicarboxy lic acid dimethyl ester (1,4-DHP-isothiocyanate) or, e.g., (+/-)-nicardipine. (-)-[3H]Azidopine was photoincorporated covalently into bands of 165 +/- 8, 39 +/- 2 and 35 +/- 3 kDa, as determined by SDS/polyacrylamide-gel electrophoresis. Labelling of the 165 kDa band is protected stereoselectively by 1,4-dihydropyridine enantiomers at low (nM) concentrations and by (-)- and (+)-Bay K 8644, whereas the lower-Mr bands are not. Thus, only the 165 kDa band is the calcium-channel-linked 1,4-dihydropyridine receptor. Photolabelling of the 39 or 35 kDa bands was only blocked by 10 microM-1,4-DHP-isothiocyanate or 50 microM-(+/-)-nicardipine but not by 10 microM-(-)-Bay K 8644. [3H]-1,4-DHP-isothiocyanate binds to guinea-pig heart membranes with a KD of 0.35 nM and dissociates with a k-1 of 0.2 min-1 at 30 degrees C. [3H]-1,4 DHP-isothiocyanate irreversibly labels bands of 39 and 35 kDa which are protected by greater than 10 microM-(+/-)-nicardipine or unlabelled ligand but not by 10 microM-(-)-Bay K 8644. Thus, [3H]-1,4-DHP-isothiocyanate is not an affinity probe for the calcium channel.

Contemporary concepts of dissociation.

PubMed

Avdibegović, Esmina

2012-10-01

The concept of dissociation was developed in the late 19th century by Pierre Janet for conditions of "double consciousness" in hypnosis, hysteria, spirit possession and mediumship. He defined dissociation as a deficit in the capacity of integration of two or more different "systems of ideas and functions that constitute personality", and suggested that it can be related to a genetic component, to severe illness and fatigue, and particularly to experiencing adverse, potentially traumatizing events. By the late 20th century, various and often contradictory concepts of dissociation were suggested, which were either insufficient or exceedingly including when compared to the original idea. Currently, dissociation is used to describe a wide range of normal and abnormal phenomena as a process in which behaviour, thoughts and emotions can become separated one from another. A complete presentation of mechanisms involved in dissociation is still unknown. Scientific research on basic processes of dissociation is derived mainly from studies of hypnosis and post-traumatic stress disorder. Given the controversies in modern concepts of dissociation, some researchers and theorists suggest return to the original understanding of dissociation as a basic premise for the further development of the concept of dissociation.

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.

Dissociation of a Dynamic Protein Complex Studied by All-Atom Molecular Simulations.

PubMed

Zhang, Liqun; Borthakur, Susmita; Buck, Matthias

2016-02-23

The process of protein complex dissociation remains to be understood at the atomic level of detail. Computers now allow microsecond timescale molecular-dynamics simulations, which make the visualization of such processes possible. Here, we investigated the dissociation process of the EphA2-SHIP2 SAM-SAM domain heterodimer complex using unrestrained all-atom molecular-dynamics simulations. Previous studies on this system have shown that alternate configurations are sampled, that their interconversion can be fast, and that the complex is dynamic by nature. Starting from different NMR-derived structures, mutants were designed to stabilize a subset of configurations by swapping ion pairs across the protein-protein interface. We focused on two mutants, K956D/D1235K and R957D/D1223R, with attenuated binding affinity compared with the wild-type proteins. In contrast to calculations on the wild-type complexes, the majority of simulations of these mutants showed protein dissociation within 2.4 μs. During the separation process, we observed domain rotation and pivoting as well as a translation and simultaneous rolling, typically to alternate and weaker binding interfaces. Several unsuccessful recapturing attempts occurred once the domains were moderately separated. An analysis of protein solvation suggests that the dissociation process correlates with a progressive loss of protein-protein contacts. Furthermore, an evaluation of internal protein dynamics using quasi-harmonic and order parameter analyses indicates that changes in protein internal motions are expected to contribute significantly to the thermodynamics of protein dissociation. Considering protein association as the reverse of the separation process, the initial role of charged/polar interactions is emphasized, followed by changes in protein and solvent dynamics. The trajectories show that protein separation does not follow a single distinct pathway, but suggest that the mechanism of dissociation is common in

ab initio calculation of the rate of vibrational relaxation and thermal dissociation of hydrogen by helium at high temperatures

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dove, J.E.; Raynor, S.

The master equation for the thermal dissociation of para-H/sub 2/ infinitely dilute in He, was solved for temperatures of 1000 to 10,000/sup 0/K. Transition probabilities, used in the master equation, were obtained, in the case of energy transfer transitions, from distorted wave and quasi-classical trajectory calculations and, for dissociative processes, from trajectory calculations alone. An ab initio potential was used. From the solution, values of the dissociation rate constant, vibrational relaxation times, and incubation times for dissociation and vibrational relaxation were calculated. The sensitivity of the calculated results to variations in the transition probabilities was examined. Vibrational relaxation is mostmore » sensitive to simultaneous transitions in vibration and rotation (VRT processes); pure rotational (RT) transitions also have a substantial effect. Dissociation is most strongly affected by RT processes, but changes in VRT and groups of dissociative transitions also have a significant effect. However complete suppression of all dissociative transitions except those from levels immediately next to the continuum lowers the dissociation rates only by a factor of about 2. The location of the dissociation ''bottleneck'' is discussed. 5 figures, 3 tables.« less

The co-occurrence of PTSD and dissociation: differentiating severe PTSD from dissociative-PTSD.

PubMed

Armour, Cherie; Karstoft, Karen-Inge; Richardson, J Don

2014-08-01

A dissociative-posttraumatic stress disorder (PTSD) subtype has been included in the DSM-5. However, it is not yet clear whether certain socio-demographic characteristics or psychological/clinical constructs such as comorbid psychopathology differentiate between severe PTSD and dissociative-PTSD. The current study investigated the existence of a dissociative-PTSD subtype and explored whether a number of trauma and clinical covariates could differentiate between severe PTSD alone and dissociative-PTSD. The current study utilized a sample of 432 treatment seeking Canadian military veterans. Participants were assessed with the Clinician Administered PTSD Scale (CAPS) and self-report measures of traumatic life events, depression, and anxiety. CAPS severity scores were created reflecting the sum of the frequency and intensity items from each of the 17 PTSD and 3 dissociation items. The CAPS severity scores were used as indicators in a latent profile analysis (LPA) to investigate the existence of a dissociative-PTSD subtype. Subsequently, several covariates were added to the model to explore differences between severe PTSD alone and dissociative-PTSD. The LPA identified five classes: one of which constituted a severe PTSD group (30.5 %), and one of which constituted a dissociative-PTSD group (13.7 %). None of the included, demographic, trauma, or clinical covariates were significantly predictive of membership in the dissociative-PTSD group compared to the severe PTSD group. In conclusion, a significant proportion of individuals report high levels of dissociation alongside their PTSD, which constitutes a dissociative-PTSD subtype. Further investigation is needed to identify which factors may increase or decrease the likelihood of membership in a dissociative-PTSD subtype group compared to a severe PTSD only group.

A Force-Activated Trip Switch Triggers Rapid Dissociation of a Colicin from Its Immunity Protein

PubMed Central

Farrance, Oliver E.; Hann, Eleanore; Kaminska, Renata; Housden, Nicholas G.; Derrington, Sasha R.; Kleanthous, Colin; Radford, Sheena E.; Brockwell, David J.

2013-01-01

Colicins are protein antibiotics synthesised by Escherichia coli strains to target and kill related bacteria. To prevent host suicide, colicins are inactivated by binding to immunity proteins. Despite their high avidity (Kd≈fM, lifetime ≈4 days), immunity protein release is a pre-requisite of colicin intoxication, which occurs on a timescale of minutes. Here, by measuring the dynamic force spectrum of the dissociation of the DNase domain of colicin E9 (E9) and immunity protein 9 (Im9) complex using an atomic force microscope we show that application of low forces (<20 pN) increases the rate of complex dissociation 106-fold, to a timescale (lifetime ≈10 ms) compatible with intoxication. We term this catastrophic force-triggered increase in off-rate a trip bond. Using mutational analysis, we elucidate the mechanism of this switch in affinity. We show that the N-terminal region of E9, which has sparse contacts with the hydrophobic core, is linked to an allosteric activator region in E9 (residues 21–30) whose remodelling triggers immunity protein release. Diversion of the force transduction pathway by the introduction of appropriately positioned disulfide bridges yields a force resistant complex with a lifetime identical to that measured by ensemble techniques. A trip switch within E9 is ideal for its function as it allows bipartite complex affinity, whereby the stable colicin:immunity protein complex required for host protection can be readily converted to a kinetically unstable complex whose dissociation is necessary for cellular invasion and competitor death. More generally, the observation of two force phenotypes for the E9:Im9 complex demonstrates that force can re-sculpt the underlying energy landscape, providing new opportunities to modulate biological reactions in vivo; this rationalises the commonly observed discrepancy between off-rates measured by dynamic force spectroscopy and ensemble methods. PMID:23431269

The effects of pH and temperature on fluorescent calcium indicators as determined with Chelex-100 and EDTA buffer systems.

PubMed

Lattanzio, F A

1990-08-31

A novel method of determining the apparent dissociation constants of fluorescent calcium indicators is described which utilizes Chelex-100 ion exchange resin and 45Ca. The affinity for calcium of indicators fluo-3, fura-2 and indo-1 measured at either 22 degrees or 37 degrees C decreases as pH is decreased from 7.4 to 5.5. These measurements agree with determinations made using EDTA-calcium buffers. The 1:1 calcium:indicator complex is maintained under all conditions. The necessity to correct dissociation constants during intracellular acidification to properly interpret fluorescence measurements is illustrated by indo-1 measurements in the ischemic rat heart.

K*-charmonium dissociation cross sections and charmonium dissociation rates in hadronic matter

NASA Astrophysics Data System (ADS)

Liu, Feng-Rong; Ji, Shi-Tao; Xu, Xiao-Ming

2016-08-01

K*-charmonium dissociation reactions in hadronic matter are studied in the Born approximation, in the quark-interchange mechanism, and with a temperature-dependent quark potential. We obtain the temperature dependence of the unpolarized cross sections for the reactions K^* J/ψ to bar DD_s^ + ,bar D^* D_s^ + ,bar DD_s^{* + } , and bar D^* D_s^{* + } ; K^* χ _c to bar DD_s^ + ,bar D^* D_s^ + ,bar DD_s^{* + } , and bar D^* D_s^{* + } . We use the cross sections for charmonium dissociation in collisions with pions, ρ mesons, kaons, vector kaons, and η mesons to calculate the dissociation rates of charmonium with five types of mesons. Because of the temperature dependence of the meson masses, dissociation cross sections, and meson distribution functions, the charmonium dissociation rates generally increase with increasing temperature and decrease with increasing charmonium momentum from 2.2 GeV/c. We find that the first derivative of the dissociation rate with respect to the charmonium momentum is zero when the charmonium is at rest. While the η + ψ' and the η + χ c dissociation reactions can be neglected, the J/ ψ, ψ', and χ c dissociations are caused by collisions with pions, ρ mesons, kaons, vector kaons, and η mesons.

Dissociation in Psychiatric Disorders: A Meta-Analysis of Studies Using the Dissociative Experiences Scale.

PubMed

Lyssenko, Lisa; Schmahl, Christian; Bockhacker, Laura; Vonderlin, Ruben; Bohus, Martin; Kleindienst, Nikolaus

2018-01-01

Dissociation is a complex, ubiquitous construct in psychopathology. Symptoms of dissociation are present in a variety of mental disorders and have been connected to higher burden of illness and poorer treatment response, and not only in disorders with high levels of dissociation. This meta-analysis offers a systematic and evidence-based study of the prevalence and distribution of dissociation, as assessed by the Dissociative Experiences Scale, within different categories of mental disorders, and it updates an earlier meta-analysis. More than 1,900 original publications were screened, and 216 were included in the meta-analysis, comprising 15,219 individuals in 19 diagnostic categories. The largest mean dissociation scores were found in dissociative disorders (mean scores >35), followed by posttraumatic stress disorder, borderline personality disorder, and conversion disorder (mean scores >25). Somatic symptom disorder, substance-related and addictive disorders, feeding and eating disorders, schizophrenia, anxiety disorder, OCD, and most affective disorders also showed mean dissociation scores >15. Bipolar disorders yielded the lowest dissociation scores (mean score, 14.8). The findings underline the importance of careful psychopathological assessment of dissociative symptoms in the entire range of mental disorders.

Water channel in the binding site of a high affinity anti-methotrexate antibody.

PubMed

Gayda, Susan; Longenecker, Kenton L; Manoj, Sharmila; Judge, Russell A; Saldana, Sylvia C; Ruan, Qiaoqiao; Swift, Kerry M; Tetin, Sergey Y

2014-06-17

In the present study, we report the structure of the free and drug-bound Fab fragment of a high affinity anti-methotrexate antibody and perform a thermodynamic analysis of the binding process. The anti-methotrexate Fab fragment features a remarkably rigid tunnel-like binding site that extends into a water channel serving as a specialized route to move solvent out and into the site upon ligand binding and dissociation. This new finding in antibody structure-function relationships directly relates to the fast association (1 × 10⁷ M⁻¹ s⁻¹) and slow dissociation (4 × 10⁻⁵ s⁻¹) rates determined for mAb ADD056, resulting in a very strong binding with a K(D) ~ 3.6 pM at 20 °C. As follows from the X-ray data analysis, the methotrexate-antibody complex is stabilized by an extended network of hydrogen bonds and stacking interactions. The analysis also shows structural involvement of the CDR H3 in formation of the water channel revealing another important role of this hypervariable region. This suggests a new direction in natural affinity maturation and opens a new possibility in antibody engineering. Methotrexate is a widely used therapeutic agent for many malignant diseases and inflammatory disorders. Unfortunately, it may also interfere with central aspects of metabolism and thereby cause inevitable side effects. Therefore, methotrexate therapy requires careful monitoring of drug blood levels, which is traditionally done by immunoassays. An understanding of the structure-function properties of antibodies selected for drug monitoring substantiates the performance and robustness of such tests.

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.

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.

Prediction of Rate Constant for Supramolecular Systems with Multiconfigurations.

PubMed

Guo, Tao; Li, Haiyan; Wu, Li; Guo, Zhen; Yin, Xianzhen; Wang, Caifen; Sun, Lixin; Shao, Qun; Gu, Jingkai; York, Peter; Zhang, Jiwen

2016-02-25

The control of supramolecular systems requires a thorough understanding of their dynamics, especially on a molecular level. It is extremely difficult to determine the thermokinetic parameters of supramolecular systems, such as drug-cyclodextrin complexes with fast association/dissociation processes by experimental techniques. In this paper, molecular modeling combined with novel mathematical relationships integrating the thermodynamic/thermokinetic parameters of a series of isomeric multiconfigurations to predict the overall parameters in a range of pH values have been employed to study supramolecular dynamics at the molecular level. A suitable form of Eyring's equation was derived and a two-stage model was introduced. The new approach enabled accurate prediction of the apparent dissociation/association (k(off)/k(on)) and unbinding/binding (k-r/kr) rate constants of the ubiquitous multiconfiguration complexes of the supramolecular system. The pyronine Y (PY) was used as a model system for the validation of the presented method. Interestingly, the predicted k(off) value ((40 ± 1) × 10(5) s(-1), 298 K) of PY is largely in agreement with that previously determined by fluorescence correlation spectroscopy ((5 ± 3) × 10(5) s(-1), 298 K). Moreover, the k(off)/k(on) and k-r/kr for flurbiprofen-β-cylcodextrin and ibuprofen-β-cyclodextrin systems were also predicted and suggested that the association processes are diffusion-controlled. The methodology is considered to be especially useful in the design and selection of excipients for a supramolecular system with preferred association and dissociation rate constants and understanding their mechanisms. It is believed that this new approach could be applicable to a wide range of ligand-receptor supramolecular systems and will surely help in understanding their complex mechanism.

Effect of pressure on the dissociation constant of boric acid in water and seawater

NASA Astrophysics Data System (ADS)

Millero, Frank J.; Ward, Gary K.; Surdo, Antonio Lo; Huang, Fen

2012-01-01

The sound speeds of boric acid and sodium borate in water and 0.725 m NaCl have been measured from 0 to 50 °C and to near 1 molal. These results have been used to determine the partial molal adiabatic compressibilities of B(OH) 3 and NaB(OH) 4. The partial molal volumes, v¯(i), and compressibilities, κ¯(i), have been used to estimate the changes in the volume (Δ V) and compressibility (Δ κ) for the dissociation of boric acid in water and average seawater (0.725 m NaCl, SA ˜ 35 g/kg) B(OH)3+H2O=H++B(OH)4- where

Multiple exciton dissociation in CdSe quantum dots by ultrafast electron transfer to adsorbed methylene blue.

PubMed

Huang, Jier; Huang, Zhuangqun; Yang, Ye; Zhu, Haiming; Lian, Tianquan

2010-04-07

Multiexciton generation in quantum dots (QDs) may provide a new approach for improving the solar-to-electric power conversion efficiency in QD-based solar cells. However, it remains unclear how to extract these excitons before the ultrafast exciton-exciton annihilation process. In this study we investigate multiexciton dissociation dynamics in CdSe QDs adsorbed with methylene blue (MB(+)) molecules by transient absorption spectroscopy. We show that excitons in QDs dissociate by ultrafast electron transfer to MB(+) with an average time constant of approximately 2 ps. The charge separated state is long-lived (>1 ns), and the charge recombination rate increases with the number of dissociated excitons. Up to three MB(+) molecules per QD can be reduced by exciton dissociation. Our result demonstrates that ultrafast interfacial charge separation can effectively compete with exciton-exciton annihilation, providing a viable approach for utilizing short-lived multiple excitons in QDs.

Measuring Norfloxacin Binding to Trypsin Using a Fluorescence Quenching Assay in an Upper-Division, Integrated Laboratory Course

ERIC Educational Resources Information Center

Hicks, Katherine A.

2016-01-01

Fluorescence quenching assays are often used to measure dissociation constants that quantify the binding affinity between small molecules and proteins. In an upper-division undergraduate laboratory course, where students work on projects using a guided inquiry-based approach, a binding titration experiment at physiological pH is performed to…

Affinity of C-Reactive Protein toward FcγRI Is Strongly Enhanced by the γ-Chain

PubMed Central

Röcker, Carlheinz; Manolov, Dimitar E.; Kuzmenkina, Elza V.; Tron, Kyrylo; Slatosch, Holger; Torzewski, Jan; Nienhaus, G. Ulrich

2007-01-01

C-reactive protein (CRP), the prototype human acute phase protein, is widely regarded as a key player in cardiovascular disease, but the identity of its cellular receptor is still under debate. By using ultrasensitive confocal imaging analysis, we have studied CRP binding to transfected COS-7 cells expressing the high-affinity IgG receptor FcγRI. Here we show that CRP binds to FcγRI on intact cells, with a kd of 10 ± 3 μmol/L. Transfection of COS-7 cells with a plasmid coding for both FcγRI and its functional counterpart, the γ-chain, markedly increases CRP affinity to FcγRI, resulting in a kd of 0.35 ± 0.10 μmol/L. The affinity increase results from an ∼30-fold enhanced association rate coefficient. The pronounced enhancement of affinity by the γ-chain suggests its crucial involvement in the CRP receptor interaction, possibly by mediating interactions between the transmembrane moieties of the receptors. Dissociation of CRP from the cell surfaces cannot be detected throughout the time course of several hours and is thus extremely slow. Considering the pentameric structure of CRP, this result indicates that multivalent binding and receptor clustering are crucially involved in the interaction of CRP with nucleated cells. PMID:17255341

Role of Reversible Histidine Coordination in Hydroxylamine Reduction by Plant Hemoglobins (Phytoglobins).

PubMed

Athwal, Navjot Singh; Alagurajan, Jagannathan; Andreotti, Amy H; Hargrove, Mark S

2016-10-18

Reduction of hydroxylamine to ammonium by phytoglobin, a plant hexacoordinate hemoglobin, is much faster than that of other hexacoordinate hemoglobins or pentacoordinate hemoglobins such as myoglobin, leghemoglobin, and red blood cell hemoglobin. The reason for differences in reactivity is not known but could be intermolecular electron transfer between protein molecules in support of the required two-electron reduction, hydroxylamine binding, or active site architecture favoring the reaction. Experiments were conducted with phytoglobins from rice, tomato, and soybean along with human neuroglobin and soybean leghemoglobin that reveal hydroxylamine binding as the rate-limiting step. For hexacoordinate hemoglobins, binding is limited by the dissociation rate constant for the distal histidine, while leghemoglobin is limited by an intrinsically low affinity for hydroxylamine. When the distal histidine is removed from rice phytoglobin, a hydroxylamine-bound intermediate is formed and the reaction rate is diminished, indicating that the distal histidine imidazole side chain is critical for the reaction, albeit not for electron transfer but rather for direct interaction with the substrate. Together, these results demonstrate that phytoglobins are superior at hydroxylamine reduction because they have distal histidine coordination affinity constants near 1, and facile rate constants for binding and dissociation of the histidine side chain. Hexacoordinate hemoglobins such as neuroglobin are limited by tighter histidine coordination that blocks hydroxylamine binding, and pentacoordinate hemoglobins have intrinsically lower hydroxylamine affinities.

Dissociative disorders in DSM-5.

PubMed

Spiegel, David; Loewenstein, Richard J; Lewis-Fernández, Roberto; Sar, Vedat; Simeon, Daphne; Vermetten, Eric; Cardeña, Etzel; Brown, Richard J; Dell, Paul F

2011-12-21

We present recommendations for revision of the diagnostic criteria for the Dissociative Disorders (DDs) for DSM-5. The periodic revision of the DSM provides an opportunity to revisit the assumptions underlying specific diagnoses and the empirical support, or lack of it, for the defining diagnostic criteria. This paper reviews clinical, phenomenological, epidemiological, cultural, and neurobiological data related to the DDs in order to generate an up-to-date, evidence-based set of DD diagnoses and diagnostic criteria for DSM-5. First, we review the definitions of dissociation and the differences between the definitions of dissociation and conceptualization of DDs in the DSM-IV-TR and the ICD-10, respectively. Also, we review more general conceptual issues in defining dissociation and dissociative disorders. Based on this review, we propose a revised definition of dissociation for DSM-5 and discuss the implications of this definition for understanding dissociative symptoms and disorders. We make the following recommendations for DSM-5: 1. Depersonalization Disorder (DPD) should include derealization symptoms as well. 2. Dissociative Fugue should become a subtype of Dissociative Amnesia (DA). 3. The diagnostic criteria for DID should be changed to emphasize the disruptive nature of the dissociation and amnesia for everyday as well as traumatic events. The experience of possession should be included in the definition of identity disruption. 4. Dissociative Trance Disorder should be included in the Unspecified Dissociative Disorder (UDD) category. There is a growing body of evidence linking the dissociative disorders to a trauma history, and to specific neural mechanisms. © 2011 Wiley Periodicals, Inc.

Dissociative disorders in DSM-5.

PubMed

Spiegel, David; Loewenstein, Richard J; Lewis-Fernández, Roberto; Sar, Vedat; Simeon, Daphne; Vermetten, Eric; Cardeña, Etzel; Dell, Paul F

2011-09-01

We present recommendations for revision of the diagnostic criteria for the Dissociative Disorders (DDs) for DSM-5. The periodic revision of the DSM provides an opportunity to revisit the assumptions underlying specific diagnoses and the empirical support, or lack of it, for the defining diagnostic criteria. This paper reviews clinical, phenomenological, epidemiological, cultural, and neurobiological data related to the DDs in order to generate an up-to-date, evidence-based set of DD diagnoses and diagnostic criteria for DSM-5. First, we review the definitions of dissociation and the differences between the definitions of dissociation and conceptualization of DDs in the DSM-IV-TR and the ICD-10, respectively. Also, we review more general conceptual issues in defining dissociation and dissociative disorders. Based on this review, we propose a revised definition of dissociation for DSM-5 and discuss the implications of this definition for understanding dissociative symptoms and disorders. We make the following recommendations for DSM-5: 1. Depersonalization Disorder (DPD) should derealization symptoms as well. 2. Dissociative Fugue should become a subtype of Dissociative Amnesia (DA). 3. The diagnostic criteria for DID should be changed to emphasize the disruptive nature of the dissociation and amnesia for everyday as well as traumatic events. The experience of possession should be included in the definition of identity disruption. 4. Should Dissociative Trance Disorder should be included in the Unspecified Dissociative Disorder (UDD) category. There is a growing body of evidence linking the dissociative disorders to a trauma history, and to specific neural mechanisms. © 2011 Wiley-Liss, Inc.

Evaluation of affinity and pseudo-affinity adsorption processes for penicillin acylase purification.

PubMed

Fonseca, L P; Cabral, J M

1996-01-01

Affinity ligand (6-Aminopenicillanic acid, Amoxycillin, Ampicillin, Benzylpenicillin and 4-Phenylbutylanzine) of penicillin acylase (EC 3.5.1.11) were attached to hydrophilic gels like Sepharose 4B-CNBr and Minileak 'medium'. Ampicillin and 4-Phenylbutylamine were the affinity ligands that presented the higher concentrations attached to both gels. Penicillin acylase adsorption on these affinity gels was mainly dependent on the activated group of the gel, the affinity ligand attached and the experimental conditions of enzyme adsorption. Under affinity conditions only the ligands Amoxycillin, Ampicillin and 4-Phenylbutylamine, immobilized on Minileak, adsorbed the enzyme from osmotic shock extracts at different pH values. These affinity ligand systems were characterized by low adsorption capacities of penicillin acylase activity (1.2-2.1 IU mL-1 gel) and specific activity (1.5-2.9 IU mg-1 prot). Under pseudo-affinity conditions all the ligands attached both activated to gels (Sepharose 4B-CNBr and Minileak) adsorbed the enzyme. The affinity gels were characterized by higher values of adsorption capacity (3.7 and 55.6 IU mL-1 gel) and adsorbed specific activity (2.0 and 6.1 IU mg-1 prot) than those observed under affinity conditions. The space arm of Minileak gel, shown to be fundamental to enzyme adsorption under affinity conditions, preferentially adsorbed proteins in relation to the enzyme under pseudo-affinity conditions. However, this effect was partially minimized when the gel was derivatized by the affinity ligands at concentrations higher than 6 mumol mL-1 gel. Ampicillin was the affinity ligand that presented the best results for specific adsorption of penicillin acylase under affinity and pseudo-affinity adsorption processes. The Sepharose 4B-CNBr derivatized gel also presented a good adsorption capacity of enzyme activity (26.8 IU mL-1 gel) under pseudo-affinity adsorption processes.

Predictors of trait dissociation and peritraumatic dissociation induced via cold pressor.

PubMed

Gómez-Pérez, Lydia; López-Martínez, Alicia Eva; Asmundson, Gordon John Glenn

2013-11-30

Understanding which factors predict individual dissociative response during stressful situations is important to clarify the nature of dissociation and the mechanisms associated to its use as a coping strategy. The present study examined (1) whether experiential avoidance (EA), anxiety sensitivity (AS), depressive symptoms, and state anxiety concurrently predicted trait dissociation (TD)-absorption, amnesia, depersonalization, and total TD scores-and laboratory induced dissociation (LID); and (2) whether TD and catastrophizing predicted LID. We also examined whether catastrophizing mediated the relationships between both AS and depressive symptoms and LID. A total of 101 female undergraduate students participated in a cold pressor task, which significantly induced dissociation. Results of hierarchical regression analyses showed that AS at Time 1 (9 months before the experimental session), as well as depressive symptoms and catastrophizing at the time of the experiment (Time 2), predicted LID at Time 2. Depressive symptoms at Time 2 predicted total TD, absorption, and amnesia scores. AS at Time 1 and depressive symptoms at Time 2 predicted depersonalization. AS, depressive symptoms, and catastrophizing seem to facilitate the use of dissociative strategies by healthy individuals, even in response to non-traumatic but discomforting stress. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

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.

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.

Peptide nucleic acid probe for protein affinity purification based on biotin-streptavidin interaction and peptide nucleic acid strand hybridization.

PubMed

Tse, Jenny; Wang, Yuanyuan; Zengeya, Thomas; Rozners, Eriks; Tan-Wilson, Anna

2015-02-01

We describe a new method for protein affinity purification that capitalizes on the high affinity of streptavidin for biotin but does not require dissociation of the biotin-streptavidin complex for protein retrieval. Conventional reagents place both the selectively reacting group (the "warhead") and the biotin on the same molecule. We place the warhead and the biotin on separate molecules, each linked to a short strand of peptide nucleic acid (PNA), synthetic polymers that use the same bases as DNA but attached to a backbone that is resistant to attack by proteases and nucleases. As in DNA, PNA strands with complementary base sequences hybridize. In conditions that favor PNA duplex formation, the warhead strand (carrying the tagged protein) and the biotin strand form a complex that is held onto immobilized streptavidin. As in DNA, the PNA duplex dissociates at moderately elevated temperature; therefore, retrieval of the tagged protein is accomplished by a brief exposure to heat. Using iodoacetate as the warhead, 8-base PNA strands, biotin, and streptavidin-coated magnetic beads, we demonstrate retrieval of the cysteine protease papain. We were also able to use our iodoacetyl-PNA:PNA-biotin probe for retrieval and identification of a thiol reductase and a glutathione transferase from soybean seedling cotyledons. Copyright © 2014 Elsevier Inc. All rights reserved.

Point mutation increases a form of the NK1 receptor with high affinity for neurokinin A and B and septide

PubMed Central

Ciucci, Alessandra; Palma, Carla; Manzini, Stefano; Werge, Thomas M

1998-01-01

The binding modalities of substance P and neurokinin A on the wild type and Gly166 to-Cys mutant NK1 receptors expressed on CHO cells were investigated in homologous and heterologous binding experiments using both radiolabelled substance P and neurokinin A.On the wild type NK1 receptor NKA displaces radiolabelled substance P with very low apparent affinity, despite its high-affinity binding constant (determined in homologous binding experiments). The Gly166 to-Cys substitution in the NK1 tachykinin receptor greatly enhances the apparent affinity of neurokinin A in competition for radiolabelled substance P, but it does not change the binding constant of neurokinin A. The mutation, thereby, eliminates the discrepancy between the low apparent affinity and the high binding constant of neurokinin A.On the wild type receptor the binding capacity of neurokinin A is significantly smaller than that of substance P. In contrast, the two tachykinins bind to approximately the same number of sites on the mutant receptor.Simultaneous mass action law analysis of binding data in which multiple radioligands were employed in parallel demonstrated that a one-site model was unable to accommodate all the experimental data, whereas a two-site model provided a dramatically better description.These two receptor-sites display equally high affinity for substance P, while neurokinin A strongly discriminates between a high and a low affinity component. The binding affinities of neurokinin A are not affected by the mutation, which instead specifically alters the distribution between receptor sites in favour of a high affinity neurokinin A binding form.The low apparent affinity and binding capacity of neurokinin A on the wild type receptor results from neurokinin A binding with high affinity only to a fraction of the sites labelled by substance P. The mutation increases the proportion of this site, and consequently enhances the apparent affinity and binding capacity of neurokinin A.The binding

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.

Determination of metal-hydrogen bond dissociation energies by the deprotonation of transition metal hydride ions: application to MnH +

NASA Astrophysics Data System (ADS)

Stevens, Amy E.; Beauchamp, J. L.

1981-03-01

ICR trapped ion techniques are used to examine the kinetics of proton transfer from MnH + (formed as a fragment ion from HMn (CO) 5 by electron impact) to bases of varying strength. Deprotonation is rapid with bases whose proton affinity exceeds 196±3 kcal mol -1. This value for PA (Mn) yields the homolytic bond dissociation energy D0(Mn +-H) = 53±5 kcal mol -1.

Posttranslational modifications of Rab proteins cause effective displacement of GDP dissociation inhibitor.

PubMed

Oesterlin, Lena K; Goody, Roger S; Itzen, Aymelt

2012-04-10

Intracellular vesicular trafficking is regulated by approximately 60 members of the Rab subfamily of small Ras-like GDP/GTP binding proteins. Rab proteins cycle between inactive and active states as well as between cytosolic and membrane bound forms. Membrane extraction/delivery and cytosolic distribution of Rabs is mediated by interaction with the protein GDP dissociation inhibitor (GDI) that binds to prenylated inactive (GDP-bound) Rab proteins. Because the Rab:GDP:GDI complex is of high affinity, the question arises of how GDI can be displaced efficiently from Rab protein in order to allow the necessary recruitment of the Rab to its specific target membrane. While there is strong evidence that DrrA, as a bacterially encoded GDP/GTP exchange factor, contributes to this event, we show here that posttranslational modifications of Rabs can also modulate the affinity for GDI and thus cause effective displacement of GDI from Rab:GDI complexes. These activities have been found associated with the phosphocholination and adenylylation activities of the enzymes AnkX and DrrA/SidM, respectively, from the pathogenic bacterium Legionella pneumophila. Both modifications occur after spontaneous dissociation of Rab:GDI complexes within their natural equilibrium. Therefore, the effective GDI displacement that is observed is caused by inhibition of reformation of Rab:GDI complexes. Interestingly, in contrast to adenylylation by DrrA, AnkX can covalently modify inactive Rabs with high catalytic efficiency even when GDP is bound to the GTPase and hence can inhibit binding of GDI to Rab:GDP complexes. We therefore speculate that human cells could employ similar mechanisms in the absence of infection to effectively displace Rabs from GDI.

Assessment of complex dissociative disorder patients and simulated dissociation in forensic contexts.

PubMed

Brand, Bethany L; Webermann, Aliya R; Frankel, A Steven

Few assessors receive training in assessing dissociation and complex dissociative disorders (DDs). Potential differential diagnoses include anxiety, mood, psychotic, substance use, and personality disorders, as well as exaggeration and malingering. Individuals with DDs typically elevate on many clinical and validity scales on psychological tests, yet research indicates that they can be distinguished from DD simulators. Becoming informed about the testing profiles of DD individuals and DD simulators can improve the accuracy of differential diagnoses in forensic settings. In this paper, we first review the testing profiles of individuals with complex DDs and contrast them with DD simulators on assessment measures used in forensic contexts, including the Minnesota Multiphasic Personality Inventory-2 (MMPI-2), Personality Assessment Inventory (PAI), and the Structured Inventory of Reported Symptoms (SIRS), as well as dissociation-specific measures such as the Dissociative Experiences Scale (DES) and Structured Clinical Interview for DSM-IV Dissociative Disorders (SCID-D-R). We then provide recommendations for assessing complex trauma and dissociation through the aforementioned assessments. Copyright © 2016 Elsevier Ltd. All rights reserved.

A chemical model of seawater including dissolved ammonia and the stoichiometric dissociation constant of ammonia in estuarine water and seawater from -2 to 40°C

NASA Astrophysics Data System (ADS)

Clegg, Simon L.; Whitfield, Michael

1995-06-01

The calculation of the percentage of un-ionised ammonia in estuarine water and seawater requires values of the stoichiometric dissociation constant of ammonia, defined by: K*a/mol kg -1 = mNH 3mH +/ mNH +4, where m denotes molality. A thermodynamic model of seawater, including dissolved NH 3 and NH +4, is developed using an extended Pitzer formalism parameterised from available data. The model is validated using emf measurements for cells containing artificial seawater with added HCl, and NH 4Cl, and NH 3 over a range of temperatures and salinities. Calculated values of K*a are tabulated from 0 to 40 ppt salinity and -2 to 40°C, on both a free ( mH +) and total ( mH + + mHSO -4) hydrogen ion basis for use with pH measurements made on the corresponding scales. Accuracy (in K*a) is likely to be better than 5% at all temperatures and salinities.

Frequency-constant Q, unity and disorder

DOE Office of Scientific and Technical Information (OSTI.GOV)

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 Qmore » = {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.« less

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

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. Copyright © by the American Society of Agronomy, Crop Science Society of America, and Soil Science Society of America, Inc.

Analysis of molecular determinants of affinity and relative efficacy of a series of R- and S-2-(dipropylamino)tetralins at the 5-HT1A serotonin receptor

PubMed Central

Alder, J Tracy; Hacksell, Uli; Strange, Philip G

2003-01-01

Factors influencing agonist affinity and relative efficacy have been studied for the 5-HT1A serotonin receptor using membranes of CHO cells expressing the human form of the receptor and a series of R-and S-2-(dipropylamino)tetralins (nonhydroxylated and monohydroxylated (5-OH, 6-OH, 7-OH, 8-OH) species). Ligand binding studies were used to determine dissociation constants for agonist binding to the 5-HT1A receptor: Ki values for agonists were determined in competition versus the binding of the agonist [3H]-8-OH DPAT. Competition data were all fitted best by a one-binding site model.Ki values for agonists were also determined in competition versus the binding of the antagonist [3H]-NAD-199. Competition data were all fitted best by a two-binding site model, and agonist affinities for the higher (Kh) and lower affinity (Kl) sites were determined. The ability of the agonists to activate the 5-HT1A receptor was determined using stimulation of [35S]-GTPγS binding. Maximal effects of agonists (Emax) and their potencies (EC50) were determined from concentration/response curves for stimulation of [35S]-GTPγS binding. Kl/Kh determined from ligand binding assays correlated with the relative efficacy (relative Emax) of agonists determined in [35S]-GTPγS binding assays. There was also a correlation between Kl/Kh and Kl/EC50 for agonists determined from ligand binding and [35S]-GTPγS binding assays. Simulations of agonist binding and effect data were performed using the Ternary Complex Model in order to assess the use of Kl/Kh for predicting the relative efficacy of agonists. PMID:12684269

Dissociation of Rac1(GDP)·RhoGDI Complexes by the Cooperative Action of Anionic Liposomes Containing Phosphatidylinositol 3,4,5-Trisphosphate, Rac Guanine Nucleotide Exchange Factor, and GTP*

PubMed Central

Ugolev, Yelena; Berdichevsky, Yevgeny; Weinbaum, Carolyn; Pick, Edgar

2008-01-01

Rac plays a pivotal role in the assembly of the superoxide-generating NADPH oxidase of phagocytes. In resting cells, Rac is found in the cytosol in complex with Rho GDP dissociation inhibitor (RhoGDI). NADPH oxidase assembly involves dissociation of the Rac·RhoGDI complex and translocation of Rac to the membrane. We reported that liposomes containing high concentrations of monovalent anionic phospholipids cause Rac·RhoGDI complex dissociation (Ugolev, Y., Molshanski-Mor, S., Weinbaum, C., and Pick, E. (2006) J. Biol. Chem.281 ,19204 -1921916702219). We now designed an in vitro model mimicking membrane phospholipid remodeling during phagocyte stimulation in vivo. We showed that liposomes of “resting cell membrane” composition (less than 20 mol % monovalent anionic phospholipids), supplemented with 1 mol % of polyvalent anionic phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P3) in conjunction with constitutively active forms of the guanine nucleotide exchange factors (GEFs) for Rac, Trio, or Tiam1 and a non-hydrolyzable GTP analogue, cause dissociation of Rac1(GDP)·RhoGDI complexes, GDP to GTP exchange on Rac1, and binding of Rac1(GTP) to the liposomes. Complexes were not dissociated in the absence of GEF and GTP, and optimal dissociation required the presence of PtdIns(3,4,5)P3 in the liposomes. Dissociation of Rac1(GDP)·RhoGDI complexes was correlated with the affinity of particular GEF constructs, via the N-terminal pleckstrin homology domain, for PtdIns(3,4,5)P3 and involved GEF-mediated GDP to GTP exchange on Rac1. Phagocyte membranes enriched in PtdIns(3,4,5)P3 responded by NADPH oxidase activation upon exposure in vitro to Rac1(GDP)·RhoGDI complexes, p67phox, GTP, and Rac GEF constructs with affinity for PtdIns(3,4,5)P3 at a level superior to that of native membranes. PMID:18505730

High affinity binding of amyloid β-peptide to calmodulin: Structural and functional implications.

PubMed

Corbacho, Isaac; Berrocal, María; Török, Katalin; Mata, Ana M; Gutierrez-Merino, Carlos

2017-05-13

Amyloid β-peptides (Aβ) are a major hallmark of Alzheimer's disease (AD) and their neurotoxicity develop with cytosolic calcium dysregulation. On the other hand, calmodulin (CaM), a protein which plays a major multifunctional role in neuronal calcium signaling, has been shown to be involved in the regulation of non-amyloidogenic processing of amyloid β precursor protein (APP). Using fluorescent 6-bromoacetyl-2-dimethylaminonaphthalene derivatives of CaM, Badan-CaM, and human amyloid β(1-42) HiLyte™-Fluor555, we show in this work that Aβ binds with high affinity to CaM through the neurotoxic Aβ25-35 domain. In addition, the affinity of Aβ for calcium-saturated CaM conformation is approximately 20-fold higher than for CaM conformation in the absence of calcium (apo-CaM). Moreover, the value of K d of 0.98 ± 0.11 nM obtained for Aβ1-42 dissociation from CaM saturated by calcium points out that CaM is one of the cellular targets with highest affinity for neurotoxic Aβ peptides. A major functional consequence of Aβ-CaM interaction is that it slowdowns Aβ fibrillation. The novel and high affinity interaction between calmodulin and Aβ shown in this work opens a yet-unexplored gateway to further understand the neurotoxic effect of Aβ in different neural cells and also to address the potential of calmodulin and calmodulin-derived peptides as therapeutic agents in AD. Copyright © 2017 Elsevier Inc. All rights reserved.

The β subunit of the high-conductance calcium-activated potassium channel contributes to the high-affinity receptor for charybdotoxin

PubMed Central

Hanner, Markus; Schmalhofer, William A.; Munujos, Petraki; Knaus, Hans-Günther; Kaczorowski, Gregory J.; Garcia, Maria L.

1997-01-01

Transient expression of either α or α+β subunits of the high-conductance Ca2+-activated K+ (maxi-K) channel has been achieved in COS-1 cells. Expression has been studied using charybdotoxin (ChTX), a peptidyl inhibitor that binds in the pore on the α subunit. Although some properties of monoiodotyrosine-ChTX (125I-ChTX) binding to membranes derived from each type of transfected cells appear to be identical, other parameters of the binding reaction are markedly different. Under low ionic strength conditions, the affinity constant for 125I-ChTX measured under equilibrium binding conditions is increased ca. 50-fold in the presence of the β subunit. The rate constant for 125I-ChTX association is enhanced ca. 5-fold, whereas the dissociation rate constant is decreased more than 7-fold when the β subunit is present. These data indicate that functional coassembly of maxi-K channel subunits can be obtained in a transient expression system, and that the β subunit has profound effects on 125I-ChTX binding. We postulate that certain negatively charged residues in the large extracellular loop of β attract the positively charged 125I-ChTX to its binding site on α through electrostatic interactions, and account for effects observed on ligand association kinetics. Moreover, another residue(s) in the loop of β must contribute to stabilization of the toxin-bound state, either by a direct interaction with toxin, or through an allosteric effect on the α subunit. Certain regions in the extracellular loop of the β subunit may be in close proximity to the pore of the channel, and could play an important role in maxi-K channel function. PMID:9096310

Solitons and the energy-momentum tensor for affine Toda theory

NASA Astrophysics Data System (ADS)

Olive, D. I.; Turok, N.; Underwood, J. W. R.

1993-07-01

Following Leznov and Saveliev, we present the general solution to Toda field theories of conformal, affine or conformal affine type, associated with a simple Lie algebra g. These depend on a free massless field and on a group element. By putting the former to zero, soliton solutions to the affine Toda theories with imaginary coupling constant result with the soliton data encoded in the group element. As this requires a reformulation of the affine Kac-Moody algebra closely related to that already used to formulate the physical properties of the particle excitations, including their scattering matrices, a unified treatment of particles and solitons emerges. The physical energy—momentum tensor for a general solution is broken into a total derivative plus a part dependent only on the derivatives of the free field. Despite the non-linearity of the field equations and their complex nature the energy and momentum of the N-soliton solution is shown to be real, equalling the sum of contributions from the individual solitons. There are rank-g species of soliton, with masses given by a generalisation of a formula due to Hollowood, being proportional to the components of the left Perron-Frobenius eigenvector of the Cartan matrix of g.

[Translating Hysteria: PTSD dissociative subtype].

PubMed

Levy Yeyati, Elena

2016-03-01

To address the symptoms of depersonalization and derealization a specific dissociative subtype of posttraumatic stress disorder has been included in DSM-5. Depersonalization and derealization have been defined as acute and chronic symptoms of dissociative disorders since DSM-III. Dissociative disorders with chronic features are often diagnosed in North America, Canada and other European countries, but this is not the case in Argentina. Dissociative disorders have clinical and historical features that are strongly connected with hysteria. The latter is included in ICD-10 within the guidelines for diagnoses of dissociative disorders. This would be one of the reasons for not using dissociative disorders with chronic presentations of symptoms like depersonalization and derealization in countries other than North America or Canada: instead hysteria keeps on been used. The relation between trauma, dissociation and hysteria has been explained in different ways depending on the underlying theory (i.e. Janet's or Freud's). Janet's conceptualizations have had decisive influence in DSM formulations in chronic forms of dissociative disorders. Janet's and Freud's theories lead to different consequences on psychotherapy: hypnosis or psychoanalysis. The aim of this article is to show that the authors of PTSD dissociative subtype are in search of a cross-cultural validity of the construct which might be able to exert a more global influence.

[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. Copyright © 2014 L’Encéphale. Published by Elsevier Masson SAS.. All rights reserved.

[Dissociated learning with GABAergic drugs].

PubMed

Azarashvili, A A; Kaĭmachnikova, I E

2008-01-01

The possibility of dissociated learning was investigated using drugs which act directly on GABAB receptors of the brain. The earlier proposed suggestion that the cholinergic system plays a key role in the mechanisms of dissociated learning was tested. It was shown in male Wistar rats that dissociated learning was possible with GABAergic drugs. The dissociated state was induced by injecting the animals with both GABA agonist Baclofen and GABA antagonist 5-aminovaleric acid. Thus, dissociated learning is possible with drugs which act on either cholinergic or GABAergic transmitter systems.

Childhood Traumatic Experiences, Dissociative Symptoms, and Dissociative Disorder Comorbidity Among Patients With Panic Disorder: A Preliminary Study.

PubMed

Ural, Cenk; Belli, Hasan; Akbudak, Mahir; Tabo, Abdulkadir

2015-01-01

This study assessed childhood trauma history, dissociative symptoms, and dissociative disorder comorbidity in patients with panic disorder (PD). A total of 92 psychotropic drug-naive patients with PD, recruited from outpatient clinics in the psychiatry department of a Turkish hospital, were involved in the study. Participants were assessed using the Structured Clinical Interview for DSM-IV Dissociative Disorders (SCID-D), Dissociation Questionnaire, Panic and Agoraphobia Scale, Panic Disorder Severity Scale, and Childhood Trauma Questionnaire. Of the patients with PD, 18 (19%) had a comorbid dissociative disorder diagnosis on screening with the SCID-D. The most prevalent disorders were dissociative disorder not otherwise specified, dissociative amnesia, and depersonalization disorders. Patients with a high degree of dissociation symptoms and dissociative disorder comorbidity had more severe PD than those without (p < .05). All of the childhood trauma subscales used were correlated with the severity of symptoms of dissociation and PD. Among all of the subscales, the strongest relationship was with childhood emotional abuse. Logistic regression analysis showed that emotional abuse and severity of PD were independently associated with dissociative disorder. In our study, a significant proportion of the patients with PD had concurrent diagnoses of dissociative disorder. We conclude that the predominance of PD symptoms at admission should not lead the clinician to overlook the underlying dissociative process and associated traumatic experiences among these patients.

Binding site and affinity prediction of general anesthetics to protein targets using docking.

PubMed

Liu, Renyu; Perez-Aguilar, Jose Manuel; Liang, David; Saven, Jeffery G

2012-05-01

The protein targets for general anesthetics remain unclear. A tool to predict anesthetic binding for potential binding targets is needed. In this study, we explored whether a computational method, AutoDock, could serve as such a tool. High-resolution crystal data of water-soluble proteins (cytochrome C, apoferritin, and human serum albumin), and a membrane protein (a pentameric ligand-gated ion channel from Gloeobacter violaceus [GLIC]) were used. Isothermal titration calorimetry (ITC) experiments were performed to determine anesthetic affinity in solution conditions for apoferritin. Docking calculations were performed using DockingServer with the Lamarckian genetic algorithm and the Solis and Wets local search method (http://www.dockingserver.com/web). Twenty general anesthetics were docked into apoferritin. The predicted binding constants were compared with those obtained from ITC experiments for potential correlations. In the case of apoferritin, details of the binding site and their interactions were compared with recent cocrystallization data. Docking calculations for 6 general anesthetics currently used in clinical settings (isoflurane, sevoflurane, desflurane, halothane, propofol, and etomidate) with known 50% effective concentration (EC(50)) values were also performed in all tested proteins. The binding constants derived from docking experiments were compared with known EC(50) values and octanol/water partition coefficients for the 6 general anesthetics. All 20 general anesthetics docked unambiguously into the anesthetic binding site identified in the crystal structure of apoferritin. The binding constants for 20 anesthetics obtained from the docking calculations correlate significantly with those obtained from ITC experiments (P = 0.04). In the case of GLIC, the identified anesthetic binding sites in the crystal structure are among the docking predicted binding sites, but not the top ranked site. Docking calculations suggest a most probable binding site

Binding Site and Affinity Prediction of General Anesthetics to Protein Targets Using Docking

PubMed Central

Liu, Renyu; Perez-Aguilar, Jose Manuel; Liang, David; Saven, Jeffery G.

2012-01-01

Background The protein targets for general anesthetics remain unclear. A tool to predict anesthetic binding for potential binding targets is needed. In this study, we explore whether a computational method, AutoDock, could serve as such a tool. Methods High-resolution crystal data of water soluble proteins (cytochrome C, apoferritin and human serum albumin), and a membrane protein (a pentameric ligand-gated ion channel from Gloeobacter violaceus, GLIC) were used. Isothermal titration calorimetry (ITC) experiments were performed to determine anesthetic affinity in solution conditions for apoferritin. Docking calculations were performed using DockingServer with the Lamarckian genetic algorithm and the Solis and Wets local search method (https://www.dockingserver.com/web). Twenty general anesthetics were docked into apoferritin. The predicted binding constants are compared with those obtained from ITC experiments for potential correlations. In the case of apoferritin, details of the binding site and their interactions were compared with recent co-crystallization data. Docking calculations for six general anesthetics currently used in clinical settings (isoflurane, sevoflurane, desflurane, halothane, propofol, and etomidate) with known EC50 were also performed in all tested proteins. The binding constants derived from docking experiments were compared with known EC50s and octanol/water partition coefficients for the six general anesthetics. Results All 20 general anesthetics docked unambiguously into the anesthetic binding site identified in the crystal structure of apoferritin. The binding constants for 20 anesthetics obtained from the docking calculations correlate significantly with those obtained from ITC experiments (p=0.04). In the case of GLIC, the identified anesthetic binding sites in the crystal structure are among the docking predicted binding sites, but not the top ranked site. Docking calculations suggest a most probable binding site located in the

Dissociation and Memory Fragmentation in Posttraumatic Stress Disorder: An Evaluation of the Dissociative Encoding Hypothesis

PubMed Central

Bedard-Gilligan, Michele; Zoellner, Lori A.

2012-01-01

Several prominent theories of posttraumatic stress disorder (PTSD) posit that peritraumatic dissociation results in insufficient encoding of the trauma memory and that persistent dissociation prevents memory elaboration, resulting in memory fragmentation and PTSD. In this review, we summarize the empirical literature on peritraumatic and trait dissociation and trauma narrative fragmentation as measured by meta-memory and rater/objective coding. Across 16 studies to date, the association between dissociation and fragmentation was most prominent when examining peritraumatic dissociation and patient's own ratings of memory fragmentation. This relationship did not hold when examining trait dissociation or rater-coded or computer-generated measures of fragmentation. Thus, initial evidence points more toward a strong self-reported association between constructs that is not supported on more objective fragmentation coding. Measurement overlap, construct ambiguity, and exclusion of potential confounds may underlie lack of a strong association between dissociation and objective-rated fragmentation. PMID:22348400

Does phasic trauma treatment make patients with dissociative identity disorder treatment more dissociative?

PubMed

Brand, Bethany; Loewenstein, Richard J

2014-01-01

Proponents of the iatrogenic model of the etiology of dissociative identity disorder (DID) have expressed concern that treatment focused on direct engagement and interaction with dissociated self-states harms DID patients. However, empirical data have shown that this type of DID treatment is beneficial. Analyzing data from the prospective Treatment of Patients With Dissociative Disorders (TOP DD) Study, we test empirically whether DID treatment is associated with clinically adverse manifestations of dissociated self-states: acting so differently that one feels like different people, hearing voices, and dissociative amnesia. We show that, over the course of the study, there were significant decreases in feeling like different people and hearing voices. These results indicate that this form of DID treatment does not lead to symptomatic worsening in these dimensions, as predicted by the iatrogenic model. Indeed, treatment provided by TOP DD therapists reduced, rather than increased, the extent to which patients experienced manifestations of pathological dissociation. Because severe symptomatology and impairment are associated with DID, iatrogenic harm may come from depriving DID patients of treatment that targets DID symptomatology.

Collision-induced dissociation of diazirine-labeled peptide ions. Evidence for Brønsted-acid assisted elimination of nitrogen.

PubMed

Marek, Aleš; Tureček, František

2014-05-01

Gas-phase dissociations were investigated for several peptide ions containing the Gly-Leu* N-terminal motif where Leu* was a modified norleucine residue containing the photolabile diazirine ring. Collisional activation of gas-phase peptide cations resulted in facile N₂ elimination that competed with backbone dissociations. A free lysine ammonium group can act as a Brønsted acid to facilitate N₂ elimination. This dissociation was accompanied by insertion of a lysine proton in the side chain of the photoleucine residue, as established by deuterium labeling and gas-phase sequencing of the products. Electron structure calculations were used to provide structures and energies of reactants, intermediates, and transition states for Gly-Leu*-Gly-Gly-Lys amide ions that were combined with RRKM calculations of unimolecular rate constants. The calculations indicated that Brønsted acid-catalyzed eliminations were kinetically preferred over direct loss of N₂ from the diazirine ring. Mechanisms are proposed to explain the proton-initiated reactions and discuss the reaction products. The non-catalyzed diazirine ring cleavage and N₂ loss is proposed as a thermometer dissociation for peptide ion dissociations.

Pair interactions in polyelectrolyte-nanoparticle systems: Influence of dielectric inhomogeneities and the partial dissociation of polymers and nanoparticles.

PubMed

Pryamitsyn, Victor; Ganesan, Venkat

2015-10-28

We study the effective pair interactions between two charged spherical particles in polyelectrolyte solutions using polymer self-consistent field theory. In a recent study [V. Pryamitsyn and V. Ganesan, Macromolecules 47, 6095 (2015)], we considered a model in which the particles possess fixed charge density, the polymers contain a prespecified amount of dissociated charges and, the dielectric constant of the solution was assumed to be homogeneous in space and independent of the polymer concentration. In this article, we present results extending our earlier model to study situations in which either or both the particle and the polymers possess partially dissociable groups. Additionally, we also consider the case when the dielectric constant of the solution depends on the local concentration of the polymers and when the particle's dielectric constant is lower than that of the solvent. For each case, we quantify the polymer-mediated interactions between the particles as a function of the polymer concentrations and the degree of dissociation of the polymer and particles. Consistent with the results of our previous study, we observe that the polymer-mediated interparticle interactions consist of a short-range attraction and a long-range repulsion. The partial dissociablity of the polymer and particles was seen to have a strong influence on the strength of the repulsive portion of the interactions. Rendering the dielectric permittivity to be inhomogeneous has an even stronger effect on the repulsive interactions and results in changes to the qualitative nature of interactions in some parametric ranges.

Complexes of DOTA-bisphosphonate conjugates: probes for determination of adsorption capacity and affinity constants of hydroxyapatite.

PubMed

Vitha, Tomas; Kubícek, Vojtech; Hermann, Petr; Kolar, Zvonimir I; Wolterbeek, Hubert Th; Peters, Joop A; Lukes, Ivan

2008-03-04

The adsorption on hydroxyapatite of three conjugates of a bisphosphonate and a macrocycle having C1, C2, and C3 spacers and their terbium complexes was studied by the radiotracer method using 160Tb as the label. The radiotracer-containing complex of the conjugate with the C3 spacer was used as a probe for the determination of the adsorption parameters of other bisphosphonates that lack a DOTA unit. A physicochemical model describing the competitive adsorption was successfully applied in the fitting of the obtained data. The maximum adsorption capacity of bisphosphonates containing bulky substituents is determined mainly by their size. For bisphosphonates having no DOTA moiety, the maximum adsorption capacity is determined by the electrostatic repulsion between negatively charged bisphosphonate groups. Compounds with a hydroxy or amino group attached to the alpha-carbon atom show higher affinities. Macrocyclic compounds containing a short spacer between the different bisphosphonic acid groups and the macrocyclic unit exhibit high affinities, indicating a synergic effect of the bisphosphonic and the macrocyclic groups during adsorption. The competition method described uses a well-characterized complex and allows a simple evaluation of the adsorption behavior of bisphosphonates. The application of the macrocycle-bisphosphonate conjugates allows easy radiolabeling via complexation of a suitable metal isotope.

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.

Dissociative diffusion mechanism in vacancy-rich materials according to mass action kinetics

DOE Office of Scientific and Technical Information (OSTI.GOV)

Biderman, N. J.; Sundaramoorthy, R.; Haldar, Pradeep

We conducted two sets of diffusion-reaction numerical simulations using a finite difference method (FDM) in order to investigate fast impurity diffusion via interstitial sites in vacancy-rich materials such as Cu(In,Ga)Se 2 (CIGS) and Cu 2ZnSn(S, Se) 4 (CZTSSe or CZTS) via the dissociative diffusion mechanism where the interstitial diffuser ultimately reacts with a vacancy to produce a substitutional. The first set of simulations extends the standard interstitial-limited dissociative diffusion theory to vacancy-rich material conditions where vacancies are annihilated in large amounts, introducing non-equilibrium vacancy concentration profiles. The second simulation set explores the vacancy-limited dissociative diffusion where impurity incorporation increases themore » equilibrium vacancy concentration. In addition to diffusion profiles of varying concentrations and shapes that were obtained in all simulations, some of the profiles can be fitted with the constant- and limited-source solutions of Fick’s second law despite the non-equilibrium condition induced by the interstitial-vacancy reaction. The first set of simulations reveals that the dissociative diffusion coefficient in vacancy-rich materials is inversely proportional to the initial vacancy concentration. In the second set of numerical simulations, impurity-induced changes in the vacancy concentration lead to distinctive diffusion profile shapes. The simulation results are also compared with published data of impurity diffusion in CIGS. And according to the characteristic properties of diffusion profiles from the two set of simulations, experimental detection of the dissociative diffusion mechanism in vacancy-rich materials may be possible.« less

Dissociative diffusion mechanism in vacancy-rich materials according to mass action kinetics

DOE PAGES

Biderman, N. J.; Sundaramoorthy, R.; Haldar, Pradeep; ...

2016-05-13

We conducted two sets of diffusion-reaction numerical simulations using a finite difference method (FDM) in order to investigate fast impurity diffusion via interstitial sites in vacancy-rich materials such as Cu(In,Ga)Se 2 (CIGS) and Cu 2ZnSn(S, Se) 4 (CZTSSe or CZTS) via the dissociative diffusion mechanism where the interstitial diffuser ultimately reacts with a vacancy to produce a substitutional. The first set of simulations extends the standard interstitial-limited dissociative diffusion theory to vacancy-rich material conditions where vacancies are annihilated in large amounts, introducing non-equilibrium vacancy concentration profiles. The second simulation set explores the vacancy-limited dissociative diffusion where impurity incorporation increases themore » equilibrium vacancy concentration. In addition to diffusion profiles of varying concentrations and shapes that were obtained in all simulations, some of the profiles can be fitted with the constant- and limited-source solutions of Fick’s second law despite the non-equilibrium condition induced by the interstitial-vacancy reaction. The first set of simulations reveals that the dissociative diffusion coefficient in vacancy-rich materials is inversely proportional to the initial vacancy concentration. In the second set of numerical simulations, impurity-induced changes in the vacancy concentration lead to distinctive diffusion profile shapes. The simulation results are also compared with published data of impurity diffusion in CIGS. And according to the characteristic properties of diffusion profiles from the two set of simulations, experimental detection of the dissociative diffusion mechanism in vacancy-rich materials may be possible.« less

Direction-dependent force-induced dissociation dynamics of an entropic-driven lock-and-key assembly.

PubMed

Chen, Yen-Fu; Chen, Hsuan-Yi; Sheng, Yu-Jane; Tsao, Heng-Kwong

2017-09-01

The unbinding dynamics of a nanosized sphere-and-cavity assembly under the pulling of constant force and constant loading rate is explored by dissipative particle dynamics simulations. The formation of this matched lock-and-key pair in a polymer solution is driven by the depletion attraction. The two-dimensional free energy landscape U(x,z) associated with this assembly is constructed. Our results indicate that the unbinding pathway along the orientation of the assembly is unfavorable due to the relatively high energy barrier compared to that along the tortuous minimum path whose energy barrier is not high. It is also found that the dissociation rate depends on the direction of the external force (θ) with respect to the assembly orientation. The presence of the force component perpendicular to the assembly orientation can reduce the bond lifetime significantly by driving the key particle to approach the minimum path. Moreover, the dissociation dynamics can be facilitated even by a pushing force compared to the spontaneous dissociation (without forces). To elucidate the effective pathway under pulling, the escaping position is analyzed and its mean direction with respect to the assembly orientation rises generally with increasing θ, revealing that the presence of the force component along the minimum pathway is helpful. The importance of the direction of the external pulling has been demonstrated in our simple system. Therefore, this effect should be considered in more complicated unbinding experiments.

Direction-dependent force-induced dissociation dynamics of an entropic-driven lock-and-key assembly

NASA Astrophysics Data System (ADS)

Chen, Yen-Fu; Chen, Hsuan-Yi; Sheng, Yu-Jane; Tsao, Heng-Kwong

2017-09-01

The unbinding dynamics of a nanosized sphere-and-cavity assembly under the pulling of constant force and constant loading rate is explored by dissipative particle dynamics simulations. The formation of this matched lock-and-key pair in a polymer solution is driven by the depletion attraction. The two-dimensional free energy landscape U (x ,z ) associated with this assembly is constructed. Our results indicate that the unbinding pathway along the orientation of the assembly is unfavorable due to the relatively high energy barrier compared to that along the tortuous minimum path whose energy barrier is not high. It is also found that the dissociation rate depends on the direction of the external force (θ ) with respect to the assembly orientation. The presence of the force component perpendicular to the assembly orientation can reduce the bond lifetime significantly by driving the key particle to approach the minimum path. Moreover, the dissociation dynamics can be facilitated even by a pushing force compared to the spontaneous dissociation (without forces). To elucidate the effective pathway under pulling, the escaping position is analyzed and its mean direction with respect to the assembly orientation rises generally with increasing θ , revealing that the presence of the force component along the minimum pathway is helpful. The importance of the direction of the external pulling has been demonstrated in our simple system. Therefore, this effect should be considered in more complicated unbinding experiments.

Dissociation of Laboratory-Synthesized Methane Hydrate in Coarse-Grained Sediments by Slow Depressurization

NASA Astrophysics Data System (ADS)

Phillips, S. C.; You, K.; Borgfeldt, T.; Meyer, D.; Dong, T.; Flemings, P. B.

2016-12-01

We performed four dissociation experiments in which experimentally-formed methane hydrate was dissociated via slow, stepwise depressurization, revealing in situ salinity conditions. Overall, these results suggest the occurrence of local pore water freshening around dissociating hydrate in which bulk equilibrium behavior is limited by salt diffusion. Depressurization was performed at a constant confining temperature over 1 to 3 weeks by releasing small volumes of methane gas from the top of a vertically-oriented sample into an inverted graduated cylinder. We identify three distinct regimes of depressurization based on pressure drop behavior: (1) release of free gas down to initial hydrate dissociation at 3.3 MPa in NaBr or 4.64 MPa in NaCl, (2) dissociation of methane hydrate characterized by a slow, logarithmic increase in pressure after each gas release and (3) residual free gas release. Initial hydrate dissociation in NaCl brine at 4.64 MPa corresponds to the phase boundary for hydrate in 9.6 wt% NaCl. In the NaCl experiment, pressure increases of 0.16 MPa while the sample was shut in over 3 days likely correspond to a recovery in salinity of 0.7 wt. %. Salt ions likely diffuse from brine ahead of the hydrate front, based on a length scale for diffusion of NaCl of 6.3 cm for 3 days. In this experiment dissociation at bulk equilibrium is expected to decline from 4.54 to 4.04 MPa; however actual dissociation during 73 gas releases over 15 days, results in a pressure drop from 4.64 to 3.25 MPa. Hydrate samples were formed by injection of methane gas at 1 ºC and 12.24 MPa within a cylinder packed with medium-grained quartz sand and initially saturated in a 7 wt% NaBr or NaCl solution. In two experiments in which the system was thoroughly leak tested, total methane consumed during formation and recovered during depressurization match within 7% indicating this approach to be relatively accurate for determining total methane in experimental or pressure core samples.

The 5-HT1A Receptor PET Radioligand 11C-CUMI-101 Has Significant Binding to α1-Adrenoceptors in Human Cerebellum, Limiting Its Use as a Reference Region.

PubMed

Shrestha, Stal S; Liow, Jeih-San; Jenko, Kimberly; Ikawa, Masamichi; Zoghbi, Sami S; Innis, Robert B

2016-12-01

Prazosin, a potent and selective α 1 -adrenoceptor antagonist, displaces 25% of 11 C-CUMI-101 ([O-methyl- 11 C]2-(4-(4-(2-methoxyphenyl)piperazin-1-yl)butyl)-4-methyl-1,2,4-triazine-3,5(2H,4H)dione) binding in monkey cerebellum. We sought to estimate the percentage contamination of 11 C-CUMI-101 binding to α 1 -adrenoceptors in human cerebellum under in vivo conditions. In vitro receptor-binding techniques were used to measure α 1 -adrenoceptor density and the affinity of CUMI-101 for these receptors in human, monkey, and rat cerebellum. Binding potential (maximum number of binding sites × affinity [(1/dissociation constant]) was determined using in vitro homogenate binding assays in human, monkey, and rat cerebellum. 3 H-prazosin was used to determine the maximum number of binding sites, as well as the dissociation constant of 3 H-prazosin and the inhibition constant of CUMI-101. α 1 -adrenoceptor density and the affinity of CUMI-101 for these receptors were similar across species. Cerebellar binding potentials were 3.7 for humans, 2.3 for monkeys, and 3.4 for rats. Reasoning by analogy, 25% of 11 C-CUMI-101 uptake in human cerebellum reflects binding to α 1 -adrenoceptors, suggesting that the cerebellum is of limited usefulness as a reference tissue for quantification in human studies. © 2016 by the Society of Nuclear Medicine and Molecular Imaging, Inc.

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

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

PubMed

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, K d) 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 K d (range 2.0-7.3 nM for 16 Richardson's ground squirrels, Urocitellus richardsonii). The K d 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 K d 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.

Low-energy collision induced dissociation (low-energy CID), collision induced dissociation (CID) and higher-energy collision dissociation (HCD) mass spectrometry for structural elucidation of saccharides and clarification of their dissolution mechanism in DMAc/LiCl.

PubMed

Bayat, Parisa; Lesage, Denis; Cole, Richard B

2018-05-29

The dissolution mechanism of oligosaccharides in N,N-dimethylacetamide/lithium chloride (DMAc/LiCl), a solvent used for cellulose dissolution, and the capabilities of low-energy collision induced dissociation (low-energy CID), collision induced dissociation (CID) and higher-energy collision dissociation (HCD) for structural analysis of carbohydrates were investigated. Comparing the spectra obtained using three techniques shows that, generally, when working with mono-lithiated sugars, CID spectra provide more structurally informative fragments, and glycosidic bond cleavage is the main pathway. However, when working with di-lithiated sugars, HCD spectra can be more informative providing predominately cross-ring cleavage fragments. This is because HCD is a non-resonant activation technique and it allows a higher amount of energy to be deposited in a short time, giving access to more endothermic decomposition pathways as well as consecutive fragmentations. The difference in preferred dissociation pathways of mono-lithiated and di-lithiated sugars indicates that the presence of the second lithium strongly influences the relative rate constants for cross-ring cleavages (rearrangement) vs. direct glycosidic bond cleavages, and disfavors the latter. Regarding the dissolution mechanism of sugars in DMAc/LiCl, CID and HCD experiments on di-lithiated and tri-lithiated sugars reveal that intensities of product ions containing two Li + or three Li + , respectively, are higher than those bearing only one Li + . In addition, comparing the fragmentation spectra (both HCD and CID) of LiCl adducted lithiated sugar and NaCl adducted sodiated sugar shows that while, in the latter case, loss of NaCl is dominant, in the former case, loss of HCl occurs preferentially. The compiled evidence implies that there is a strong and direct interaction between lithium and the saccharide during the dissolution process in the DMAc/LiCl solvent system. This article is protected by copyright. All

Dissociated learning using GABAergic drugs.

PubMed

Azarashvili, A A; Kaimachnikova, I E

2009-02-01

Experiments on Wistar rats addressed the possibility of dissociated learning using drugs acting directly on brain GABA(B) receptors. A previously suggested hypothesis was tested: that the cholinergic system of the brain plays the decisive role in the mechanisms of dissociative learning. The data obtained here provided evidence that dissociated learning an occur with compounds acting on the GABAergic transmitter system of the brain. Dissociated states arose on treatment of animals with both the GABA-mimetic baclofen and the GABA receptor antagonist 5-aminovaleric acid. Thus, these results show that dissociated learning can occur using drugs acting on both the cholinergic and the GABAergic transmitter systems of the brain.

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". Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Dissociative symptoms and dissociative disorders comorbidity in obsessive compulsive disorder: Symptom screening, diagnostic tools and reflections on treatment

PubMed Central

Belli, Hasan

2014-01-01

Borderline personality disorder, conversion disorder and obsessive compulsive disorder frequently have dissociative symptoms. The literature has demonstrated that the level of dissociation might be correlated with the severity of obsessive compulsive disorder (OCD) and that those not responding to treatment had high dissociative symptoms. The structured clinical interview for DSM-IV dissociative disorders, dissociation questionnaire, somatoform dissociation questionnaire and dissociative experiences scale can be used for screening dissociative symptoms and detecting dissociative disorders in patients with OCD. However, a history of neglect and abuse during childhood is linked to a risk factor in the pathogenesis of dissociative psychopathology in adults. The childhood trauma questionnaire-53 and childhood trauma questionnaire-40 can be used for this purpose. Clinicians should not fail to notice the hidden dissociative symptoms and childhood traumatic experiences in OCD cases with severe symptoms that are resistant to treatment. Symptom screening and diagnostic tools used for this purpose should be known. Knowing how to treat these pathologies in patients who are diagnosed with OCD can be crucial. PMID:25133142

Dissociative symptoms and dissociative disorders comorbidity in obsessive compulsive disorder: Symptom screening, diagnostic tools and reflections on treatment.

PubMed

Belli, Hasan

2014-08-16

Borderline personality disorder, conversion disorder and obsessive compulsive disorder frequently have dissociative symptoms. The literature has demonstrated that the level of dissociation might be correlated with the severity of obsessive compulsive disorder (OCD) and that those not responding to treatment had high dissociative symptoms. The structured clinical interview for DSM-IV dissociative disorders, dissociation questionnaire, somatoform dissociation questionnaire and dissociative experiences scale can be used for screening dissociative symptoms and detecting dissociative disorders in patients with OCD. However, a history of neglect and abuse during childhood is linked to a risk factor in the pathogenesis of dissociative psychopathology in adults. The childhood trauma questionnaire-53 and childhood trauma questionnaire-40 can be used for this purpose. Clinicians should not fail to notice the hidden dissociative symptoms and childhood traumatic experiences in OCD cases with severe symptoms that are resistant to treatment. Symptom screening and diagnostic tools used for this purpose should be known. Knowing how to treat these pathologies in patients who are diagnosed with OCD can be crucial.

Molybdenum-carbon bond dissociation energies in Mo(CO) sub 6

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ganske, J.A.; Rosenfeld, R.N.

1990-05-17

The pressure dependence of the recombination rate constants for Mo(CO){sub n} (n = 3,4 and 5) with CO has been studied by time-resolved infrared laser absorption spectroscopy. These data, in conjunction with an RRKM model for unimolecular decay of the activated molecules (Mo(CO){sub 6})*, (Mo(CO){sub 5})*, and (Mo(CO){sub 4})*, have allowed the determination of the bond dissociation energies for several of the Mo-C bonds in Mo(CO){sub 6}. The first Mo-C bond dissociation energies for Mo(CO){sub 6}, Mo(CO){sub 5}, and Mo(CO){sub 4} are found to be DH{degree}((CO){sub 5}Mo{hor ellipsis}CO) = 35 {plus minus} 5 kcal/mol, DH{degree}((CO){sub 4}Mo{hor ellipsis}CO) = 27 {plusmore » minus} 5 kcal/mol, and DH{degree}((CO){sub 3}Mo{hor ellipsis}CO) = 31 {plus minus} 5 kcal/mol.« less

Dissociative experiences in patients with epilepsy.

PubMed

Özdemir, Osman; Cilingir, Vedat; Özdemir, Pınar Güzel; Milanlioglu, Aysel; Hamamci, Mehmet; Yilmaz, Ekrem

2016-03-01

A few studies have explored dissociative experiences in epilepsy patients. We investigated dissociative experiences in patients with epilepsy using the dissociative experiences scale (DES). Ninety-eight patients with epilepsy and sixty healthy controls were enrolled in this study. A sociodemographic questionnaire and the Dissociative Experiences Scale (DES), Beck Depression Inventory (BDI) and Beck Anxiety Inventory (BAI) were administered to the participants. The DES scores were significantly higher for the patients with epilepsy than the healthy individuals. The number of individuals with pathological dissociation (DES ≥ 30) was higher in the epilepsy group (n = 28) than in the control group (n = 8). Also, higher levels of dissociation were significantly associated with frequency of seizures, but were not associated with duration of epilepsy and age at onset of the disorder. These findings demonstrate that patients with epilepsy are more prone to dissociation than controls. The high rate of dissociative experiences among patients with epilepsy suggest that some epilepsy-related factors are present.

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.

Engineering an Affinity-Enhanced Peptide through Optimization of Cyclization Chemistry.

PubMed

Ngambenjawong, Chayanon; Pineda, Julio Marco B; Pun, Suzie H

2016-12-21

Peptide cyclization is a strategy used to improve stability and activity of peptides. The most commonly used cyclization method is disulfide bridge formation of cysteine-containing peptides, as is typically found in nature. Over the years, an increasing number of alternative chemistries for peptide cyclization with improved efficiency, kinetics, orthogonality, and stability have been reported. However, there has been less appreciation for the opportunity to fine-tune peptide activity via the diverse chemical entities introduced at the site of linkage by different cyclization strategies. Here, we demonstrate how cyclization optimization of an M2 "anti-inflammatory" macrophage-binding peptide (M2pep) resulted in a significant increase in binding affinity of the optimized analog to M2 macrophages while maintaining binding selectivity compared to M1 "pro-inflammatory" macrophages. In this study, we report synthesis and evaluation of four cyclic M2pep(RY) analogs with diverse cyclization strategies: (1) Asp-[amide]-Lys, (2) azido-Lys-[triazole(copper(I)-catalyzed alkyne-azide cycloaddition (CuAAC))]-propargyl-Gly, (3) Cys-[decafluorobiphenyl (DFBP)]-Cys, and (4) Cys-[decafluorobiphenyl sulfone (DFS)]-Cys, whereby the chemical entity or linker at the linkage site is shown in the square bracket and is between the residues involved in cyclization. These peptides are compared to a disulfide-cyclized M2pep(RY) that we previously reported as a serum-stable, affinity-enhanced analog to the original linear M2pep. DFBP-cyclized M2pep(RY) exhibits the highest binding activity to M2 macrophages with apparent dissociation constant (K D ) about 2.03 μM compared to 36.3 μM for the original disulfide-cyclized M2pep(RY) and 220 μM for the original linear peptide. DFS-cyclized M2pep(RY) also binds more strongly than the original cyclized analog, whereas amide- and triazole-cyclized M2pep(RY) analogs bind less strongly. We verified that DFBP alone has negligible binding to M2

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.

Multiphoton dissociation and thermal unimolecular reactions induced by infrared lasers. [REAMPA code

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dai, H.L.

1981-04-01

Multiphoton dissociation (MPD) of ethyl chloride was studied using a tunable 3.3 ..mu..m laser to excite CH stretches. The absorbed energy increases almost linearly with fluence, while for 10 ..mu..m excitation there is substantial saturation. Much higher dissociation yields were observed for 3.3 ..mu..m excitation than for 10 ..mu..m excitation, reflecting bottlenecking in the discrete region of 10 ..mu..m excitation. The resonant nature of the excitation allows the rate equations description for transitions in the quasicontinuum and continuum to be extended to the discrete levels. Absorption cross sections are estimated from ordinary ir spectra. A set of cross sections whichmore » is constant or slowly decreasing with increasing vibrational excitation gives good fits to both absorption and dissociation yield data. The rate equations model was also used to quantitatively calculate the pressure dependence of the MPD yield of SF/sub 6/ caused by vibrational self-quenching. Between 1000-3000 cm/sup -1/ of energy is removed from SF/sub 6/ excited to approx. > 60 kcal/mole by collision with a cold SF/sub 6/ molecule at gas kinetic rate. Calculation showed the fluence dependence of dissociation varies strongly with the gas pressure. Infrared multiphoton excitation was applied to study thermal unimolecular reactions. With SiF/sub 4/ as absorbing gas for the CO/sub 2/ laser pulse, transient high temperature pulses were generated in a gas mixture. IR fluorescence from the medium reflected the decay of the temperature. The activation energy and the preexponential factor of the reactant dissociation were obtained from a phenomenological model calculation. Results are presented in detail. (WHK)« less

Threshold collision-induced dissociation and theoretical study of protonated azobenzene

NASA Astrophysics Data System (ADS)

Rezaee, Mohammadreza; McNary, Christopher P.; Armentrout, P. B.

2017-10-01

Protonated azobenzene (AB), H+(C6H5N2C6H5), has been studied using threshold collision-induced dissociation in a guided ion beam tandem mass spectrometer. Product channels observed are C6H5N2+ + C6H6 and C6H5+ + N2 + C6H6. The experimental kinetic energy-dependent cross sections were analyzed using a statistical model that accounts for internal and kinetic energy distributions of the reactants, multiple collisions, and kinetic shifts. From this analysis, the activation energy barrier height of 2.02 ± 0.11 eV for benzene loss is measured. To identify the transition states (TSs) and intermediates (IMs) for these dissociations, relaxed potential energy surface (PES) scans were performed at the B3LYP/aug-cc-pVTZ level of theory. The PES indicates that there is a substantial activation energy along the dissociation reaction coordinate that is the rate-limiting step for benzene loss and at some levels of theory, for subsequent N2 loss as well. Relative energies of the reactant, TSs, IMs, and products were calculated at B3LYP, wB97XD, M06, PBEPBE, and MP2(full) levels of theory using both 6-311++G(2d,2p) and aug-cc-pVTZ basis sets. Comparison of the experimental results with theoretical values from various computational methods indicates how well these theoretical methods can predict thermochemical properties. In addition to these density functional theory and MP2 methods, several high accuracy multi-level calculations such as CBS-QB3, G3, G3MP2, G3B3MP2, G4, and G4MP2 were performed to determine the thermochemical properties of AB including the proton affinity and gas-phase basicity, and to compare the performance of different theoretical methods.

Trends in electron-ion dissociative recombination of benzene analogs with functional group substitutions: Negative Hammett σpara values

NASA Astrophysics Data System (ADS)

Osborne, David; Lawson, Patrick Andrew; Adams, Nigel; Dotan, Itzhak

2014-06-01

An in-depth study of the effects of functional group substitution on benzene's electron-ion dissociative recombination (e-IDR) rate constant has been conducted. The e-IDR rate constants for benzene, biphenyl, toluene, ethylbenzene, anisole, phenol, and aniline have been measured using a Flowing Afterglow equipped with an electrostatic Langmuir probe (FALP). These measurements have been made over a series of temperatures from 300 to 550 K. A relationship between the Hammett σpara values for each compound and rate constant has indicated a trend in the e-IDR rate constants and possibly in their temperature dependence data. The Hammett σpara value is a method to describe the effect a functional group substituted to a benzene ring has upon the reaction rate constant.

Isotherm-Based Thermodynamic Models for Solute Activities of Organic Acids with Consideration of Partial Dissociation.

PubMed

Nandy, Lucy; Ohm, Peter B; Dutcher, Cari S

2016-06-23

Organic acids make up a significant fraction of the organic mass in atmospheric aerosol particles. The calculation of gas-liquid-solid equilibrium partitioning of the organic acid is therefore critical for accurate determination of atmospheric aerosol physicochemical properties and processes such as new particle formation and activation to cloud condensation nuclei. Previously, an adsorption isotherm-based statistical thermodynamic model was developed for capturing solute concentration-activity relationships for multicomponent aqueous solutions over the entire concentration range (Dutcher et al. J. Phys. Chem. C/A 2011, 2012, 2013), with model parameters for energies of adsorption successfully related to dipole-dipole electrostatic forces in solute-solvent and solvent-solvent interactions for both electrolytes and organics (Ohm et al. J. Phys. Chem. A 2015). However, careful attention is needed for weakly dissociating semivolatile organic acids. Dicarboxylic acids, such as malonic acid and glutaric acid are treated here as a mixture of nondissociated organic solute (HA) and dissociated solute (H(+) + A(-)). It was found that the apparent dissociation was greater than that predicted by known dissociation constants alone, emphasizing the effect of dissociation on osmotic and activity coefficient predictions. To avoid additional parametrization from the mixture approach, an expression was used to relate the Debye-Hückel hard-core collision diameter to the adjustable solute-solvent intermolecular distance. An improved reference state treatment for electrolyte-organic aqueous mixtures, such as that observed here with partial dissociation, has also been proposed. This work results in predictive correlations for estimation of organic acid and water activities for which there is little or no activity data.

Lp-dual affine surface area

NASA Astrophysics Data System (ADS)

Wei, Wang; Binwu, He

2008-12-01

According to the notion of Lp-affine surface area by Lutwak, in this paper, we introduce the concept of Lp-dual affine surface area. Further, we establish the affine isoperimetric inequality and the Blaschke-Santaló inequality for Lp-dual affine surface area. Besides, the dual Brunn-Minkowski inequality for Lp-dual affine surface area is presented.

Experimentally-induced dissociation impairs visual memory.

PubMed

Brewin, Chris R; Mersaditabari, Niloufar

2013-12-01

Dissociation is a phenomenon common in a number of psychological disorders and has been frequently suggested to impair memory for traumatic events. In this study we explored the effects of dissociation on visual memory. A dissociative state was induced experimentally using a mirror-gazing task and its short-term effects on memory performance were investigated. Sixty healthy individuals took part in the experiment. Induced dissociation impaired visual memory performance relative to a control condition; however, the degree of dissociation was not associated with lower memory scores in the experimental group. The results have theoretical and practical implications for individuals who experience frequent dissociative states such as patients with posttraumatic stress disorder (PTSD). Copyright © 2013 Elsevier Inc. All rights reserved.

Speech-Language Dissociations, Distractibility, and Childhood Stuttering

PubMed Central

Conture, Edward G.; Walden, Tedra A.; Lambert, Warren E.

2015-01-01

Purpose This study investigated the relation among speech-language dissociations, attentional distractibility, and childhood stuttering. Method Participants were 82 preschool-age children who stutter (CWS) and 120 who do not stutter (CWNS). Correlation-based statistics (Bates, Appelbaum, Salcedo, Saygin, & Pizzamiglio, 2003) identified dissociations across 5 norm-based speech-language subtests. The Behavioral Style Questionnaire Distractibility subscale measured attentional distractibility. Analyses addressed (a) between-groups differences in the number of children exhibiting speech-language dissociations; (b) between-groups distractibility differences; (c) the relation between distractibility and speech-language dissociations; and (d) whether interactions between distractibility and dissociations predicted the frequency of total, stuttered, and nonstuttered disfluencies. Results More preschool-age CWS exhibited speech-language dissociations compared with CWNS, and more boys exhibited dissociations compared with girls. In addition, male CWS were less distractible than female CWS and female CWNS. For CWS, but not CWNS, less distractibility (i.e., greater attention) was associated with more speech-language dissociations. Last, interactions between distractibility and dissociations did not predict speech disfluencies in CWS or CWNS. Conclusions The present findings suggest that for preschool-age CWS, attentional processes are associated with speech-language dissociations. Future investigations are warranted to better understand the directionality of effect of this association (e.g., inefficient attentional processes → speech-language dissociations vs. inefficient attentional processes ← speech-language dissociations). PMID:26126203

Objective documentation of child abuse and dissociation in 12 murderers with dissociative identity disorder.

PubMed

Lewis, D O; Yeager, C A; Swica, Y; Pincus, J H; Lewis, M

1997-12-01

The skepticism regarding the existence of dissociative identity disorder as well as the abuse that engenders it persists for lack of objective documentation. This is doubly so for the disorder in murderers because of issues of suspected malingering. This article presents objective verification of both dissociative symptoms and severe abuse during childhood in a series of adult murderers with dissociative identity disorder. This study consisted of a review of the clinical records of 11 men and one woman with DSM-IV-defined dissociative identity disorder who had committed murder. Data were gathered from medical, psychiatric, social service, school, military, and prison records and from records of interviews with subjects' family members and others. Handwriting samples were also examined. Data were analyzed qualitatively. Signs and symptoms of dissociative identity disorder in childhood and adulthood were corroborated independently and from several sources in all 12 cases; objective evidence of severe abuse was obtained in 11 cases. The subjects had amnesia for most of the abuse and underreported it. Marked changes in writing style and/or signatures were documented in 10 cases. This study establishes, once and for all, the linkage between early severe abuse and dissociative identity disorder. Further, the data demonstrate that the disorder can be distinguished from malingering and from other disorders. The study shows that it is possible, with great effort, to obtain objective evidence of both the symptoms of dissociative identity disorder and the abuse that engenders it.

Dissociation in the laboratory: a comparison of strategies.

PubMed

Leonard, K N; Telch, M J; Harrington, P J

1999-01-01

Several methods for inducing dissociation in the laboratory were examined in a sample of 78 undergraduate students. Participants scoring high or low on the Dissociative Experiences Scale participated in three dissociation challenge conditions: (a) dot-staring task, (b) administration of pulsed photic and audio stimulation and (c) stimulus deprivation. Participants recorded their dissociative experiences both before and after each of the three challenge conditions. Across conditions, high DES participants reported significantly more dissociative sensations than low DES participants, even after controlling for pre-challenge dissociation. Moreover, regardless of DES status, pulsed photo and audio stimulation produced the greatest level of dissociative symptoms. The findings suggest that the induction of dissociative symptoms in a nonclinical sample is easily accomplished in the laboratory and that those who report more dissociative symptoms in their day-to-day life exhibit more pronounced dissociative symptoms when undergoing dissociative challenge in the laboratory. Implications for the study and treatment of dissociative symptoms are discussed.

Dissociative, depressive, and PTSD symptom severity as correlates of nonsuicidal self-injury and suicidality in dissociative disorder patients.

PubMed

Webermann, Aliya R; Myrick, Amie C; Taylor, Christina L; Chasson, Gregory S; Brand, Bethany L

2016-01-01

The present study investigates whether symptom severity can distinguish patients diagnosed with dissociative identity disorder and dissociative disorder not otherwise specified with a recent history of nonsuicidal self-injury (NSSI) and suicide attempts from those patients without recent self-harm. A total of 241 clinicians reported on recent history of patient NSSI and suicide attempts. Of these clinicians' patients, 221 completed dissociative, depressive, and posttraumatic stress disorder symptomatology measures. Baseline cross-sectional data from a naturalistic and prospective study of dissociative disorder patients receiving community treatment were utilized. Analyses evaluated dissociative, depressive, and posttraumatic stress disorder symptom severity as methods of classifying patients into NSSI and suicide attempt groupings. Results indicated that dissociation severity accurately classified patients into NSSI and suicidality groups, whereas depression severity accurately classified patients into NSSI groups. These findings point to dissociation and depression severity as important correlates of NSSI and suicidality in patients with dissociative disorders and have implications for self-harm prevention and treatment.

Label-free Growth Receptor-2 Detection and Dissociation Constant Assessment in Diluted Human Serum Using a Longitudinal Extension Mode of a Piezoelectric Microcantilever Sensor

PubMed Central

Capobianco, Joseph A.; Shih, Wan Y.; Adams, Gregory P.; Shih, Wei-Heng

2011-01-01

We have investigated real-time, label-free, in-situ detection of human epidermal growth factor receptor 2 (Her2) in diluted serum using the first longitudinal extension mode of a lead zirconate-lead titanate (PZT)/glass piezoelectric microcantilever sensor (PEMS) with H3 single-chain variable fragment (scFv) immobilized on the 3-mercaptopropyltrimethoxysilane (MPS) insulation layer of the PEMS surface. We showed that with the longitudinal extension mode, the PZT/glass PEMS consisting of a 1 mm long and 127 μm thick PZT layer bonded with a 75 μm thick glass layer with a 1.8 mm long glass tip could detect Her2 at a concentration of 6-60 ng/ml (or 0.06-0.6 nM) in diluted human serum, about 100 times lower than the concentration limit obtained using the lower-frequency flexural mode of a similar PZT/glass PEMS. We further showed that with the longitudinal mode, the PZT/glass PEMS determined the equilibrium H3-Her2 dissociation constant Kd to be 3.3±0.3 × 10-8 M consistent with the value, 3.2±0.28 ×10-8 M deduced by the surface plasmon resonance method (BIAcore). PMID:22888196

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(+).

Blackbody infrared radiative dissociation of oligonucleotide anions.

PubMed

Klassen, J S; Schnier, P D; Williams, E R

1998-11-01

The dissociation kinetics of a series of doubly deprotonated oligonucleotide 7-mers [d(A)7(2-), d(AATTAAT)2-, d(TTAATTA)2-, and d(CCGGCCG)2-] were measured using blackbody infrared radiative dissociation in a Fourier-transform mass spectrometer. The oligonucleotides dissociate first by cleavage at the glycosidic bond leading to the loss of a neutral nucleobase, followed by cleavage at the adjacent (5') phosphodiester bond to produce structurally informative a-base and w type ions. From the temperature dependence of the unimolecular dissociation rate constants, Arrhenius activation parameters in the zero-pressure limit are obtained for the loss of base. The measured Arrhenius parameters are dependent on the identity of the nucleobase. The process involving the loss of an adenine base from the dianions, d(A)7(2-), d(AATTAAT)2-, and d(TTAATTA)2- has an average activation energy (Ea) of approximately 1.0 eV and a preexponential factor (A) of 10(10) s-1. Both guanine and cytosine base loss occurs for d(CCGGCCG)2-. The average Arrhenius parameters for the loss of cytosine and guanine are Ea = 1.32 +/- 0.03 eV and A = 10(13.3 +/- 0.3) s-1. No loss of thymine was observed for mixed adenine-thymine oligonucleotides. Neither base loss nor any other fragmentation reactions occur for d(T)7(2-) over a 600 s reaction delay at 207 degrees C, a temperature close to the upper limit accessible with our instrument. The Arrhenius parameters indicate that the preferred cleavage sites for mixed oligonucleotides of similar mass-to-charge ratio will be strongly dependent on the internal energy of the precursor ions. At low internal energies (effective temperatures below 475 K), loss of adenine and subsequent cleavage of the adjacent phosphoester bonds will dominate, whereas at higher energies, preferential cleavage at C and G residues will occur. The magnitude of the A factors < or = 10(13) s-1 measured for the loss of the three nucleobases (A, G, and C) is indicative of an entropically

Blackbody Infrared Radiative Dissociation of Oligonucleotide Anions

PubMed Central

Klassen, John S.; Schnier, Paul D.; Williams, Evan R.

2005-01-01

The dissociation kinetics of a series of doubly deprotonated oligonucleotide 7-mers [ d(A)72-, d(AATTAAT)2−, d(TTAATTA)2−, and d(CCGGCCG)2−] were measured using blackbody infrared radiative dissociation in a Fourier-transform mass spectrometer. The oligonucleotides dissociate first by cleavage at the glycosidic bond leading to the loss of a neutral nucleobase, followed by cleavage at the adjacent (5′) phosphodiester bond to produce structurally informative a-base and w type ions. From the temperature dependence of the unimolecular dissociation rate constants, Arrhenius activation parameters in the zero-pressure limit are obtained for the loss of base. The measured Arrhenius parameters are dependent on the identity of the nucleobase. The process involving the loss of an adenine base from the dianions, d(A)72-, d(AATTAAT)2−, and d(TTAATTA)2− has an average activation energy (Ea) of ~1.0 eV and a preexponential factor (A) of 1010 s−1. Both guanine and cytosine base loss occurs for d(CCGGCCG)2−. The average Arrhenius parameters for the loss of cytosine and guanine are Ea = 1.32 ± 0.03 eV and A = 1013.3±0.3 s−1. No loss of thymine was observed for mixed adenine–thymine oligonucleotides. Neither base loss nor any other fragmentation reactions occur for d(T)72- over a 600 s reaction delay at 207 °C, a temperature close to the upper limit accessible with our instrument. The Arrhenius parameters indicate that the preferred cleavage sites for mixed oligonucleotides of similar mass-to-charge ratio will be strongly dependent on the internal energy of the precursor ions. At low internal energies (effective temperatures below 475 K), loss of adenine and subsequent cleavage of the adjacent phosphoester bonds will dominate, whereas at higher energies, preferential cleavage at C and G residues will occur. The magnitude of the A factors ≤1013 s−1 measured for the loss of the three nucleobases (A, G, and C) is indicative of an entropically neutral or

Exploring Thermoresponsive Affinity Agents to Enhance Microdialysis Sampling Efficiency of Proteins

NASA Astrophysics Data System (ADS)

Vasicek, Thaddeus

Affinity agents increase microdialysis protein relative recovery, yet they have not seen widespread use within the microdialysis community due to their additional instrumentation requirements and prohibitive cost. This dissertation describes new affinity agents for microdialysis that require no additional instrumentation to use, have nearly 100% particle recovery, are 7 times more cost efficient than alternatives, and have low specificity enabling their use for a wide variety of proteins. Initially gold nanoparticles were chosen as an affinity ligand support due to their high surface area/volume ratio and colloidal stability. Poly (N-isopropylacrylamide) was immobilized to the gold nanoparticles, which served to sterically stabilize the particles and to act as a generic, reversible protein capture agent. A method was developed to reproducibly vary and quantify poly (N-isopropylacrylamide) graft density from 0.09 to 0.40 ligands/nm2 on gold nanoparticles. During characterization of the polymer coated gold nanoparticles, irreversible particle agglomeration was observed at low polymer graft density in ionic solutions, which prevented further development as a protein capture agent. Poly (N-isopropylacrylamide) nanogels, which have low nonspecific adsorption, low interparticle attractive forces owing to the low curvature of the particle, and a low Hamaker constant, were synthesized to overcome the agglomeration problem. A generic protein affinity ligand cibacron blue, was immobilized to the nanogels, which enabled rapid determination of particle recovery. The perfusion of the nanogels through a microdialysis probe was optimized yielding 100% particle recovery using a combination of a syringe and peristaltic pump. The microdialysis collection efficiency of CCL2, a physiologically relevant cytokine, was increased 3-fold with addition of the nanogel to the microdialysis perfusion fluid. The reduction in instrumentation requirements, low cost, and low specificity obtained

Fatty acids bind tightly to the N-terminal domain of angiopoietin-like protein 4 and modulate its interaction with lipoprotein lipase.

PubMed

Robal, Terje; Larsson, Mikael; Martin, Miina; Olivecrona, Gunilla; Lookene, Aivar

2012-08-24

Angiopoietin-like protein 4 (Angptl4), a potent regulator of plasma triglyceride metabolism, binds to lipoprotein lipase (LPL) through its N-terminal coiled-coil domain (ccd-Angptl4) inducing dissociation of the dimeric enzyme to inactive monomers. In this study, we demonstrate that fatty acids reduce the inactivation of LPL by Angptl4. This was the case both with ccd-Angptl4 and full-length Angptl4, and the effect was seen in human plasma or in the presence of albumin. The effect decreased in the sequence oleic acid > palmitic acid > myristic acid > linoleic acid > linolenic acid. Surface plasmon resonance, isothermal titration calorimetry, fluorescence, and chromatography measurements revealed that fatty acids bind with high affinity to ccd-Angptl4. The interactions were characterized by fast association and slow dissociation rates, indicating formation of stable complexes. The highest affinity for ccd-Angptl4 was detected for oleic acid with a subnanomolar equilibrium dissociation constant (K(d)). The K(d) values for palmitic and myristic acid were in the nanomolar range. Linoleic and linolenic acid bound with much lower affinity. On binding of fatty acids, ccd-Angptl4 underwent conformational changes resulting in a decreased helical content, weakened structural stability, dissociation of oligomers, and altered fluorescence properties of the Trp-38 residue that is located close to the putative LPL-binding region. Based on these results, we propose that fatty acids play an important role in modulating the effects of Angptl4.

Investigation of the Mechanism of Electron Capture and Electron Transfer Dissociation of Peptides with a Covalently Attached Free Radical Hydrogen Atom Scavenger.

PubMed

Sohn, Chang Ho; Yin, Sheng; Peng, Ivory; Loo, Joseph A; Beauchamp, J L

2015-11-15

The mechanisms of electron capture and electron transfer dissociation (ECD and ETD) are investigated by covalently attaching a free-radical hydrogen atom scavenger to a peptide. The 2,2,6,6-tetramethylpiperidin-l-oxyl (TEMPO) radical was chosen as the scavenger due to its high hydrogen atom affinity (ca. 280 kJ/mol) and low electron affinity (ca. 0.45 ev), and was derivatized to the model peptide, FQX TEMPO EEQQQTEDELQDK. The X TEMPO residue represents a cysteinyl residue derivatized with an acetamido-TEMPO group. The acetamide group without TEMPO was also examined as a control. The gas phase proton affinity (882 kJ/mol) of TEMPO is similar to backbone amide carbonyls (889 kJ/mol), minimizing perturbation to internal solvation and sites of protonation of the derivatized peptides. Collision induced dissociation (CID) of the TEMPO tagged peptide dication generated stable odd-electron b and y type ions without indication of any TEMPO radical induced fragmentation initiated by hydrogen abstraction. The type and abundance of fragment ions observed in the CID spectra of the TEMPO and acetamide tagged peptides are very similar. However, ECD of the TEMPO labeled peptide dication yielded no backbone cleavage. We propose that a labile hydrogen atom in the charge reduced radical ions is scavenged by the TEMPO radical moiety, resulting in inhibition of N-C α backbone cleavage processes. Supplemental activation after electron attachment (ETcaD) and CID of the charge-reduced precursor ion generated by electron transfer of the TEMPO tagged peptide dication produced a series of b + H (b H ) and y + H (y H ) ions along with some c ions having suppressed intensities, consistent with stable O-H bond formation at the TEMPO group. In summary, the results indicate that ECD and ETD backbone cleavage processes are inhibited by scavenging of a labile hydrogen atom by the localized TEMPO radical moiety. This observation supports the conjecture that ECD and ETD processes involve long

Covalent attachment of Arc repressor subunits by a peptide linker enhances affinity for operator DNA.

PubMed

Robinson, C R; Sauer, R T

1996-01-09

By designing a recombinant gene containing tandem copies of the arc coding sequence with intervening DNA encoding the linker sequence GGGSGGGTGGGSGGG, the two subunits of the P22 Are repressor dimer have been covalently linked to form a single-chain protein called Arc-L1-Arc. The 15-residue linker joins the C-terminus of one monomer to the N-terminus of the second, a distance of approximately 45 A in the Arc-operator cocrystal structure. Arc-L1-Arc is expressed at high levels in Escherichia coli, with no evidence of degradation or proteolytic clipping of the linker, and is more active than wild-type Arc in repression assays. The purified Arc-L1-Arc protein has the molecular weight expected for the designed protein and unfolds cooperatively, reversibly, and with no concentration dependence in thermal-denaturation studies. Arc-L1-Arc protects operator DNA in a manner indistinguishable from that of wild-type Arc in DNase I and copper-phenanthroline footprinting studies, but the covalent attachment of the two monomers results in enhanced affinity for operator DNA. Arc-L1-Arc binds operator DNA half-maximally at a concentration of 1.7 pM, compared with the wild-type value of 185 pM, and also binds DNA fragments containing the left or right operator half-sites more tightly than wild type. Because wild-type Arc is monomeric at sub-nanomolar concentrations and must dimerize before binding to the operator, it was anticipated that Arc-L1-Arc would exhibit a lower half-maximal binding concentration. However, even when the change from a monomeric to a dimeric species is taken into account, the affinity of Arc-L1-Arc for operator and half-operator DNA is greater than the wild-type affinity. This tighter binding appears to result from slower dissociation, as Arc-L1-Arc DNA complexes with full or half-site operators dissociate at rates 5-10 times slower than the corresponding Arc--DNA complexes. Hence, the activity of the designed Arc-L1-Arc protein is substantially increased

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

The Dissociative Subtype of Posttraumatic Stress Disorder (PTSD) Among Adolescents: Co-Occurring PTSD, Depersonalization/Derealization, and Other Dissociation Symptoms.

PubMed

Choi, Kristen R; Seng, Julia S; Briggs, Ernestine C; Munro-Kramer, Michelle L; Graham-Bermann, Sandra A; Lee, Robert C; Ford, Julian D

2017-12-01

The purpose of this study was to examine the co-occurrence of posttraumatic stress disorder (PTSD) and dissociation in a clinical sample of trauma-exposed adolescents by evaluating evidence for the depersonalization/derealization dissociative subtype of PTSD as defined by the DSM-5 and then examining a broader set of dissociation symptoms. A sample of treatment-seeking, trauma-exposed adolescents 12 to 16 years old (N = 3,081) from the National Child Traumatic Stress Network Core Data Set was used to meet the study objectives. Two models of PTSD/dissociation co-occurrence were estimated using latent class analysis, one with 2 dissociation symptoms and the other with 10 dissociation symptoms. After model selection, groups within each model were compared on demographics, trauma characteristics, and psychopathology. Model A, the depersonalization/derealization model, had 5 classes: dissociative subtype/high PTSD; high PTSD; anxious arousal; dysphoric arousal; and a low symptom/reference class. Model B, the expanded dissociation model, identified an additional class characterized by dissociative amnesia and detached arousal. These 2 models provide new information about the specific ways PTSD and dissociation co-occur and illuminate some differences between adult and adolescent trauma symptom expression. A dissociative subtype of PTSD can be distinguished from PTSD alone in adolescents, but assessing a wider range of dissociative symptoms is needed to fully characterize adolescent traumatic stress responses. Copyright © 2017 American Academy of Child and Adolescent Psychiatry. Published by Elsevier Inc. All rights reserved.

Intrinsic Thermodynamics and Structures of 2,4- and 3,4-Substituted Fluorinated Benzenesulfonamides Binding to Carbonic Anhydrases.

PubMed

Zubrienė, Asta; Smirnov, Alexey; Dudutienė, Virginija; Timm, David D; Matulienė, Jurgita; Michailovienė, Vilma; Zakšauskas, Audrius; Manakova, Elena; Gražulis, Saulius; Matulis, Daumantas

2017-01-20

The goal of rational drug design is to understand structure-thermodynamics correlations in order to predict the chemical structure of a drug that would exhibit excellent affinity and selectivity for a target protein. In this study we explored the contribution of added functionalities of benzenesulfonamide inhibitors to the intrinsic binding affinity, enthalpy, and entropy for recombinant human carbonic anhydrases (CA) CA I, CA II, CA VII, CA IX, CA XII, and CA XIII. The binding enthalpies of compounds possessing similar chemical structures and affinities were found to be very different, spanning a range from -90 to +10 kJ mol -1 , and are compensated by a similar opposing entropy contribution. The intrinsic parameters of binding were determined by subtracting the linked protonation reactions. The sulfonamide group pK a values of the compounds were measured spectrophotometrically, and the protonation enthalpies were measured by isothermal titration calorimetry (ITC). Herein we describe the development of meta- or ortho-substituted fluorinated benzenesulfonamides toward the highly potent compound 10 h, which exhibits an observed dissociation constant value of 43 pm and an intrinsic dissociation constant value of 1.1 pm toward CA IX, an anticancer target that is highly overexpressed in various tumors. Fluorescence thermal shift assays, ITC, and X-ray crystallography were all applied in this work. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

The computational analysis and modelling of substitution effects on hydrolysis of formanilides in acidic aqueous solutions

NASA Astrophysics Data System (ADS)

Lukeš, Vladimír; Škorňa, Peter; Michalík, Martin; Klein, Erik

2017-11-01

Various para, meta and ortho substituted formanilides have been theoretically studied. For trans and cis-isomers of non-substituted formanilide, the calculated B3LYP vibration normal modes were analyzed. Substituent effect on the selected normal modes was described and the comparison with the available experimental data is presented. The calculated B3LYP proton affinities were correlated with Hammett constants, Fujita-Nishioka equation and the rate constants of the hydrolysis in 1 M HCl. Found linear dependences allow predictions of dissociation constants (pKBH+) and hydrolysis rate constants. Obtained results indicate that protonation of amide group may represent the rate determining step of acid catalyzed hydrolysis.

Accelerated dissociation of IgE:FcεRI complexes by disruptive inhibitors actively desensitizes allergic effector cells

PubMed Central

Eggel, Alexander; Baravalle, Günther; Hobi, Gabriel; Kim, Beomkyu; Buschor, Patrick; Forrer, Patrik; Shin, Jeoung-Sook; Vogel, Monique; Stadler, Beda M.; Dahinden, Clemens A.; Jardetzky, Theodore S.

2014-01-01

Background The remarkably stable interaction of immunoglobulin E (IgE) with its high-affinity receptor FcεRI on basophils and mast cells is critical for the induction of allergic hypersensitivity reactions. Due to the exceptionally slow dissociation rate of IgE:FcεRI complexes such allergic effector cells permanently display allergen-specific IgE on their surface and immediately respond to allergen challenge by releasing inflammatory mediators. We have recently described a novel macromolecular inhibitor that actively promotes the dissociation of IgE from FcεRI through a molecular mechanism termed facilitated dissociation. Objective Here, we assessed the therapeutic potential of this non-immunoglobulin based IgE inhibitor DARPin E2_79 as well as a novel engineered biparatopic DARPin bi53_79 and directly compared them to the established anti-IgE antibody omalizumab. Methods: IgE:FcεRI complex dissociation was analyzed in vitro using recombinant proteins in ELISA and surface plasmon resonance, ex vivo using human primary basophils with flow cytometry and in vivo using human FcεRI transgenic mice in a functional passive cutaneous anaphylaxis test. Results We show that E2_79 mediated removal of IgE from primary human basophils fully abrogates IgE-dependent cell activation and release of pro-inflammatory mediators ex vivo. Furthermore, we report that omalizumab also accelerates the dissociation of IgE from FcεRI albeit much less efficiently than E2_79. Using the biparatopic IgE targeting approach we further improved the disruptive potency of E2_79 by ~100 fold and show that disruptive IgE inhibitors efficiently prevent passive cutaneous anaphylaxis in mice expressing the human FcεRI alpha chain. Conclusion Our findings highlight the potential of such novel IgE inhibitors as important diagnostic and therapeutic tools to managing allergic diseases. PMID:24642143

Gallium metal affinity capture tandem mass spectrometry for the selective detection of phosphopeptides in complex mixtures

PubMed Central

Blacken, Grady R.; Sadílek, Martin; Tureček, František

2008-01-01

Metal affinity capture tandem mass spectrometry (MAC-MSMS) is evaluated in a comparative study of a lysine-derived nitrilotriacetic acid (Nα, Nα-bis-(carboxymethyl)lysine, LysNTA) and an aspartic-acid-related iminodiacetic acid (N-(4-aminobutyl)aspartic acid, AspIDA) as selective phosphopeptide detection reagents. Both LysNTA and AspIDA spontaneously form ternary complexes with GaIII and phosphorylated amino acids and phosphopeptides upon mixing in solution. Collision-induced dissociation of positive complex ions produced by electrospray produces common fragments (LysNTA + H)+ or (AspIDA + H)+ at m/z 263 and 205, respectively. MSMS precursor scans using these fragments as reporter ions allow one to selectively detect multiple charge states of phosphopeptides in mixtures. It follows from this comparative study that LysNTA is superior to AspIDA in detecting phosphopeptides, possibly because of the higher coordination number and greater stability constant for GaIII – phosphopeptide complexation of the former reagent. In a continuing development of MAC-MSMS for proteomics applications, we demonstrate its utility in a post-column reaction format. Using a simple post-column-reaction ‘T’ and syringe pump to deliver our chelating reagents, α-casein tryptic phosphopeptides can be selectively analyzed from a solution containing a twofold molar excess of bovine serum albumin. The MAC-MSMS method is shown to be superior to the commonly used neutral loss scan for the common loss of phosphoric acid. PMID:18265438

Dissociative amnesia in dissociative disorders and borderline personality disorder: self-rating assessment in a college population.

PubMed

Sar, Vedat; Alioğlu, Firdevs; Akyuz, Gamze; Karabulut, Sercan

2014-01-01

Dissociative amnesia (DA) among subjects with a dissociative disorder and/or borderline personality disorder (BPD) recruited from a nonclinical population was examined. The Steinberg Dissociative Amnesia Questionnaire (SDAQ), the Childhood Trauma Questionnaire, and the self-report screening tool of the BPD section of the Structured Clinical Interview for DSM-IV(SCID-BPD) were administered to 1,301 college students. A total of 80 participants who were diagnosed with BPD according to the clinician-administered SCID-BPD and 111 nonborderline controls were evaluated using the Structured Clinical Interview for DSM-IV Dissociative Disorders (SCID-D) by a psychiatrist blind to diagnosis and scale scores. Internal consistency analyses and test-retest evaluations suggested that the SDAQ is a reliable instrument for the population studied. Of the participants, 20.6% reported an SDAQ score of 20 or above and impairment by DA. Those who had both dissociative disorder and BPD (n = 78) had the highest SDAQ scores. Both disorders had significant effects on the SCID-D total and amnesia scores in the variance analysis. On SDAQ scores, however, only BPD had this effect. There was a significant interaction between the 2 disorders for the SCID-D total but not for the SDAQ or SCID-D amnesia scores. BPD represented the severity of dissociation and childhood trauma in this study group. However, in contrast to the dissociative disorders, BPD was characterized by better awareness of DA in self-report. The discrepancies between self-report and clinical interview associated with BPD and dissociative disorders are discussed in the context of betrayal theory (J. J. Freyd, 1994) of BPD and perceptual theory (D. B. Beere, 2009) of dissociative disorders.

Traceless affinity labeling of endogenous proteins for functional analysis in living cells.

PubMed

Hayashi, Takahiro; Hamachi, Itaru

2012-09-18

Protein labeling and imaging techniques have provided tremendous opportunities to study the structure, function, dynamics, and localization of individual proteins in the complex environment of living cells. Molecular biology-based approaches, such as GFP-fusion tags and monoclonal antibodies, have served as important tools for the visualization of individual proteins in cells. Although these techniques continue to be valuable for live cell imaging, they have a number of limitations that have only been addressed by recent progress in chemistry-based approaches. These chemical approaches benefit greatly from the smaller probe sizes that should result in fewer perturbations to proteins and to biological systems as a whole. Despite the research in this area, so far none of these labeling techniques permit labeling and imaging of selected endogenous proteins in living cells. Researchers have widely used affinity labeling, in which the protein of interest is labeled by a reactive group attached to a ligand, to identify and characterize proteins. Since the first report of affinity labeling in the early 1960s, efforts to fine-tune the chemical structures of both the reactive group and ligand have led to protein labeling with excellent target selectivity in the whole proteome of living cells. Although the chemical probes used for affinity labeling generally inactivate target proteins, this strategy holds promise as a valuable tool for the labeling and imaging of endogenous proteins in living cells and by extension in living animals. In this Account, we summarize traceless affinity labeling, a technique explored mainly in our laboratory. In our overview of the different labeling techniques, we emphasize the challenge of designing chemical probes that allow for dissociation of the affinity module (often a ligand) after the labeling reaction so that the labeled protein retains its native function. This feature distinguishes the traceless labeling approach from the traditional

Constants and thermodynamics of the acid-base equilibria of triglycine in water-ethanol solutions containing sodium perchlorate at 298 K

NASA Astrophysics Data System (ADS)

Pham Tkhi, L.; Usacheva, T. R.; Tukumova, N. V.; Koryshev, N. E.; Khrenova, T. M.; Sharnin, V. A.

2016-02-01

The acid-base equilibrium constants for glycyl-glycyl-glycine (triglycine) in water-ethanol solvents containing 0.0, 0.1, 0.3, and 0.5 mole fractions of ethanol are determined by potentiometric titration at 298.15 K and an ionic strength of 0.1, maintained with sodium perchlorate. It is established that an increase in the ethanol content in the solvent reduces the dissociation constant of the carboxyl group of triglycine (increases p K 1) and increases the dissociation constant of the amino group of triglycine (decreases p K 2). It is noted that the weakening of the acidic properties of a triglycinium ion upon an increase of the ethanol content in the solvent is due to the attenuation of the solvation shell of the zwitterionic form of triglycine, and to the increased solvation of triglycinium ions. It is concluded that the acid strength of triglycine increases along with a rise in the EtOH content in the solvent, due to the desolvation of the tripeptide zwitterion and the enhanced solvation of protons.

Self-Injury and Suicide Attempt in Relation with Trauma and Dissociation among Adolescents with Dissociative and Non-Dissociative Disorders.

PubMed

Kılıç, Filiz; Coşkun, Murat; Bozkurt, Hasan; Kaya, İlyas; Zoroğlu, Salih

2017-03-01

To explore the role of trauma and dissociation over self-injurious behaviors (SIB) and suicide attempts (SA) in adolescents. A total of 207 adolescents participated in the study. After conducting diagnostic interview, participants were divided into five groups as subjects with dissociative disorders (DD), attention deficit hyperactivity disorder (ADHD), major depressive disorder (MDD) and anxiety disorders (AD), and a control group (CG) without any psychiatric disorder. ADHD, MDD and AD groups were considered as non-dissociative disorders (non-DD group) in the present study. There is no significant difference between groups in terms of number and age of the subjects (p>0.05). Among all participants SIB was reported in 32.2% of females (n=37) and 25% of males (n=23) while SA was reported in 29.6% of females (n=34) and 4.4% of males (n=4). Adolescents with DD were found to experience higher rates of SIB and SA than the other groups. Dissociation was the most important variable contributing to SIB and female gender was the most efficient variable for SA. Total trauma scores were also found to be significantly higher in DD group followed by non-DD and CG respectively. SIB and SA are complex behavioral problems which may be associated with many psychiatric factors. However higher level dissociation seems as an important mediating factor, even regardless of psychiatric diagnosis, in the development of SIB and SA. More research is needed to further explore the factors effective over SIB and SA in adolescents.

[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

Dissociative Experiences and Vividness of Auditory Imagery

ERIC Educational Resources Information Center

Pérez-Fabello, María José; Campos, Alfredo

2017-01-01

The relationship between dissociation and auditory imagery were assessed, 2 variables that sometime influence on artistic creativity. A total of 170 fine arts undergraduates (94 women and 76 men) received 2 dissociation questionnaires--the Dissociative Ability Scale (DAS), and the Dissociative Experiences Scale (DES)--and 2 auditory imagery…

Kinetic Characterisation of a Single Chain Antibody against the Hormone Abscisic Acid: Comparison with Its Parental Monoclonal

PubMed Central

Badescu, George O.; Marsh, Andrew; Smith, Timothy R.; Thompson, Andrew J.; Napier, Richard M.

2016-01-01

A single-chain Fv fragment antibody (scFv) specific for the plant hormone abscisic acid (ABA) has been expressed in the bacterium Escherichia coli as a fusion protein. The kinetics of ABA binding have been measured using surface plasmon resonance spectrometry (BIAcore 2000) using surface and solution assays. Care was taken to calculate the concentration of active protein in each sample using initial rate measurements under conditions of partial mass transport limitation. The fusion product, parental monoclonal antibody and the free scFv all have low nanomolar affinity constants, but there is a lower dissociation rate constant for the parental monoclonal resulting in a three-fold greater affinity. Analogue specificity was tested and structure-activity binding preferences measured. The biologically-active (+)-ABA enantiomer is recognised with an affinity three orders of magnitude higher than the inactive (-)-ABA. Metabolites of ABA including phaseic acid, dihydrophaseic acid and deoxy-ABA have affinities over 100-fold lower than that for (+)-ABA. These properties of the scFv make it suitable as a sensor domain in bioreporters specific for the naturally occurring form of ABA. PMID:27023768

A quantitative analysis of the effects of 2,3-diphosphoglycerate, adenosine triphosphate and inositol hexaphosphate on the oxygen dissociation curve of human haemoglobin.

PubMed Central

Goodford, P J; St-Louis, J; Wootton, R

1978-01-01

1. Oxygen dissociation curves have been measured for human haemoglobin solutions with different concentrations of the allosteric effectors 2,3-diphosphoglycerate, adenosine triphosphate and inositol hexaphosphate. 2. Each effector produces a concentration dependent right shift of the oxygen dissociation curve, but a point is reached where the shift is maximal and increasing the effector concentration has no further effect. 3. Mathematical models based on the Monod, Wyman & Changeux (1965) treatment of allosteric proteins have been fitted to the data. For each compound the simple two-state model and its extension to take account of subunit inequivalence were shown to be inadequate, and a better fit was obtained by allowing the effector to lower the oxygen affinity of the deoxy conformational state as well as binding preferentially to this conformation. PMID:722582

Reconsidering the autohypnotic model of the dissociative disorders.

PubMed

Dell, Paul F

2018-03-22

The dissociative disorders field and the hypnosis field currently reject the autohypnotic model of the dissociative disorders, largely because many correlational studies have shown hypnotizability and dissociation to be minimally related (r = .12). Curiously, it is also widely accepted that dissociative patients are highly hypnotizable. If dissociative patients are highly hypnotizable because only highly hypnotizable individuals can develop a dissociative disorder - as the author proposes - then the methodology of correlational studies of hypnotizability and dissociation in random clinical and community samples would necessarily be constitutively unable to detect, and statistically unable to reflect, that fact. That is, the autohypnotic, dissociative distancing of that small subset of highly hypnotizable individuals who repeatedly encountered intolerable circumstances is statistically lost among the data of (1) the highly hypnotizable subjects who do not dissociate and (2) subjects (of all levels of hypnotizability) who manifest other kinds of dissociation. The author proposes that, when highly hypnotizable individuals repeatedly engage in autohypnotic distancing from intolerable circumstances, they develop an overlearned, highly-motivated, automatized pattern of dissociative self-protection (i.e., a dissociative disorder). The author urges that theorists of hypnosis and the dissociative disorders explicitly include in their theories (a) the trait of high hypnotizability, (b) the phenomena of autohypnosis, and (c) the manifestations of systematized, autohypnotic pathology. Said differently, the author is suggesting that autohypnosis and autohypnotic pathology are unacknowledged nodes in the nomothetic networks of both hypnosis and dissociation.

Testing the diagnosis of dissociative identity disorder through measures of dissociation, absorption, hypnotizability and PTSD: a Norwegian pilot study.

PubMed

Dale, Karl Yngvar; Berg, Renate; Elden, Ake; Ødegård, Atle; Holte, Arne

2009-01-01

A total of 14 women meeting criteria for dissociative identity disorder (DID) based on the Diagnostic and Statistical Manual of Mental Disorders (4th ed. [DSM-IV]) were compared to a group of women (n = 10) with other dissociative diagnoses and a group of normal controls (n = 14) with regard to dissociativity, absorption, trauma related symptoms and hypnotizability. Both of the clinical groups reported histories of childhood trauma and attained high PTSD scores. The DID group differed significantly from the group with other dissociative diagnoses and the non-diagnosed comparison group with regard to hypnotizability, the variety of dissociative symptomatology, and the magnitude of dissociative symptomatology. However, no significant differences between the two clinical groups were detected with regard to absorption, general dissociative level, or symptoms related to traumatic stress. Results support the notion that DID can be regarded as a clinical entity which is separable from other dissociative disorders. Results also indicated that hypnotizability is the most important clinical feature of DID.

Cabazitaxel is more active than first-generation taxanes in ABCB1(+) cell lines due to its reduced affinity for P-glycoprotein.

PubMed

Duran, George E; Derdau, Volker; Weitz, Dietmar; Philippe, Nicolas; Blankenstein, Jörg; Atzrodt, Jens; Sémiond, Dorothée; Gianolio, Diego A; Macé, Sandrine; Sikic, Branimir I

2018-04-19

The primary aim of this study was to determine cabazitaxel's affinity for the ABCB1/P-glycoprotein (P-gp) transporter compared to first-generation taxanes. We determined the kinetics of drug accumulation and retention using [ 14 C]-labeled taxanes in multidrug-resistant (MDR) cells. In addition, membrane-enriched fractions isolated from doxorubicin-selected MES-SA/Dx5 cells were used to determine sodium orthovanadate-sensitive ATPase stimulation after exposure to taxanes. Custom [ 3 H]-azido-taxane analogues were synthesized for the photoaffinity labeling of P-gp. The maximum intracellular drug concentration was achieved faster with [ 14 C]-cabazitaxel (5 min) than [ 14 C]-docetaxel (15-30 min). MDR cells accumulated twice as much cabazitaxel than docetaxel, and these levels could be restored to parental levels in the presence of the P-gp inhibitor PSC-833 (valspodar). Efflux in drug-free medium confirmed that MDR cells retained twice as much cabazitaxel than docetaxel. There was a strong association (r 2  = 0.91) between the degree of taxane resistance conferred by P-gp expression and the accumulation differences observed with the two taxanes. One cell model expressing low levels of P-gp was not cross-resistant to cabazitaxel while demonstrating modest resistance to docetaxel. Furthermore, there was a 1.9 × reduction in sodium orthovanadate-sensitive ATPase stimulation resulting from treatment with cabazitaxel compared to docetaxel. We calculated a dissociation constant (Kd) value of 1.7 µM for [ 3 H]-azido-docetaxel and ~ 7.5 µM for [ 3 H]-azido-cabazitaxel resulting in a 4.4 × difference in P-gp labeling, and cold docetaxel was a more effective competitor than cabazitaxel. Our studies confirm that cabazitaxel is more active in ABCB1(+) cell models due to its reduced affinity for P-gp compared to docetaxel.

Dissociative tendencies and facilitated emotional processing.

PubMed

Oathes, Desmond J; Ray, William J

2008-10-01

Dissociation is a process linked to lapses of attention, history of abuse or trauma, compromised emotional memory, and a disintegrated sense of self. It is theorized that dissociation stems from avoiding emotional information, especially negative emotion, to protect a fragile psyche. The present study tested whether or not dissociaters do actually avoid processing emotion by asking groups scoring high or low on the Dissociative Experiences Scale to judge the affective valence of several types of emotional stimuli. Manipulations of valence, modality (pictures or words), task complexity, and personal relevance lead to results suggesting that dissociation is linked to facilitated rather than deficient emotional processing. Our results are consistent with a theory that sensitivity to emotional material may be a contributing factor in subsequent dissociation to avoid further elaboration of upsetting emotion in these individuals. The findings for dissociation further exemplify the influence of individual differences in the link between cognition and emotion. (c) 2008 APA, all rights reserved

Dissociation in undergraduate students: disruptions in executive functioning.

PubMed

Giesbrecht, Timo; Merckelbach, Harald; Geraerts, Elke; Smeets, Ellen

2004-08-01

The concept of dissociation refers to disruptions in attentional control. Attentional control is an executive function. Few studies have addressed the link between dissociation and executive functioning. Our study investigated this relationship in a sample of undergraduate students (N = 185) who completed the Dissociative Experiences Scale and the Random Number Generation Task. We found that minor disruptions in executive functioning were related to a subclass of dissociative experiences, notably dissociative amnesia and the Dissociative Experiences Scale Taxon. However, the two other subscales of the Dissociative Experiences Scale, measuring depersonalization and absorption, were unrelated to executive functioning. Our findings suggest that a failure to inhibit previous responses might contribute to the pathological memory manifestations of dissociation.

[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.

Deuterium kinetic isotope effects on the dissociation of a protein-fatty acid complex in the gas phase.

PubMed

Liu, Lan; Michelsen, Klaus; Kitova, Elena N; Schnier, Paul D; Brown, Alex; Klassen, John S

2012-04-04

Deuterium kinetic isotope effects (KIEs) are reported for the first time for the dissociation of a protein-ligand complex in the gas phase. Temperature-dependent rate constants were measured for the loss of neutral ligand from the deprotonated ions of the 1:1 complex of bovine β-lactoglobulin (Lg) and palmitic acid (PA), (Lg + PA)(n-) → Lg(n-) + PA, at the 6- and 7- charge states. At 25 °C, partial or complete deuteration of the acyl chain of PA results in a measurable inverse KIE for both charge states. The magnitude of the KIEs is temperature dependent, and Arrhenius analysis of the rate constants reveals that deuteration of PA results in a decrease in activation energy. In contrast, there is no measurable deuterium KIE for the dissociation of the (Lg + PA) complex in aqueous solution at pH 8. Deuterium KIEs were calculated using conventional transition-state theory with an assumption of a late dissociative transition state (TS), in which the ligand is free of the binding pocket. The vibrational frequencies of deuterated and non-deuterated PA in the gas phase and in various solvents (n-hexane, 1-chlorohexane, acetone, and water) were established computationally. The KIEs calculated from the corresponding differences in zero-point energies account qualitatively for the observation of an inverse KIE but do not account for the magnitude of the KIEs nor their temperature dependence. It is proposed that the dissociation of the (Lg + PA) complex in aqueous solution also proceeds through a late TS in which the acyl chain is extensively hydrated such that there is no significant differential change in the vibrational frequencies along the reaction coordinate and, consequently, no significant KIE.

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)

Photolabeling of tonoplast from sugar beet cell suspensions by [h]5-(N-methyl-N-isobutyl)-amiloride, an inhibitor of the vacuolar na/h antiport.

PubMed

Barkla, B J; Charuk, J H; Cragoe, E J; Blumwald, E

1990-07-01

The effects of 5-(N-methyl-N-isobutyl)-amiloride (MIA), an amiloride analog, was tested on the Na(+)/H(+) antiport activity of intact vacuoles and tonoplast vesicles isolated from sugar beet (Beta vulgaris L.) cell suspension cultures. MIA inhibited Na(+)/H(+) exchange in a competitive manner with a K(i) of 2.5 and 5.9 micromolar for DeltapH-dependent (22)Na(+) influx in tonoplast vesicles and Na(+)-dependent H(+) efflux in intact vacuoles, respectively. Scatchard analysis of the binding of [(3)H]MIA to tonoplast membranes revealed a high affinity binding component with a K(d) of 1.3 micromolar. The close relationship between the dissociation constant value obtained and the constants of inhibition for MIA obtained by fluorescence quenching and isotope exchange suggests that the high affinity component represents a class of sites associated with the tonoplast Na(+)/H(+) antiport. Photolabeling of the tonoplast with [(3)H]MIA revealed two sets of polypeptides with a different affinity to amiloride and its analog.

Engineering an antibody with picomolar affinity to DOTA chelates of multiple radionuclides for pretargeted radioimmunotherapy and imaging

PubMed Central

Orcutt, Kelly Davis; Slusarczyk, Adrian L; Cieslewicz, Maryelise; Ruiz-Yi, Benjamin; Bhushan, Kumar R; Frangioni, John V; Wittrup, K Dane

2014-01-01

Introduction In pretargeted radioimmunotherapy (PRIT), a bifunctional antibody is administered and allowed to pre-localize to tumor cells. Subsequently, a chelated radionuclide is administered and captured by cell-bound antibody while unbound hapten clears rapidly from the body. We aim to engineer high-affinity binders to DOTA chelates for use in PRIT applications. Methods We mathematically modeled antibody and hapten pharmacokinetics to analyze hapten tumor retention as a function of hapten binding affinity. Motivated by model predictions, we used directed evolution and yeast surface display to affinity mature the 2D12.5 antibody to 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA), reformatted as a single chain variable fragment (scFv). Results Modeling predicts that for high antigen density and saturating bsAb dose, a hapten binding affinity of 100 picomolar (pM) is needed for near-maximal hapten retention. We affinity matured 2D12.5 with an initial binding constant of about 10 nanomolar (nM) to DOTA-yttrium chelates. Affinity maturation resulted in a 1000-fold affinity improvement to biotinylated DOTA-yttrium, yielding an 8.2 ± 1.9 picomolar binder. The high-affinity scFv binds DOTA complexes of lutetium and gadolinium with similar picomolar affinity and indium chelates with low nanomolar affinity. When engineered into a bispecific antibody construct targeting carcinoembryonic antigen (CEA), pretargeted high-affinity scFv results in significantly higher tumor retention of a 111In-DOTA hapten compared to pretargeted wild-type scFv in a xenograft mouse model. Conclusions We have engineered a versatile, high-affinity DOTA-chelate-binding scFv. We anticipate it will prove useful in developing pretargeted imaging and therapy protocols to exploit the potential of a variety of radiometals. PMID:21315278

Dissociative symptomatology in cancer patients.

PubMed

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

2015-01-01

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. 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. 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. These findings allow us to hypothesize a specific psychological reaction which may be ascribed to the traumatic spectrum within the context of cancer

Proton dissociation properties of arylphosphonates: Determination of accurate Hammett equation parameters.

PubMed

Dargó, Gergő; Bölcskei, Adrienn; Grün, Alajos; Béni, Szabolcs; Szántó, Zoltán; Lopata, Antal; Keglevich, György; Balogh, György T

2017-09-05

Determination of the proton dissociation constants of several arylphosphonic acid derivatives was carried out to investigate the accuracy of the Hammett equations available for this family of compounds. For the measurement of the pK a values modern, accurate methods, such as the differential potentiometric titration and NMR-pH titration were used. We found our results significantly different from the pK a values reported before (pK a1 : MAE = 0.16 pK a2 : MAE=0.59). Based on our recently measured pK a values, refined Hammett equations were determined that might be used for predicting highly accurate ionization constants of newly synthesized compounds (pK a1 =1.70-0.894σ, pK a2 =6.92-0.934σ). Copyright © 2017 Elsevier B.V. All rights reserved.

Empirical testing of criteria for dissociative schizophrenia.

PubMed

Laferrière-Simard, Marie-Christine; Lecomte, Tania; Ahoundova, Lola

2014-01-01

This study examined the validity of dissociative schizophrenia diagnostic criteria. In the first phase, 50 participants with a psychotic disorder were administered the Dissociative Experiences Scale and the Childhood Trauma Questionnaire to identify those with dissociative characteristics. In the second phase, we selected those who had a score of 15 or above on the Dissociative Experiences Scale. Fifteen of these participants were evaluated thoroughly with the Structured Clinical Interview for DSM-IV Axis I, Structured Clinical Interview for DSM-IV Axis II, and Structured Clinical Interview for DSM-IV Dissociative Disorders to determine whether they met the criteria for dissociative schizophrenia and to generate a clinical description. Our results indicated that 24% of the individuals we tested met these criteria. We propose making mandatory 1 of the 3 dissociative symptoms of the criteria to eliminate people with only nonspecific symptoms (e.g., extensive comorbidity). According to this modified criterion, 14% of our sample would receive a diagnosis of dissociative schizophrenia. However, a more comprehensive look at the clinical picture begs the question of whether dissociative schizophrenia is truly present in every person meeting the criteria. We discuss the relevance of creating a new schizophrenia subtype and offer recommendations for clinicians.

Comparing the symptoms and mechanisms of "dissociation" in dissociative identity disorder and borderline personality disorder.

PubMed

Laddis, Andreas; Dell, Paul F; Korzekwa, Marilyn

2017-01-01

A total of 75 patients were diagnosed with the Structured Clinical Interview for DSM-IV Dissociative Disorders-Revised as having dissociative identity disorder (DID), and 100 patients were diagnosed with the Structured Interview for DSM-IV Personality as having borderline personality disorder (BPD). Both groups were administered the Multidimensional Inventory of Dissociation (MID). DID patients had significantly higher MID scores than BPD patients, different distributions of MID scores, and different MID subscale profiles in 3 ranges of MID scores (0-15, 15-30, 30-45). The core MID symptoms-exhibited at all ranges of MID scores-for DID patients (the presence of alters, identity confusion, and memory problems) and BPD patients (flashbacks, identity confusion, and memory problems) were ostensibly similar but were considered to be mostly produced by different underlying processes. Multiple regression analyses showed that the core MID symptoms of DID patients had different predictors than did the core MID symptoms of BPD patients. Alter identities seemed to generate most-but not all-dissociative phenomena in DID patients, whereas only the 24% highest scoring BPD patients (MID ≥45) seemed to manifest alter-driven dissociative experiences. Most BPD dissociative experiences appeared to be due to 5 other mechanisms: (a) BPD-specific, stress-driven, rapid shifts of self-state; (b and c) nondefensive disruptions of the framework of perceptual organization with or without an accompanying BPD-specific, dissociation-like disintegration of affective/neurocognitive functioning; (d) a defensive distancing or detachment from distress (i.e., simple depersonalization); and (e) Allen, Console, and Lewis's (1999) severe absorptive detachment.

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.

[Dissociative phenomena in a sample of outpatients].

PubMed

Cantone, Daniela; Sperandeo, Raffaele; Maldonato, Mauro Nelson; Cozzolino, Pasquale; Perris, Francesco

2012-01-01

The study describes the frequency and the quality of dissociative phenomena and their relationship with axis I disorders and the psychopathological severity in outpatients. The sample (N=383) was subjected to MINI diagnostic interview and self-assessment scales DES and SCL-90. The data were analysed using SPSS. The 11,0% of subjects has a score ≥20 on DES. The 5,2% has no dissociative symptoms. The absorption images is the most frequent dissociative phenomenon, the less common is the dissociation amnesia. A relationship between dissociative phenomena and conditions unemployment, marital separation and single parties and an inverse relationship with age founded. Dissociative phenomena are more frequent in participants who have been diagnosed at least one axis I disorder and their severity is positively correlated with the number of diagnosed diseases and scores to the General Symptomatic Index. Our results point towards the existence of three types of dissociative experiences. The first type, represented by the factor absorption/imaginative involvement, is expressed along a continuum from normal to pathological; a second type, represented by the factor depersonalization/derealization, occurs in a significantly more intense and specific among subjects with axis I disorders; the latest manifestation dissociative, described by the dissociation amnesia, seems to have a predominantly typological feature that qualifies it as an experience not commonly distributed in the general population. The identifying of dissociative symptoms is necessary for the psychopathologic evaluation and to improve the effectiveness of treatment programs.

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

DOE Office of Scientific and Technical Information (OSTI.GOV)

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 andmore » 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.« less

Partition functions and equilibrium constants for diatomic molecules and atoms of astrophysical interest

NASA Astrophysics Data System (ADS)

Barklem, P. S.; Collet, R.

2016-04-01

Partition functions and dissociation equilibrium constants are presented for 291 diatomic molecules for temperatures in the range from near absolute zero to 10 000 K, thus providing data for many diatomic molecules of astrophysical interest at low temperature. The calculations are based on molecular spectroscopic data from the book of Huber & Herzberg (1979, Constants of Diatomic Molecules) with significant improvements from the literature, especially updated data for ground states of many of the most important molecules by Irikura (2007, J. Phys. Chem. Ref. Data, 36, 389). Dissociation energies are collated from compilations of experimental and theoretical values. Partition functions for 284 species of atoms for all elements from H to U are also presented based on data collected at NIST. The calculated data are expected to be useful for modelling a range of low density astrophysical environments, especially star-forming regions, protoplanetary disks, the interstellar medium, and planetary and cool stellar atmospheres. The input data, which will be made available electronically, also provides a possible foundation for future improvement by the community. Full Tables 1-8 are only available at the CDS via anonymous ftp to http://cdsarc.u-strasbg.fr (ftp://130.79.128.5) or via http://cdsarc.u-strasbg.fr/viz-bin/qcat?J/A+A/588/A96

MDMA, cannabis, and cocaine produce acute dissociative symptoms.

PubMed

van Heugten-Van der Kloet, Dalena; Giesbrecht, Timo; van Wel, Janelle; Bosker, Wendy M; Kuypers, Kim P C; Theunissen, Eef L; Spronk, Desirée B; Jan Verkes, Robbert; Merckelbach, Harald; Ramaekers, Johannes G

2015-08-30

Some drugs of abuse may produce dissociative symptoms, but this aspect has been understudied. We explored the dissociative potential of three recreational drugs (3,4-methylenedioxymethamphetamine (MDMA), cannabis, and cocaine) during intoxication and compared their effects to literature reports of dissociative states in various samples. Two placebo-controlled studies were conducted. In Study 1 (N=16), participants received single doses of 25, 50, and 100 mg of MDMA, and placebo. In Study 2 (N=21), cannabis (THC 300 µg/kg), cocaine (HCl 300 mg), and placebo were administered. Dissociative symptoms as measured with the Clinician-Administered Dissociative States Scale (CADSS) significantly increased under the influence of MDMA and cannabis. To a lesser extent, this was also true for cocaine. Dissociative symptoms following MDMA and cannabis largely exceeded those observed in schizophrenia patients, were comparable with those observed in Special Forces soldiers undergoing survival training, but were lower compared with ketamine-induced dissociation. Cocaine produced dissociative symptoms that were comparable with those observed in schizophrenia patients, but markedly less than those in Special Forces soldiers and ketamine users. Thus, MDMA and cannabis can produce dissociative symptoms that resemble dissociative pathology. The study of drug induced dissociation is important, because it may shed light on the mechanisms involved in dissociative psychopathology. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Dissociation and purification of the endogenous membrane-bound Vo complex from Pichia pastoris.

PubMed

Li, Sumei; Hong, Tao; Wang, Kun; Lu, Yinghong; Zhou, Min

2017-10-01

Most proteins occur and function in complexes rather than as isolated entities in membranes. In most cases macromolecules with multiple subunits are purified from endogenous sources. In this study, an endogenous membrane-protein complex was obtained from Pichia pastoris, which can be grown at high densities to significantly improve the membrane protein yield. We successfully isolated the membrane-bound Vo complex of V-ATPase from P. pastoris using a fusion FLAG tag attached to the C-terminus of subunit a to generate the vph-tag strain, which was used for dissociation and purification. After FLAG affinity and size exclusion chromatography purification, the production quantity and purity of the membrane-bound Vo complex was 20 μg l -1 and >98%, respectively. The subunits of the endogenous membrane-bound Vo complex observed in P. pastoris were similar to those obtained from S. cerevisiae, as demonstrated by liquid chromatography-tandem mass spectrometry (LC-MS-MS). Therefore, successful dissociation and purification of the membrane-bound Vo complex at a high purity and sufficient quantity was achieved via a rapid and simple procedure that can be used to obtain the endogenous membrane-protein complexes from P. pastoris. Copyright © 2017 Elsevier Inc. All rights reserved.

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.

The role of equilibrium and kinetic properties in the dissociation of Gd[DTPA-bis(methylamide)] (Omniscan) at near to physiological conditions.

PubMed

Baranyai, Zsolt; Brücher, Ernő; Uggeri, Fulvio; Maiocchi, Alessandro; Tóth, Imre; Andrási, Melinda; Gáspár, Attila; Zékány, László; Aime, Silvio

2015-03-16

[Gd(DTPA-BMA)] is the principal constituent of Omniscan, a magnetic resonance imaging (MRI) contrast agent. In body fluids, endogenous ions (Zn(2+), Cu(2+), and Ca(2+)) may displace the Gd(3+). To assess the extent of displacement at equilibrium, the stability constants of DTPA-BMA(3-) complexes of Gd(3+), Ca(2+), Zn(2+), and Cu(2+) have been determined at 37 °C in 0.15 M NaCl. The order of these stability constants is as follows: GdL≈CuL>ZnL≫CaL. Applying a simplified blood plasma model, the extent of dissociation of Omniscan (0.35 mM [Gd(DTPA-BMA)]) was found to be 17% by the formation of Gd(PO4), [Zn(DTPA-BMA)](-) (2.4%), [Cu(DTPA-BMA)](-) (0.2%), and [Ca(DTPA-BMA)](-) (17.7%). By capillary electrophoresis, the formation of [Ca(DTPA-BMA)](-) has been detected in human serum spiked with [Gd(DTPA-BMA)] (2.0 mM) at pH 7.4. Transmetallation reactions between [Gd(DTPA-BMA)] and Cu(2+) at 37 °C in the presence of citrate, phosphate, and bicarbonate ions occur by dissociation of the complex assisted by the endogenous ligands. At physiological concentrations of citrate, phosphate, and bicarbonate ions, the half-life of dissociation of [Gd(DTPA-BMA)] was calculated to be 9.3 h at pH 7.4. Considering the rates of distribution and dissociation of [Gd(DTPA-BMA)] in the extracellular space of the body, an open two-compartment model has been developed, which allows prediction of the extent of dissociation of the Gd(III) complex in body fluids depending on the rate of elimination of the contrast agent. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Gene Therapy Vectors with Enhanced Transfection Based on Hydrogels Modified with Affinity Peptides

PubMed Central

Shepard, Jaclyn A.; Wesson, Paul J.; Wang, Christine E.; Stevans, Alyson C.; Holland, Samantha J.; Shikanov, Ariella; Grzybowski, Bartosz A.; Shea, Lonnie D.

2011-01-01

Regenerative strategies for damaged tissue aim to present biochemical cues that recruit and direct progenitor cell migration and differentiation. Hydrogels capable of localized gene delivery are being developed to provide a support for tissue growth, and as a versatile method to induce the expression of inductive proteins; however, the duration, level, and localization of expression isoften insufficient for regeneration. We thus investigated the modification of hydrogels with affinity peptides to enhance vector retention and increase transfection within the matrix. PEG hydrogels were modified with lysine-based repeats (K4, K8), which retained approximately 25% more vector than control peptides. Transfection increased 5- to 15-fold with K8 and K4 respectively, over the RDG control peptide. K8- and K4-modified hydrogels bound similar quantities of vector, yet the vector dissociation rate was reduced for K8, suggesting excessive binding that limited transfection. These hydrogels were subsequently applied to an in vitro co-culture model to induce NGF expression and promote neurite outgrowth. K4-modified hydrogels promoted maximal neurite outgrowth, likely due to retention of both the vector and the NGF. Thus, hydrogels modified with affinity peptides enhanced vector retention and increased gene delivery, and these hydrogels may provide a versatile scaffold for numerous regenerative medicine applications. PMID:21514659

Theoretical and Experimental Investigation of Heat Conduction in Air, Including Effects of Oxygen Dissociation

NASA Technical Reports Server (NTRS)

Hansen, C. Frederick; Early, Richard A.; Alzofon, Frederick E.; Witteborn, Fred C.

1959-01-01

Solutions are presented for the conduction of beat through a semi-infinite gas medium having a uniform initial temperature and a constant boundary temperature. The coefficients of thermal conductivity and diffusivity are treated as variables, and the solutions are extended to the case of air at temperatures where oxygen dissociation occurs. These solutions are used together with shock-tube measurements to evaluate the integral of thermal conductivity for air as a function of temperature.

Simple dissociations for a higher-powered neuropsychology.

PubMed

McIntosh, Robert D

2018-06-01

Dissociations in cognitive neuropsychology are often investigated at the level of the single-case, and formal criteria exist for the detection of dissociations, and their sub-classification into 'classical' and 'strong' types. These criteria require a patient to show a frank deficit on one task (for a classical dissociation) or both tasks (for a strong dissociation), and a significantly extreme difference between tasks. I propose that only the significant between-task difference is logically necessary, and that if this simple criterion is met, the patient should be said to show a dissociation. Using Monte Carlo simulations, I show that this simplification increases the power to detect dissociations across a range of practically-relevant conditions, whilst retaining excellent control over Type I error. Additional testing for frank deficits on each task provides further qualifying information, but using these test outcomes to categorise dissociations as classical or strong may be too uncertain to guide theoretical inferences reliably. I suggest that we might instead characterise the strength of the dissociation using a continuous index, such as the effect size of the between-task difference. Copyright © 2018 Elsevier Ltd. All rights reserved.

Selecting for Fast Protein-Protein Association As Demonstrated on a Random TEM1 Yeast Library Binding BLIP.

PubMed

Cohen-Khait, Ruth; Schreiber, Gideon

2018-04-27

Protein-protein interactions mediate the vast majority of cellular processes. Though protein interactions obey basic chemical principles also within the cell, the in vivo physiological environment may not allow for equilibrium to be reached. Thus, in vitro measured thermodynamic affinity may not provide a complete picture of protein interactions in the biological context. Binding kinetics composed of the association and dissociation rate constants are relevant and important in the cell. Therefore, changes in protein-protein interaction kinetics have a significant impact on the in vivo activity of the proteins. The common protocol for the selection of tighter binders from a mutant library selects for protein complexes with slower dissociation rate constants. Here we describe a method to specifically select for variants with faster association rate constants by using pre-equilibrium selection, starting from a large random library. Toward this end, we refine the selection conditions of a TEM1-β-lactamase library against its natural nanomolar affinity binder β-lactamase inhibitor protein (BLIP). The optimal selection conditions depend on the ligand concentration and on the incubation time. In addition, we show that a second sort of the library helps to separate signal from noise, resulting in a higher percent of faster binders in the selected library. Fast associating protein variants are of particular interest for drug development and other biotechnological applications.

On the dissociation and conformation of gas-phase methonium ions.

PubMed

Gross, Deborah S; Williams, Evan R

1996-12-20

The dissociation pathways of both doubly and singly charged methonium ions, (CH(3))(3)N(+) -(CH(2))(n)-(+)N(CH(3))(3)·X(-) (n = 6,10; X = Br, I, and OAc), are measured using blackbody infrared radiative dissociation (BIRD) and SORI-CAD in a Fourier transform mass spectrometer. SORI-CAD of the doubly charged decamethonium ions results primarily in the formation of even electron ions by hydrogen rearrangements. In contrast, homolytic bond cleavage to form two odd electron ions is highly favored in the hexamethonium ion, presumably due to increased Coulomb repulsion in this ion. For BIRD of the singly charged salts, ions are mass selected and dissociated by heating the vacuum chamber to elevated temperatures. Under the low pressure conditions of our experiment, energy is transferred from the chamber walls to the ions by the absorption of blackbody radiation. From the temperature dependence of the unimolecular rate constants for dissociation, Arrhenius activation energies in the zero-pressure limit are obtained. The primary dissociation pathways correspond to counterion substitution reactions which result in loss of N(CH(3))(3) and CH(3)X. For hexamethonium and decamethonium with X = Br or I, the branching ratios for these pathways differ dramatically; the ratio of loss of N(CH(3))(3) and CH(3)Br is 3.8 and 0.4 for hexamethonium and decamethonium bromide, respectively. The hexamethonium acetate salt has a branching ratio of 0.1. The Arrhenius activation energies for hexamethonium (Br or I) and decamethonium (Br or I) are 0.9 and 1.0 eV, respectively. This value for hexamethonium acetate is 0.6 eV. Molecular dynamics simulations and Monte Carlo conformation searching are used to obtain the lowest energy structures of hexamethonium and decamethonium bromide. These calculations indicate that the methonium ion folds around the counterion to form a cyclic salt-bridge structure in which both quaternary nitrogens interact with the oppositely charged counterion. The

On the dissociation and conformation of gas-phase methonium ions

PubMed Central

Gross, Deborah S.

2005-01-01

The dissociation pathways of both doubly and singly charged methonium ions, (CH3)3N+ –(CH2)n–+N(CH3)3·X− (n = 6,10; X = Br, I, and OAc), are measured using blackbody infrared radiative dissociation (BIRD) and SORI–CAD in a Fourier transform mass spectrometer. SORI–CAD of the doubly charged decamethonium ions results primarily in the formation of even electron ions by hydrogen rearrangements. In contrast, homolytic bond cleavage to form two odd electron ions is highly favored in the hexamethonium ion, presumably due to increased Coulomb repulsion in this ion. For BIRD of the singly charged salts, ions are mass selected and dissociated by heating the vacuum chamber to elevated temperatures. Under the low pressure conditions of our experiment, energy is transferred from the chamber walls to the ions by the absorption of blackbody radiation. From the temperature dependence of the unimolecular rate constants for dissociation, Arrhenius activation energies in the zero-pressure limit are obtained. The primary dissociation pathways correspond to counterion substitution reactions which result in loss of N(CH3)3 and CH3X. For hexamethonium and decamethonium with X = Br or I, the branching ratios for these pathways differ dramatically; the ratio of loss of N(CH3)3 and CH3Br is 3.8 and 0.4 for hexamethonium and decamethonium bromide, respectively. The hexamethonium acetate salt has a branching ratio of 0.1. The Arrhenius activation energies for hexamethonium (Br or I) and decamethonium (Br or I) are 0.9 and 1.0 eV, respectively. This value for hexamethonium acetate is 0.6 eV. Molecular dynamics simulations and Monte Carlo conformation searching are used to obtain the lowest energy structures of hexamethonium and decamethonium bromide. These calculations indicate that the methonium ion folds around the counterion to form a cyclic salt-bridge structure in which both quaternary nitrogens interact with the oppositely charged counterion. The significantly different

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.

Adjoint affine fusion and tadpoles

DOE Office of Scientific and Technical Information (OSTI.GOV)

Urichuk, Andrew, E-mail: andrew.urichuk@uleth.ca; Walton, Mark A., E-mail: walton@uleth.ca; International School for Advanced Studies

2016-06-15

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 writtenmore » 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.« less

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(+).

Pulsed IR Heating Studies of Single-Molecule DNA Duplex Dissociation Kinetics and Thermodynamics

PubMed Central

Holmstrom, Erik D.; Dupuis, Nicholas F.; Nesbitt, David J.

2014-01-01

Single-molecule fluorescence spectroscopy is a powerful technique that makes it possible to observe the conformational dynamics associated with biomolecular processes. The addition of precise temperature control to these experiments can yield valuable thermodynamic information about equilibrium and kinetic rate constants. To accomplish this, we have developed a microscopy technique based on infrared laser overtone/combination band absorption to heat small (≈10−11 liter) volumes of water. Detailed experimental characterization of this technique reveals three major advantages over conventional stage heating methods: 1), a larger range of steady-state temperatures (20–100°C); 2), substantially superior spatial (≤20 μm) control; and 3), substantially superior temporal (≈1 ms) control. The flexibility and breadth of this spatial and temporally resolved laser-heating approach is demonstrated in single-molecule fluorescence assays designed to probe the dissociation of a 21 bp DNA duplex. These studies are used to support a kinetic model based on nucleic acid end fraying that describes dissociation for both short (<10 bp) and long (>10 bp) DNA duplexes. These measurements have been extended to explore temperature-dependent kinetics for the 21 bp construct, which permit determination of single-molecule activation enthalpies and entropies for DNA duplex dissociation. PMID:24411254

Open scapulothoracic dissociation.

PubMed

Fischer, P J; Kent, R B

2001-04-01

Scapulothoracic dissociation refers to the traumatic separation of the shoulder from the chest wall. This most commonly occurs as a closed injury. We present a case of open scapulothoracic dissociation and emphasize clinical features unique to this injury. In both closed and open scapulothoracic dissociation, the force necessary to shear the scapula from its thoracic attachments results in vascular disruption and neurologic injury to the upper extremity. As a consequence, patients have a pulseless, flail upper extremity with a significant chest wall hematoma (closed) or active bleeding (open). The first priority is to resuscitate and address life-threatening injuries. If the patient has active bleeding, immediate vascular control to prevent exsanguination is essential. Patients with ischemia and an incomplete injury or unreliable neurologic examination need revascularization. Outcome is based on the extent of brachial plexus or cervical nerve root avulsion. Patients with loss of neurologic function ultimately benefit from amputation at the appropriate level.

Activation of muscle nicotinic acetylcholine receptor channels by nicotinic and muscarinic agonists

PubMed Central

Akk, Gustav; Auerbach, Anthony

1999-01-01

The dose-response parameters of recombinant mouse adult neuromuscular acetylcholine receptor channels (nAChR) activated by carbamylcholine, nicotine, muscarine and oxotremorine were measured. Rate constants for agonist association and dissociation, and channel opening and closing, were estimated from single-channel kinetic analysis.The dissociation equilibrium constants were (mM): ACh (0.16)constants (opening/closing) were: ACh (45)>carbamylcholine (5.1)>oxotremorine M (0.6)>nicotine (0.5)>muscarine (0.15).Rat neuronal α4β2 nAChR can be activated by all of the agonists. However, detailed kinetic analysis was impossible because the recordings lacked clusters representing the activity of a single receptor complex. Thus, the number of channels in the patch was unknown and the activation rate constants could not be determined.Considering both receptor affinity and agonist efficacy, muscarine and oxotremorine are significant agonists of muscle-type nAChR. The results are discussed in terms of structure-function relationships at the nAChR transmitter binding site. PMID:10602325

Herpes Simplex Virus Processivity Factor UL42 Imparts Increased DNA-Binding Specificity to the Viral DNA Polymerase and Decreased Dissociation from Primer-Template without Reducing the Elongation Rate

PubMed Central

Weisshart, Klaus; Chow, Connie S.; Coen, Donald M.

1999-01-01

Herpes simplex virus DNA polymerase consists of a catalytic subunit, Pol, and a processivity subunit, UL42, that, unlike other established processivity factors, binds DNA directly. We used gel retardation and filter-binding assays to investigate how UL42 affects the polymerase-DNA interaction. The Pol/UL42 heterodimer bound more tightly to DNA in a primer-template configuration than to single-stranded DNA (ssDNA), while Pol alone bound more tightly to ssDNA than to DNA in a primer-template configuration. The affinity of Pol/UL42 for ssDNA was reduced severalfold relative to that of Pol, while the affinity of Pol/UL42 for primer-template DNA was increased ∼15-fold relative to that of Pol. The affinity of Pol/UL42 for circular double-stranded DNA (dsDNA) was reduced drastically relative to that of UL42, but the affinity of Pol/UL42 for short primer-templates was increased modestly relative to that of UL42. Pol/UL42 associated with primer-template DNA ∼2-fold faster than did Pol and dissociated ∼10-fold more slowly, resulting in a half-life of 2 h and a subnanomolar Kd. Despite such stable binding, rapid-quench analysis revealed that the rates of elongation of Pol/UL42 and Pol were essentially the same, ∼30 nucleotides/s. Taken together, these studies indicate that (i) Pol/UL42 is more likely than its subunits to associate with DNA in a primer-template configuration rather than nonspecifically to either ssDNA or dsDNA, and (ii) UL42 reduces the rate of dissociation from primer-template DNA but not the rate of elongation. Two models of polymerase-DNA interactions during replication that may explain these findings are presented. PMID:9847307

Dissociative disorders and dissociative symptoms among veterans of the Iraq-Iran war suffering from chronic post-traumatic disorder.

PubMed

Nejad, Alireza G; Farahati, Hamid

2007-10-01

To evaluate any dissociative disorders in patients with post traumatic stress disorder (PTSD), and find any relationship between dissociation and related variables in these patients. A case controlled study carried out among 130 male patients with PTSD, and 130 matched individuals from the normal population. The study was carried out between January and September 2005, at Beheshti Psychiatric Hospital, Kerman, Iran, Demographic data and history of childhood physical abuse and self-harm in both groups were recorded. The severity of PTSD was measured by Davidson PTSD scale in the study group. Dissociative symptoms were evaluated with the dissociative experience scale (DES). For evaluating dissociative disorders, the dissociative disorder interview schedule (DDIS) was used. Data were analyzed by SPSS software. The mean age (+/- SD) of veterans was 41.46 (+/-5.09) years, and the mean age of going to the combat area was 29.4 (+/-5.015). The mean duration of involvement in combat was 27.8 (+/-23.44) months. The mean DES score in the study group was 26.01 (+/-12.31) and was 9.58 (+/-7.23) in the control group (F=1.171, p<0.0001). The mean score (+/-SD) of Davidson PTSD scale was 54+/-96 in the study group. The most prevalent dissociative disorder in the study group was psychogenic amnesia. Many veterans with the impression of chronic PTSD may have an additional diagnosis of dissociative disorders.

Dissociative Disorders Among Chinese Inpatients Diagnosed With Schizophrenia

PubMed Central

Yu, Junhan; Ross, Colin A.; Keyes, Benjamin B.; Li, Ying; Dai, Yunfei; Zhang, Tianhong; Wang, Lanlan; Fan, Qing; Xiao, Zeping

2010-01-01

The purpose of the study was to assess the prevalence of dissociative disorders in a sample of Chinese psychiatric inpatients. Participants in the study consisted of 569 consecutively admitted inpatients at Shanghai Mental Health Center, China, of whom 84.9% had a clinical diagnosis of schizophrenia based on the Chinese Classification and Diagnostic Criteria for Mental Disorders, Version 3 (CCMD-3). All participants completed a self-report measure of dissociation, the Dissociative Experiences Scale (DES) and none had a prior diagnosis of a dissociative disorder. Ninety-six randomly selected participants were interviewed with a structured interview, the Dissociative Disorders Interview Schedule (DDIS) and a clinical interview. These 96 patients did not differ significantly from the 473 patients who were not interviewed on any demographic measures or on the self-report measure dissociation. A total of 28 (15.3%, after weighting of the data) patients received a clinical diagnosis of a dissociative disorder based on DSM-IV-TR criteria. Dissociative identity disorder was diagnosed in 2 (0.53%, after weighting) patients. Compared to the patients without a dissociative disorder, patients with dissociative disorders were significantly more likely to report childhood abuse (57.1% versus 22.1%), but the two groups did not differ significantly on any demographic measures. Dissociative disorders were readily identified in an inpatient psychiatric population in China. PMID:20603768

Dissociative disorders among Chinese inpatients diagnosed with schizophrenia.

PubMed

Yu, Junhan; Ross, Colin A; Keyes, Benjamin B; Li, Ying; Dai, Yunfei; Zhang, Tianhong; Wang, Lanlan; Fan, Qing; Xiao, Zeping

2010-01-01

The purpose of this study was to assess the prevalence of dissociative disorders in a sample of Chinese psychiatric inpatients. Participants in the study were 569 consecutively admitted inpatients at Shanghai Mental Health Center, China, of whom 84.9% had a clinical diagnosis of schizophrenia based on the Chinese Classification and Diagnostic Criteria of Mental Disorders, Version 3. All participants completed a self-report measure of dissociation (the Dissociative Experiences Scale), and none had a prior diagnosis of a dissociative disorder. A total of 96 randomly selected participants were interviewed with a structured interview (the Dissociative Disorders Interview Schedule) and a clinical interview. These 96 patients did not differ significantly from the 473 patients who were not interviewed on any demographic measures or who did not complete the self-report dissociation measure. A total of 28 patients (15.3%, after weighting of the data) received a clinical diagnosis of a dissociative disorder based on Diagnostic and Statistical Manual of Mental Disorders (4th ed., text rev.) criteria. Dissociative identity disorder was diagnosed in 2 patients (0.53%, after weighting). Compared to the patients without a dissociative disorder, patients with dissociative disorders were significantly more likely to report childhood abuse (57.1% vs. 22.1%), but the 2 groups did not differ significantly on any demographic measures. Dissociative disorders were readily identified in an inpatient psychiatric population in China.

Non-affine deformations in polymer hydrogels

PubMed Central

Wen, Qi; Basu, Anindita; Janmey, Paul A.; Yodh, A. G.

2012-01-01

Most theories of soft matter elasticity assume that the local strain in a sample after deformation is identical everywhere and equal to the macroscopic strain, or equivalently that the deformation is affine. We discuss the elasticity of hydrogels of crosslinked polymers with special attention to affine and non-affine theories of elasticity. Experimental procedures to measure non-affine deformations are also described. Entropic theories, which account for gel elasticity based on stretching out individual polymer chains, predict affine deformations. In contrast, simulations of network deformation that result in bending of the stiff constituent filaments generally predict non-affine behavior. Results from experiments show significant non-affine deformation in hydrogels even when they are formed by flexible polymers for which bending would appear to be negligible compared to stretching. However, this finding is not necessarily an experimental proof of the non-affine model for elasticity. We emphasize the insights gained from experiments using confocal rheoscope and show that, in addition to filament bending, sample micro-inhomogeneity can be a significant alternative source of non-affine deformation. PMID:23002395

Dancing with the Muses: dissociation and flow.

PubMed

Thomson, Paula; Jaque, S Victoria

2012-01-01

This study investigated dissociative psychological processes and flow (dispositional and state) in a group of professional and pre-professional dancers (n=74). In this study, high scores for global (Mdn=4.14) and autotelic (Mdn=4.50) flow suggest that dancing was inherently integrating and rewarding, although 17.6% of the dancers were identified as possibly having clinical levels of dissociation (Dissociative Experiences Scale-Taxon cutoff score≥20). The results of the multivariate analysis of variance indicated that subjects with high levels of dissociation had significantly lower levels of global flow (p<.05). Stepwise linear regression analyses demonstrated that dispositional flow negatively predicted the dissociative constructs of depersonalization and taxon (p<.05) but did not significantly predict the variance in absorption/imagination (p>.05). As hypothesized, dissociation and flow seem to operate as different mental processes.

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.

In-Vacuum Dissociator for Atomic-Hydrogen Masers

NASA Technical Reports Server (NTRS)

Vessot, R. F.

1987-01-01

Thermal control and vacuum sealing achieved while contamination avoided. Simple, relatively inexpensive molecular-hydrogen dissociator for atomic-hydrogen masers used on Earth or in vacuum of space. No air cooling required, and absence of elastomeric O-ring seals prevents contamination. In-vacuum dissociator for atomic hydrogen masers, hydrogen gas in glass dissociator dissociated by radio-frequency signal transmitted from surrounding 3-turn coil. Heat in glass conducted away by contacting metal surfaces.

Measuring Binding Affinity of Protein-Ligand Interaction Using Spectrophotometry: Binding of Neutral Red to Riboflavin-Binding Protein

ERIC Educational Resources Information Center

Chenprakhon, Pirom; Sucharitakul, Jeerus; Panijpan, Bhinyo; Chaiyen, Pimchai

2010-01-01

The dissociation constant, K[subscript d], of the binding of riboflavin-binding protein (RP) with neutral red (NR) can be determined by titrating RP to a fixed concentration of NR. Upon adding RP to the NR solution, the maximum absorption peak of NR shifts to 545 nm from 450 nm for the free NR. The change of the absorption can be used to determine…

Lp-mixed affine surface area

NASA Astrophysics Data System (ADS)

Wang, Weidong; Leng, Gangsong

2007-11-01

According to the three notions of mixed affine surface area, Lp-affine surface area and Lp-mixed affine surface area proposed by Lutwak, in this article, we give the concept of ith Lp-mixed affine surface area such that the first and second notions of Lutwak are its special cases. Further, some Lutwak's results are extended associated with this concept. Besides, applying this concept, we establish an inequality for the volumes and dual quermassintegrals of a class of star bodies.

Calculation of the rate constant for state-selected recombination of H+O2(v) as a function of temperature and pressure

NASA Astrophysics Data System (ADS)

Teitelbaum, Heshel; Caridade, Pedro J. S. B.; Varandas, António J. C.

2004-06-01

Classical trajectory calculations using the MERCURY/VENUS code have been carried out on the H+O2 reactive system using the DMBE-IV potential energy surface. The vibrational quantum number and the temperature were selected over the ranges v=0 to 15, and T=300 to 10 000 K, respectively. All other variables were averaged. Rate constants were determined for the energy transfer process, H+O2(v)-->H+O2(v''), for the bimolecular exchange process, H+O2(v)-->OH(v')+O, and for the dissociative process, H+O2(v)-->H+O+O. The dissociative process appears to be a mere extension of the process of transferring large amounts of energy. State-to-state rate constants are given for the exchange reaction, and they are in reasonable agreement with previous results, while the energy transfer and dissociative rate constants have never been reported previously. The lifetime distributions of the HO2 complex, calculated as a function of v and temperature, were used as a basis for determining the relative contributions of various vibrational states of O2 to the thermal rate coefficients for recombination at various pressures. This novel approach, based on the complex's ability to survive until it collides in a secondary process with an inert gas, is used here for the first time. Complete falloff curves for the recombination of H+O2 are also calculated over a wide range of temperatures and pressures. The combination of the two separate studies results in pressure- and temperature-dependent rate constants for H+O2(v)(+Ar)⇄HO2(+Ar). It is found that, unlike the exchange reaction, vibrational and rotational-translational energy are liabilities in promoting recombination.

The influence of the position of the oxygen dissociation curve on oxygen-dependent functions of the isolated perfused rat liver. III. Studies at different levels of anaemic hypoxia.

PubMed

Bakker, J C; Gortmaker, G C; de Vries-van Rossen, A; Offerijns, F G

1977-03-11

The influence of a 2,3-diphosphoglycerate (2,3-DPG)-induced displacement of the oxygen dissociation curve (O.D.C.) on the isolated perfused rat liver was studied at different levels of anaemic hypoxia. Rat livers were perfused either with fresh or with 2,3-DPG-depleted human erythrocytes at different haematocrit values (from 30% to 2.5%) at constant Po2 of the inflowing perfusate and at constant blood flow rate. The 2,3-DPG-induced difference in oxygen affinity of the red cells did not cause a significant difference in perfusion pressure during the perfusion experiments. Therefore, there is no evidence that 2,3-DPG did alter the vascular resistance of the liver, since blood flow rate could be adusted at equal values. The decrease in oxygen supply brought about by decrease of haematocrit caused a decrease of O2 consumption, of bile flow rate and of venous Po2 and an increase of lactate/pyruvate (L/P) ratio and of beta-hydroxybutyrate/acetoacetate (betaOH/Acac) ratio. There was no influence of a difference in 2,3-DPG content of the erythrocytes on the above-metioned parameters during severe anaemic hypoxia. At moderate anaemic hypoxia the venous Po2 was higher during perfusion with fresh erythrocytes than during perfusion with 2,3-DPG-depleted erythrocytes. Thus, although 2,3-DPG may play a compensatory role during conditions of mild anaemia, no such effects can be observed during conditions of severe hypoxia.

Kinetic studies on strand displacement in de novo designed parallel heterodimeric coiled coils.

PubMed

Groth, Mike C; Rink, W Mathis; Meyer, Nils F; Thomas, Franziska

2018-05-14

Among the protein folding motifs, which are accessible by de novo design, the parallel heterodimeric coiled coil is most frequently used in bioinspired applications and chemical biology in general. This is due to the straightforward sequence-to-structure relationships, which it has in common with all coiled-coil motifs, and the heterospecificity, which allows control of association. Whereas much focus was laid on designing orthogonal coiled coils, systematic studies on controlling association, for instance by strand displacement, are rare. As a contribution to the design of dynamic coiled-coil-based systems, we studied the strand-displacement mechanism in obligate heterodimeric coiled coils to investigate the suitability of the dissociation constants ( K D ) as parameters for the prediction of the outcome of strand-displacement reactions. We use two sets of heterodimeric coiled coils, the previously reported N-A x B y and the newly characterized C-A x B y . Both comprise K D values in the μM to sub-nM regime. Strand displacement is explored by CD titration and a FRET-based kinetic assay and is proved to be an equilibrium reaction with half-lifes from a few seconds up to minutes. We could fit the displacement data by a competitive binding model, giving rate constants and overall affinities of the underlying association and dissociation reactions. The overall affinities correlate well with the ratios of K D values determined by CD-thermal denaturation experiments and, hence, support the dissociative mechanism of strand displacement in heterodimeric coiled coils. From the results of more than 100 different displacement reactions we are able to classify three categories of overall affinities, which allow for easy prediction of the equilibrium of strand displacement in two competing heterodimeric coiled coils.

Kinetic studies on strand displacement in de novo designed parallel heterodimeric coiled coils† †Electronic supplementary information (ESI) available. See DOI: 10.1039/c7sc05342h

PubMed Central

Groth, Mike C.; Rink, W. Mathis; Meyer, Nils F.

2018-01-01

Among the protein folding motifs, which are accessible by de novo design, the parallel heterodimeric coiled coil is most frequently used in bioinspired applications and chemical biology in general. This is due to the straightforward sequence-to-structure relationships, which it has in common with all coiled-coil motifs, and the heterospecificity, which allows control of association. Whereas much focus was laid on designing orthogonal coiled coils, systematic studies on controlling association, for instance by strand displacement, are rare. As a contribution to the design of dynamic coiled-coil-based systems, we studied the strand-displacement mechanism in obligate heterodimeric coiled coils to investigate the suitability of the dissociation constants (KD) as parameters for the prediction of the outcome of strand-displacement reactions. We use two sets of heterodimeric coiled coils, the previously reported N-AxBy and the newly characterized C-AxBy. Both comprise KD values in the μM to sub-nM regime. Strand displacement is explored by CD titration and a FRET-based kinetic assay and is proved to be an equilibrium reaction with half-lifes from a few seconds up to minutes. We could fit the displacement data by a competitive binding model, giving rate constants and overall affinities of the underlying association and dissociation reactions. The overall affinities correlate well with the ratios of KD values determined by CD-thermal denaturation experiments and, hence, support the dissociative mechanism of strand displacement in heterodimeric coiled coils. From the results of more than 100 different displacement reactions we are able to classify three categories of overall affinities, which allow for easy prediction of the equilibrium of strand displacement in two competing heterodimeric coiled coils. PMID:29780562

Characterizing low affinity epibatidine binding to α4β2 nicotinic acetylcholine receptors with ligand depletion and nonspecific binding

PubMed Central

2011-01-01

Background Along with high affinity binding of epibatidine (Kd1≈10 pM) to α4β2 nicotinic acetylcholine receptor (nAChR), low affinity binding of epibatidine (Kd2≈1-10 nM) to an independent binding site has been reported. Studying this low affinity binding is important because it might contribute understanding about the structure and synthesis of α4β2 nAChR. The binding behavior of epibatidine and α4β2 AChR raises a question about interpreting binding data from two independent sites with ligand depletion and nonspecific binding, both of which can affect equilibrium binding of [3H]epibatidine and α4β2 nAChR. If modeled incorrectly, ligand depletion and nonspecific binding lead to inaccurate estimates of binding constants. Fitting total equilibrium binding as a function of total ligand accurately characterizes a single site with ligand depletion and nonspecific binding. The goal of this study was to determine whether this approach is sufficient with two independent high and low affinity sites. Results Computer simulations of binding revealed complexities beyond fitting total binding for characterizing the second, low affinity site of α4β2 nAChR. First, distinguishing low-affinity specific binding from nonspecific binding was a potential problem with saturation data. Varying the maximum concentration of [3H]epibatidine, simultaneously fitting independently measured nonspecific binding, and varying α4β2 nAChR concentration were effective remedies. Second, ligand depletion helped identify the low affinity site when nonspecific binding was significant in saturation or competition data, contrary to a common belief that ligand depletion always is detrimental. Third, measuring nonspecific binding without α4β2 nAChR distinguished better between nonspecific binding and low-affinity specific binding under some circumstances of competitive binding than did presuming nonspecific binding to be residual [3H]epibatidine binding after adding a large concentration of

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.

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. Copyright © 2015 Elsevier Ltd. All rights reserved.

Dissociation: Defining the Concept in Criminal Forensic Psychiatry.

PubMed

Bourget, Dominique; Gagné, Pierre; Wood, Stephen Floyd

2017-06-01

Claims of amnesia and dissociative experiences in association with a violent crime are not uncommon. Research has shown that dissociation is a risk factor for violence and is seen most often in crimes of extreme violence. The subject matter is most relevant to forensic psychiatry. Peritraumatic dissociation for instance, with or without a history of dissociative disorder, is quite frequently reported by offenders presenting for a forensic psychiatric examination. Dissociation or dissociative amnesia for serious offenses can have legal repercussions stemming from their relevance to the legal constructs of fitness to stand trial, criminal responsibility, and diminished capacity. The complexity in forensic psychiatric assessments often lies in the difficulty of connecting clinical symptomatology reported by violent offenders to a specific condition included in the Diagnostic and Statistical Manual of Mental Disorders (DSM). This article provides a review of diagnostic considerations with regard to dissociation across the DSM nomenclature, with a focus on the main clinical constructs related to dissociation. Forensic implications are discussed, along with some guides for the forensic evaluator of offenders presenting with dissociation. © 2017 American Academy of Psychiatry and the Law.

Collision-induced dissociation analysis of negative atmospheric ion adducts in atmospheric pressure corona discharge ionization mass spectrometry.

PubMed

Sekimoto, Kanako; Takayama, Mitsuo

2013-05-01

Collision-induced dissociation (CID) experiments were performed on atmospheric ion adducts [M + R](-) formed between various types of organic compounds M and atmospheric negative ions R(-) [such as O2(-), HCO3(-), COO(-)(COOH), NO2(-), NO3(-), and NO3(-)(HNO3)] in negative-ion mode atmospheric pressure corona discharge ionization (APCDI) mass spectrometry. All of the [M + R](-) adducts were fragmented to form deprotonated analytes [M - H](-) and/or atmospheric ions R(-), whose intensities in the CID spectra were dependent on the proton affinities of the [M - H](-) and R(-) fragments. Precursor ions [M + R](-) for which R(-) have higher proton affinities than [M - H](-) formed [M - H](-) as the dominant product. Furthermore, the CID of the adducts with HCO3(-) and NO3(-)(HNO3) led to other product ions such as [M + HO](-) and NO3(-), respectively. The fragmentation behavior of [M + R](-) for each R(-) observed was independent of analyte type (e.g., whether the analyte was aliphatic or aromatic, or possessed certain functional groups).

Dissociation Energies of the Alkaline Earth Monofluorides

DOE Office of Scientific and Technical Information (OSTI.GOV)

BLUE, GARY D.; GREEN, JOHN W.; EHLERT, THOMAS C.

1963-08-24

New results and theoretical calculations are presented that indicate consistently high dissocintion energies for all the alkaline earth monofluorides. Experimental results were obtained by utilizing a mass spectrometer to analyze the vapors from a heated Ta Knudsen cell containing an alkaline earth fluoride salt with Al present as a reducing agent. Ionization efficiency curves were obtained and temperature dependence investigations were made to determine the molecular precursor of the ions observed. Values of the equilibrium constants at different temperatures were used together with the free-energy functions to calculate the third law heats of reaction at 298 deg K. Data aremore » tabulated for the heats of various reactions for Al--MF2 systems with M = Mg, Ca, Sr, and Ba, and dissociation energies of MF molecules by various methods for Be, Mg, Ca, Sr, and Ba. (C.H.)« less

High-affinity RNA aptamers to C-reactive protein (CRP): newly developed pre-elution methods for aptamer selection

NASA Astrophysics Data System (ADS)

Orito, N.; Umekage, S.; Sato, K.; Kawauchi, S.; Tanaka, H.; Sakai, E.; Tanaka, T.; Kikuchi, Y.

2012-03-01

We have developed a modified SELEX (systematic evolution of ligands by exponential enrichment) method to obtain RNA aptamers with high affinity to C-reactive protein (CRP). CRP is a clinical biomarker present in plasma, the level of which increases in response to infections and noninfectious inflammation. The CRP level is also an important prognostic indicator in patients with several syndromes. At present, CRP content in blood is measured immunochemically using antibodies. To develop a more sensitive method using RNA aptamers, we have attempted to obtain high-affinity RNA aptamers to CRP. We succeeded in obtaining an RNA aptamer with high affinity to CRP using a CRP-immobilized Sepharose column and pre-elution procedure. Pre-elution is a method that removes the weak binding portion from a selected RNA population by washing for a short time with buffer containing CRP. By surface plasmon-resonance (SPR) analysis, the affinity constant of this aptamer for CRP was calculated to be KD = 2.25×10-9 (M). The secondary structure, contact sites with CRP protein, and application of this aptamer will be described.

Intimate Partner Violence Among Patients With Dissociative Disorders.

PubMed

Webermann, Aliya R; Brand, Bethany L; Kumar, Shaina A

2017-12-01

Childhood trauma is common among survivors and perpetrators of intimate partner violence (IPV). Although symptoms of posttraumatic stress disorder (PTSD) and dissociative disorders (DDs) are predictors of IPV victimization and perpetration, few studies explore IPV among those with DDs. The present study examined IPV and symptoms as predictors among participants in the Treatment of Patients With Dissociative Disorders (TOP DD) Network study, an educational intervention for individuals with DDs and their clinicians. Both clinicians and patients reported on patients' history of physical, emotional, and sexual IPV as both victims and perpetrators. Patients self-reported dissociative, posttraumatic (PTSD), and emotion dysregulation symptoms, as well as IPV-specific dissociative symptoms. According to patients and clinicians, patients were frequently victims of IPV, most commonly emotional IPV. Dissociative symptoms predicted IPV exposure, whereas dissociative and emotion dysregulation symptoms predicted IPV-specific dissociative symptoms.

Serotonin transporter activity of imidazolidine-2,4-dione and imidazo[2,1-f]purine-2,4-dione derivatives in aspect of their acid-base properties.

PubMed

Zagórska, Agnieszka; Czopek, Anna; Pawłowski, Maciej; Dybała, Małgorzata; Siwek, Agata; Nowak, Gabriel

2012-11-01

Affinities of arylpiperazinylalkyl derivatives of imidazo[2,1-f]purine-2,4-dione and imidazolidine-2,4-dione for serotonin transporter and their acid-base properties were evaluated. The dissociation constant (pK(a)) of compounds 1-22 were determinated by potentiometric titration and calculated using pKalc 3.1 module of the Pallas system. The data from experimental methods and computational calculations were compared and suitable conclusions were reached.

Muscarinic receptor occupation and receptor activation in the guinea-pig ileum by some acetamides related to oxotremorine.

PubMed Central

Ringdahl, B.

1984-01-01

The dissociation constants (KD values) and relative efficacies of seven acetamide analogues of oxotremorine, including two enantiomeric pairs, at muscarinic receptors in the guinea-pig isolated ileum were determined. The method used involved analysis of dose-response data before and after fractional inactivation of receptors with propylbenzilylcholine mustard. All of the compounds studied had lower affinities than oxotremorine, but some had substantially higher relative efficacies. Replacement of the pyrrolidine ring of N-methyl-N-(4- pyrrolidino -2- butynyl )acetamide(I), the parent compound in the series, by a dimethylamino or a trimethylammonium group decreased the affinity 32 and 4.5 fold, respectively, whereas the relative efficacy increased 5.7-8.3 times. There was no correlation between relative efficacies and affinities of the compounds. The structural requirements for high affinity and high efficacy appeared to be quite different. PMID:6733356

Dissociation of I II in chemical oxygen-iodine lasers: experiment, modeling, and pre-dissociation by electrical discharge

NASA Astrophysics Data System (ADS)

Katz, A.; Waichman, K.; Dahan, Z.; Rybalkin, V.; Barmashenko, B. D.; Rosenwaks, S.

2007-06-01

The dissociation of I II molecules at the optical axis of a supersonic chemical oxygen-iodine laser (COIL) was studied via detailed measurements and three dimensional computational fluid dynamics calculations. Comparing the measurements and the calculations enabled critical examination of previously proposed dissociation mechanisms and suggestion of a mechanism consistent with the experimental and theoretical results obtained in a supersonic COIL for the gain, temperature and I II dissociation fraction at the optical axis. The suggested mechanism combines the recent scheme of Azyazov and Heaven (AIAA J. 44, 1593 (2006)), where I II(A' 3Π 2u), I II(A 3Π 1u) and O II(a1Δ g, v) are significant dissociation intermediates, with the "standard" chain branching mechanism of Heidner et al. (J. Phys. Chem. 87, 2348 (1983)), involving I(2P 1/2) and I II(X1Σ + g, v). In addition, we examined a new method for enhancement of the gain and power in a COIL by applying DC corona/glow discharge in the transonic section of the secondary flow in the supersonic nozzle, dissociating I II prior to its mixing with O II(1Δ). The loss of O II(1Δ) consumed for dissociation was thus reduced and the consequent dissociation rate downstream of the discharge increased, resulting in up to 80% power enhancement. The implication of this method for COILs operating beyond the specific conditions reported here is assessed.

Structure-guided development of a high-affinity human Programmed Cell Death-1: Implications for tumor immunotherapy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lázár-Molnár, Eszter; Scandiuzzi, Lisa; Basu, Indranil

Programmed Cell Death-1 (PD-1) is an inhibitory immune receptor, which plays critical roles in T cell co-inhibition and exhaustion upon binding to its ligands PD-L1 and PD-L2. We report the crystal structure of the human PD-1 ectodomain and the mapping of the PD-1 binding interface. Mutagenesis studies confirmed the crystallographic interface, and resulted in mutant PD-1 receptors with altered affinity and ligand-specificity. In particular, a high-affinity mutant PD-1 (HA PD-1) exhibited 45 and 30-fold increase in binding to PD-L1 and PD-L2, respectively, due to slower dissociation rates. This mutant (A132L) was used to engineer a soluble chimeric Ig fusion proteinmore » for cell-based and in vivo studies. HA PD-1 Ig showed enhanced binding to human dendritic cells, and increased T cell proliferation and cytokine production in a mixed lymphocyte reaction (MLR) assay. Moreover, in an experimental model of murine Lewis lung carcinoma, HA PD-1 Ig treatment synergized with radiation therapy to decrease local and metastatic tumor burden, as well as in the establishment of immunological memory responses. Our studies highlight the value of structural considerations in guiding the design of a high-affinity chimeric PD-1 Ig fusion protein with robust immune modulatory properties, and underscore the power of combination therapies to selectively manipulate the PD-1 pathway for tumor immunotherapy.« less

Dissociative Functions in the Normal Mourning Process.

ERIC Educational Resources Information Center

Kauffman, Jeffrey

1994-01-01

Sees dissociative functions in mourning process as occurring in conjunction with integrative trends. Considers initial shock reaction in mourning as model of normal dissociation in mourning process. Dissociation is understood to be related to traumatic significance of death in human consciousness. Discerns four psychological categories of…

Binding Energies of Proton-Bound Dimers of Imidazole and n-Acetylalanine Methyl Ester Obtained by Blackbody Infrared Radiative Dissociation

PubMed Central

Jockusch, Rebecca A.; Williams*, Evan R.

2005-01-01

The dissociation kinetics of protonated n-acetyl-L-alanine methyl ester dimer (AcAlaMEd), imidazole dimer, and their cross dimer were measured using blackbody infrared radiative dissociation (BIRD). Master equation modeling of these data was used to extract threshold dissociation energies (Eo) for the dimers. Values of 1.18 ± 0.06, 1.11 ± 0.04, and 1.12 ± 0.08 eV were obtained for AcAlaMEd, imidazole dimer, and the cross dimer, respectively. Assuming that the reverse activation barrier for dissociation of the ion–molecule complex is negligible, the value of Eo can be compared to the dissociation enthalpy (ΔHd°) from HPMS data. The Eo values obtained for the imidazole dimer and the cross dimer are in agreement with HPMS values; the value for AcAlaMEd is somewhat lower. Radiative rate constants used in the master equation modeling were determined using transition dipole moments calculated at the semiempirical (AM1) level for all dimers and compared to ab initio (RHF/3-21G*) calculations where possible. To reproduce the experimentally measured dissociation rates using master equation modeling, it was necessary to multiply semiempirical transition dipole moments by a factor between 2 and 3. Values for transition dipole moments from the ab initio calculations could be used for two of the dimers but appear to be too low for AcAlaMEd. These results demonstrate that BIRD, in combination with master equation modeling, can be used to determine threshold dissociation energies for intermediate size ions that are in neither the truncated Boltzmann nor the rapid energy exchange limit. PMID:16604163

UV-Vis spectrophotometric studies of self-oxidation/dissociation of quaternary ammonium permanganates (QAP) - impact of solvent polarity

NASA Astrophysics Data System (ADS)

Bank, Suraj Prakash; Guru, Partha Sarathi; Dash, Sukalyan

2015-05-01

Self-oxidation/dissociation of some quaternary ammonium permanganates (QAPs), such as cetyltrimethylammonium permanganate (CTAP) and tetrabutylammonium permanganate (TBAP), have been studied spectrophotometrically in six different organic solvent media of different polarities wherein the compounds show good solubility and stability. The optical densities of the substrates at zero time (ODo) and first-order rate constants of dissociation (k1) have been determined from their successive scanning for 40 min. At comparable experimental conditions, absorption capabilities of the substrates are compared from the ODo values in various organic media; the stability of the solutions is compared from the successive scan spectra in those media. The ODo values and the k1 values have been plotted against some solvent parameters to understand their effects on the absorbance and reactivity of the QAPs. These data are also subjected to multiple regression analysis to explain the influence of various solvent parameters on the ion-pairing properties of the substrates, thus elucidating their effects on the process of self-oxidation/dissociation of the substrates.

Molecular modeling of the dissociation of methane hydrate in contact with a silica surface.

PubMed

Bagherzadeh, S Alireza; Englezos, Peter; Alavi, Saman; Ripmeester, John A

2012-03-15

We use constant energy, constant volume (NVE) molecular dynamics simulations to study the dissociation of the fully occupied structure I methane hydrate in a confined geometry between two hydroxylated silica surfaces between 36 and 41 Å apart, at initial temperatures of 283, 293, and 303 K. Simulations of the two-phase hydrate/water system are performed in the presence of silica, with and without a 3 Å thick buffering water layer between the hydrate phase and silica surfaces. Faster decomposition is observed in the presence of silica, where the hydrate phase is prone to decomposition from four surfaces, as compared to only two sides in the case of the hydrate/water simulations. The existence of the water layer between the hydrate phase and the silica surface stabilizes the hydrate phase relative to the case where the hydrate is in direct contact with silica. Hydrates bound between the silica surfaces dissociate layer-by-layer in a shrinking core manner with a curved decomposition front which extends over a 5-8 Å thickness. Labeling water molecules shows that there is exchange of water molecules between the surrounding liquid and intact cages in the methane hydrate phase. In all cases, decomposition of the methane hydrate phase led to the formation of methane nanobubbles in the liquid water phase. © 2012 American Chemical Society

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…

Recent developments in the theory of dissociation

PubMed Central

SPITZER, CARSTEN; BARNOW, SVEN; FREYBERGER, HARALD J; GRABE, HANS JOERGEN

2006-01-01

Although the construct of dissociation was introduced into psychiatry at the end of the 19th century by Pierre Janet, the term still lacks a coherent conceptualization, which is partially reflected by differences in the classification of dissociative and conversion disorders in ICD-10 and DSM-IV. Given the clinical significance of dissociative psychopathology in numerous clinical conditions, it is very valuable that various efforts have been made to refine and to specify current conceptualizations in recent years. The most promising and convincing approaches converge in subdividing dissociation into qualitatively different types, i.e. pathological versus non-pathological dissociation, and "detachment" versus "compartmentalization". We review these concepts and discuss their scientific and clinical potential as well as their limitations. PMID:16946940

Symptom overreporting and dissociative experiences: A qualitative review.

PubMed

Merckelbach, H; Boskovic, I; Pesy, D; Dalsklev, M; Lynn, S J

2017-03-01

We discuss a phenomenon that has received little attention to date in research on dissociative phenomena, namely that self-reports of these phenomena overlap with the tendency to overendorse eccentric items. We review the literature documenting the dissociation-overreporting link and then briefly discuss various interpretations of this link: (1) overreporting is an artifact of measuring dissociative symptoms; (2) dissociative psychopathology engenders overreporting of eccentric symptoms through fantasy proneness or impairments in internal monitoring; (3) an overreporting response style as is evident in malingerers, for example, promotes reports of dissociative symptoms. These three interpretations are not mutually exclusive. Also, the dissociation-overreporting link may have different origins among different samples. Because overreporting may introduce noise in datasets, we need more research specifically aimed at disentangling the dissociation-overreporting link. We suggest various avenues to accomplish this goal. Copyright © 2017 Elsevier Inc. All rights reserved.

Malingering dissociative identity disorder: objective and projective assessment.

PubMed

Labott, Susan M; Wallach, Heather R

2002-04-01

Verification of dissociative identity disorder presents challenges given the complex nature of the illness. This study addressed the concern that this disorder can be successfully malingered on objective and projective psychological tests. 50 undergraduate women were assigned to a Malingering or a Control condition, then completed the Rorschach Inkblot Test and the Dissociative Experiences Scale II. The Malingering group were asked to simulate dissociative identity disorder; controls received instructions to answer all materials honestly. Analysis indicated that malingerers were significantly more likely to endorse dissociative experiences on the Dissociative Experiences Scale II in the range common to patients with diagnosed dissociative identity disorder. However, on the Rorschach there were no significant differences between the two groups. Results suggest that the assessment of dissociative identity disorder requires a multifaceted approach with both objective and projective assessment tools. Research is needed to assess these issues in clinical populations.

The role of water in gas hydrate dissociation

USGS Publications Warehouse

Circone, S.; Stern, L.A.; Kirby, S.H.

2004-01-01

When raised to temperatures above the ice melting point, gas hydrates release their gas in well-defined, reproducible events that occur within self-maintained temperature ranges slightly below the ice point. This behavior is observed for structure I (carbon dioxide, methane) and structure II gas hydrates (methane-ethane, and propane), including those formed with either H2O- or D2O-host frameworks, and dissociated at either ambient or elevated pressure conditions. We hypothesize that at temperatures above the H2O (or D2O) melting point: (1) hydrate dissociation produces water + gas instead of ice + gas, (2) the endothermic dissociation reaction lowers the temperature of the sample, causing the water product to freeze, (3) this phase transition buffers the sample temperatures within a narrow temperature range just below the ice point until dissociation goes to completion, and (4) the temperature depression below the pure ice melting point correlates with the average rate of dissociation and arises from solution of the hydrate-forming gas, released by dissociation, in the water phase at elevated concentrations. In addition, for hydrate that is partially dissociated to ice + gas at lower temperatures and then heated to temperatures above the ice point, all remaining hydrate dissociates to gas + liquid water as existing barriers to dissociation disappear. The enhanced dissociation rates at warmer temperatures are probably associated with faster gas transport pathways arising from the formation of water product.

Psychotherapy and Pharmacotherapy for Patients with Dissociative Identity Disorder

PubMed Central

Gentile, Julie P.; Dillon, Kristy S.

2013-01-01

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. PMID:23556139

Functional (dissociative) retrograde amnesia.

PubMed

Markowitsch, H J; Staniloiu, A

2016-01-01

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. © 2016 Elsevier B.V. All rights reserved.

Dissociation: the clinical realities.

PubMed

Frankel, F H

1996-07-01

An attempt was made by the authors of DSM-III to restrict its focus to the experimental, the observable, and the measurable. The intention was to free the nosology from the influence of unproven theories, and the philosophy was driven largely by the importance of research being able to identify diagnostic categories to facilitate the study of homogeneous groups. So it is of interest that the authors accepted dissociation-an ambiguous event linked to an explicit theoretical concept that had been introduced by Janet-as the basis for classification of clinical presentations that were formerly included under the rubric of hysteria, a similarly unclear category. Since DSM-III, there have been an increasing number of reports of dissociative experiences and dissociative identity disorder (formerly known as multiple personality disorder), but neither of these clinical presentations seems able to withstand the concern that it is dramatically influenced by environmental cues, e.g., the expectations of the therapist. Thus, a restricted phenomenological perspective does not fully appreciate the distorting potential of suggestibility and imagination on the nature of the emerging clinical picture. These factors might well have contributed to and laid the conceptual groundwork for the growth in the number of reports of dissociation.

Anchoring the Gas-Phase Acidity Scale from Hydrogen Sulfide to Pyrrole. Experimental Bond Dissociation Energies of Nitromethane, Ethanethiol, and Cyclopentadiene.

PubMed

Ervin, Kent M; Nickel, Alex A; Lanorio, Jerry G; Ghale, Surja B

2015-07-16

A meta-analysis of experimental information from a variety of sources is combined with statistical thermodynamics calculations to refine the gas-phase acidity scale from hydrogen sulfide to pyrrole. The absolute acidities of hydrogen sulfide, methanethiol, and pyrrole are evaluated from literature R-H bond energies and radical electron affinities to anchor the scale. Relative acidities from proton-transfer equilibrium experiments are used in a local thermochemical network optimized by least-squares analysis to obtain absolute acidities of 14 additional acids in the region. Thermal enthalpy and entropy corrections are applied using molecular parameters from density functional theory, with explicit calculation of hindered rotor energy levels for torsional modes. The analysis reduces the uncertainties of the absolute acidities of the 14 acids to within ±1.2 to ±3.3 kJ/mol, expressed as estimates of the 95% confidence level. The experimental gas-phase acidities are compared with calculations, with generally good agreement. For nitromethane, ethanethiol, and cyclopentadiene, the refined acidities can be combined with electron affinities of the corresponding radicals from photoelectron spectroscopy to obtain improved values of the C-H or S-H bond dissociation energies, yielding D298(H-CH2NO2) = 423.5 ± 2.2 kJ mol(-1), D298(C2H5S-H) = 364.7 ± 2.2 kJ mol(-1), and D298(C5H5-H) = 347.4 ± 2.2 kJ mol(-1). These values represent the best-available experimental bond dissociation energies for these species.

Fundamentals of affinity cell separations.

PubMed

Zhang, Ye; Lyons, Veronica; Pappas, Dimitri

2018-03-01

Cell separations using affinity methods continue to be an enabling science for a wide variety of applications. In this review, we discuss the fundamental aspects of affinity separation, including the competing forces for cell capture and elution, cell-surface interactions, and models for cell adhesion. Factors affecting separation performance such as bond affinity, contact area, and temperature are presented. We also discuss and demonstrate the effects of nonspecific binding on separation performance. Metrics for evaluating cell separations are presented, along with methods of comparing separation techniques for cell isolation using affinity capture. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Kinetics of phloretin binding to phosphatidylcholine vesicle membranes

PubMed Central

1980-01-01

The submillisecond kinetics for phloretin binding to unilamellar phosphatidylcholine (PC) vesicles was investigated using the temperature-jump technique. Spectrophotometric studies of the equilibrium binding performed at 328 nm demonstrated that phloretin binds to a single set of independent, equivalent sites on the vesicle with a dissociation constant of 8.0 microM and a lipid/site ratio of 4.0. The temperature of the phloretin-vesicle solution was jumped by 4 degrees C within 4 microseconds producing a monoexponential, concentration-dependent relaxation process with time constants in the 30--200-microseconds time range. An analysis of the concentration dependence of relaxation time constants at pH 7.30 and 24 degrees C yielded a binding rate constant of 2.7 X 10(8) M-1 s-1 and an unbinding constant of 2,900 s-1; approximately 66 percent of total binding sites are exposed at the outer vesicle surface. The value of the binding rate constant and three additional observations suggest that the binding kinetics are diffusion limited. The phloretin analogue, naringenin, which has a diffusion coefficient similar to phloretin yet a dissociation constant equal to 24 microM, bound to PC vesicle with the same rate constant as phloretin did. In addition, the phloretin-PC system was studied in buffers made one to six times more viscous than water by addition of sucrose or glycerol to the differ. The equilibrium affinity for phloretin binding to PC vesicles is independent of viscosity, yet the binding rate constant decreases with the expected dependence (kappa binding alpha 1/viscosity) for diffusion-limited processes. Thus, the binding rate constant is not altered by differences in binding affinity, yet depends upon the diffusion coefficient in buffer. Finally, studies of the pH dependence of the binding rate constant showed a dependence (kappa binding alpha [1 + 10pH-pK]) consistent with the diffusion-limited binding of a weak acid. PMID:7391812

Low-energy collisionally activated dissociation of pentose-borate complexes

NASA Astrophysics Data System (ADS)

Pepi, Federico; Garzoli, Stefania; Tata, Alessandra; Giacomello, Pierluigi

2010-01-01

Pentose-borate 1:1 complexes were generated in the ESI source of a triple quadrupole and ion trap mass spectrometer by electrospray ionization of Na2B4O7 and pentose (arabinose, lyxose, ribose, xylose) 2:1 solution in CH3CN/H2O. The study of their low-energy collisionally activated dissociation (CAD) demonstrated that ribose and lyxose are preferentially complexed at the C2-C3 cis-diol function whereas arabinose and xylose are esterified at the C1-C2 hydroxyl groups. No evidence was found of the stronger affinity for ribose to borate. The ribose probiotic rule can be explained by considering its peculiar capability, among the investigated pentoses, to almost totally complex the borate anion at the C2-C3 hydroxyl group, thus enabling the subsequent stages of nucleotide assembly, such as phosphorylation and linkage to the nucleobases. Finally, the differences observed in the pentose-borate complex CAD spectra can be used for the mass spectrometric discrimination of isomeric pentoses in complex mixtures.

Clandestine psychopathology: unrecognized dissociative disorders in inpatient psychiatry.

PubMed

Ginzburg, Karni; Somer, Eli; Tamarkin, Gali; Kramer, Lilach

2010-05-01

Surveys among Israeli mental health professionals found that almost half of them doubt the validity of dissociative disorders (DD) and have no experience in either diagnosing or treating DD patients. These findings, in line with arguments that DDs are socially construed North American phenomena, call for the need to investigate it in Israel. Eighty-one psychiatric inpatients were screened for dissociative pathology. Participants categorized as having low levels of dissociation (n = 26) and those demonstrating high levels of dissociation (n = 22) were asked to participate in a Structured Clinical Interview for the DSM-Dissociative Disorders-Revised. One-quarter of all participants were identified as having probable dissociative psychopathology. Based on the Structured Clinical Interview for the DSM-Dissociative Disorders-Revised, estimates of DD range between 12 and 21%. None of the participants had any indication of a DD diagnosis in their medical records. Diagnosis of personality disorder and psychiatric comorbidity were related to the likelihood of a DD diagnosis and its severity.

Dissociation kinetics of Fe(III)- and Al(III)-natural organic matter complexes at pH 6.0 and 8.0 and 25 °C

NASA Astrophysics Data System (ADS)

Jones, Adele M.; Pham, A. Ninh; Collins, Richard N.; Waite, T. David

2009-05-01

The rate at which iron- and aluminium-natural organic matter (NOM) complexes dissociate plays a critical role in the transport of these elements given the readiness with which they hydrolyse and precipitate. Despite this, there have only been a few reliable studies on the dissociation kinetics of these complexes suggesting half-times of some hours for the dissociation of Fe(III) and Al(III) from a strongly binding component of NOM. First-order dissociation rate constants are re-evaluated here at pH 6.0 and 8.0 and 25 °C using both cation exchange resin and competing ligand methods for Fe(III) and a cation exchange resin method only for Al(III) complexes. Both methods provide similar results at a particular pH with a two-ligand model accounting satisfactorily for the dissociation kinetics results obtained. For Fe(III), half-times on the order of 6-7 h were obtained for dissociation of the strong component and 4-5 min for dissociation of the weak component. For aluminium, the half-times were on the order of 1.5 h and 1-2 min for the strong and weak components, respectively. Overall, Fe(III) complexes with NOM are more stable than analogous complexes with Al(III), implying Fe(III) may be transported further from its source upon dilution and dispersion.

Effects of Charge State on Fragmentation Pathways, Dynamics, and Activation Energies of Ubiquitin Ions Measured by Blackbody Infrared Radiative Dissociation

PubMed Central

Jockusch, Rebecca A.; Schnier, Paul D.; Price, William D.; Strittmatter, Eric. F.; Demirev, Plamen A.; Williams*, Evan R.

2005-01-01

Blackbody infrared radiative dissociation spectra of the (M + 5H)5+ through (M + 11H)11+ ions of the protein ubiquitin (8.6 kDa) formed by electrospray ionization were measured in a Fourier-transform mass spectrometer. The 5+ ion dissociates exclusively by loss of water and/or ammonia, whereas the 11+ charge state dissociates only by formation of complementary y and b ions. These two processes are competitive for intermediate charge state ions, with the formation of y and b ions increasingly favored for the higher charge states. The y and b ions are formed by cleavage of the backbone amide bond on the C-terminal side of acidic residues exclusively, with cleavage adjacent to aspartic acid favored. Thermal unimolecular dissociation rate constants for the dissociation of each of these charge states were measured. From the temperature dependence of these rates, Arrhenius activation parameters in the rapid energy exchange limit are obtained. The activation energies (Ea) and preexponential factors (A) for the 5+, 8+, and 9+ ions are 1.2 eV and 1012 s−1, respectively. These values for the 6+ and 7+ ions are 0.9–1.0 eV and 109 s−1, and those for the 10+ and 11+ ions are 1.6 eV and 1016–1017 s−1. Thus, with the exception of the 5+ ion, the higher charge states of ubiquitin have larger dissociation activation energies than the lower charge states. The different A factors observed for production of y and b ions from different precursor charge states indicate that they are formed by different mechanisms, ranging from relatively complex rearrangements to direct bond cleavages. These results clearly demonstrate that the relative dissociation rates of large biomolecule ions by themselves are not necessarily a reliable indicator of their relative dissociation energies, even when similar fragment ions are formed. PMID:9075403

Effects of charge state on fragmentation pathways, dynamics, and activation energies of ubiquitin ions measured by blackbody infrared radiative dissociation.

PubMed

Jockusch, R A; Schnier, P D; Price, W D; Strittmatter, E F; Demirev, P A; Williams, E R

1997-03-15

Blackbody infrared radiative dissociation spectra of the (M + 5H)5+ through (M + 11H)11+ ions of the protein ubiquitin (8.6 kDa) formed by electrospray ionization were measured in a Fourier-transform mass spectrometer. The 5+ ion dissociates exclusively by loss of water and/or ammonia, whereas the 11+ charge state dissociates only by formation of complementary y and b ions. These two processes are competitive for intermediate charge state ions, with the formation of y and b ions increasingly favored for the higher charge states. The y and b ions are formed by cleavage of the backbone amide bond on the C-terminal side of acidic residues exclusively, with cleavage adjacent to aspartic acid favored. Thermal unimolecular dissociation rate constants for the dissociation of each of these charge states were measured. From the temperature dependence of these rates, Arrhenius activation parameters in the rapid energy exchange limit are obtained. The activation energies (Ea) and preexponential factors (A) for the 5+, 8+, and 9+ ions are 1.2 eV and 10(12) s-1, respectively. These values for the 6+ and 7+ ions are 0.9-1.0 eV and 10(9) s-1, and those for the 10+ and 11+ ions are 1.6 eV and 10(16)-10(17) s-1. Thus, with the exception of the 5+ ion, the higher charge states of ubiquitin have larger dissociation activation energies than the lower charge states. The different A factors observed for production of y and b ions from different precursor charge states indicate that they are formed by different mechanisms, ranging from relatively complex rearrangements to direct bond cleavages. These results clearly demonstrate that the relative dissociation rates of large biomolecule ions by themselves are not necessarily a reliable indicator of their relative dissociation energies, even when similar fragment ions are formed.

Exploring evidence of a dissociative subtype in PTSD: Baseline symptom structure, etiology, and treatment efficacy for those who dissociate.

PubMed

Burton, Mark S; Feeny, Norah C; Connell, Arin M; Zoellner, Lori A

2018-05-01

With the inclusion of a dissociative subtype, recent changes to the DSM-5 diagnosis of posttraumatic stress disorder (PTSD) have emphasized the role of dissociation in the experience and treatment of the disorder. However, there is a lack of research exploring the clinical impact for highly dissociative groups receiving treatment for PTSD. The current study examined the presence and clinical impact of a dissociative subtype in a sample of individuals receiving treatment for chronic PTSD. This study used latent transition analyses (LTA), an expanded form of latent profile analyses (LPA), to examine latent profiles of PTSD and dissociation symptoms before and after treatment for individuals (N = 200) receiving prolonged exposure (PE) or sertraline treatment for chronic PTSD. The best fitting LTA model was one with a 4-class solution at both pretreatment and posttreatment. There was a latent class at pretreatment with higher levels of dissociative symptoms. However, this class was also marked by higher reexperiencing symptoms, and membership was not predicted by chronic child abuse. Further, although those in the class were less likely to transition to the responder class overall, this was not the case for exposure-based treatment specifically. These findings are not in line with the dissociative-subtype theoretical literature that proposes those who dissociate represent a clinically distinct group that may respond worse to exposure-based treatments for PTSD. (PsycINFO Database Record (c) 2018 APA, all rights reserved).

Determining the folding and unfolding rate constants of nucleic acids by biosensor. Application to telomere G-quadruplex.

PubMed

Zhao, Yong; Kan, Zhong-yuan; Zeng, Zhi-xiong; Hao, Yu-hua; Chen, Hua; Tan, Zheng

2004-10-20

Nucleic acid molecules may fold into secondary structures, and the formation of such structures is involved in many biological processes and technical applications. The folding and unfolding rate constants define the kinetics of conformation interconversion and the stability of these structures and is important in realizing their functions. We developed a method to determine these kinetic parameters using an optical biosensor based on surface plasmon resonance. The folding and unfolding of a nucleic acid is coupled with a hybridization reaction by immobilization of the target nucleic acid on a sensor chip surface and injection of a complementary probe nucleic acid over the sensor chip surface. By monitoring the time course of duplex formation, both the folding and unfolding rate constants for the target nucleic acid and the association and dissociation rate constants for the target-probe duplex can all be derived from the same measurement. We applied this method to determine the folding and unfolding rate constants of the G-quadruplex of human telomere sequence (TTAGGG)(4) and its association and dissociation rate constants with the complementary strand (CCCTAA)(4). The results show that both the folding and unfolding occur on the time scale of minutes at physiological concentration of K(+). We speculate that this property might be important for telomere elongation. A complete set of the kinetic parameters for both of the structures allows us to study the competition between the formation of the quadruplex and the duplex. Calculations indicate that the formation of both the quadruplex and the duplex is strand concentration-dependent, and the quadruplex can be efficiently formed at low strand concentration. This property may provide the basis for the formation of the quadruplex in vivo in the presence of a complementary strand.

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.

Identity, Affinity, Reality: Making the Case for Affinity Groups in Elementary School

ERIC Educational Resources Information Center

Parsons, Julie; Ridley, Kimberly

2012-01-01

Affinity groups are places where students build connections and process "ouch" moments from their classes. Children talk about the isolation they sometimes feel. The relationships students gain through race-based affinity groups enable them to feel less alone with their emotions and help them build a stronger sense of self. At the same…

Does dissociation offer a useful explanation for psychopathology?

PubMed

Jureidini, Jon

2004-01-01

Dissociation is often conceptualised as an altered state of consciousness, a trance-like state in which normal barriers between conscious and unconscious memories, desires and beliefs break down and other amnestic barriers emerge. This review explores whether it is likely that there is a neurophysiology of pathological dissociative processes that will elucidate management. A critical reading of current research, sourced through Medline and Psychinfo searches from 1990 to 2002, using subject headings: dissociative disorders, hypnosis and stress disorder (post-traumatic), as well as keywords: dissociation, hypnosis and trance. Current knowledge does not support the notion of dissociation as a discrete brain state or process. Psychiatric and neurophysiological research and theory development are better directed towards individual components that contribute to dissociative experience. Copyright (c) 2004 S. Karger AG, Basel.

Measurement of threshold temperature effects in dissociative electron attachment to HI and DI

NASA Technical Reports Server (NTRS)

Chutjian, A.; Alajajian, S. H.; Man, K.-F.

1990-01-01

From accurate spectroscopic constants it is found that the thermal dissociative-attachment process (DA) in DI should be exothermic only for rotational levels J greater than 8 in v = O. Here, measurement of an enhancement of DA with rotational temperature T in the range 298-468 K is reported. The effect is easily accounted for by the increase in total fractional population of excited J levels in DI relative to HI. The effect affords a rotational analog to the use of vibrationally excited molecules (e.g., HCl) in a plasma to control electron conduction.

Rate coefficients for dissociative attachment and resonant electron-impact dissociation involving vibrationally excited O{sub 2} molecules

DOE Office of Scientific and Technical Information (OSTI.GOV)

Laporta, V.; Celiberto, R.; Tennyson, J.

Rate coefficients for dissociative electron attachment and electron-impact dissociation processes, involving vibrationally excited molecular oxygen, are presented. Analytical fits of the calculated numerical data, useful in the applications, are also provided.

Glycolipid acquisition by human glycolipid transfer protein dramatically alters intrinsic tryptophan fluorescence: insights into glycolipid binding affinity.

PubMed

Zhai, Xiuhong; Malakhova, Margarita L; Pike, Helen M; Benson, Linda M; Bergen, H Robert; Sugár, István P; Malinina, Lucy; Patel, Dinshaw J; Brown, Rhoderick E

2009-05-15

Glycolipid transfer proteins (GLTPs) are small, soluble proteins that selectively accelerate the intermembrane transfer of glycolipids. The GLTP fold is conformationally unique among lipid binding/transfer proteins and serves as the prototype and founding member of the new GLTP superfamily. In the present study, changes in human GLTP tryptophan fluorescence, induced by membrane vesicles containing glycolipid, are shown to reflect glycolipid binding when vesicle concentrations are low. Characterization of the glycolipid-induced "signature response," i.e. approximately 40% decrease in Trp intensity and approximately 12-nm blue shift in emission wavelength maximum, involved various modes of glycolipid presentation, i.e. microinjection/dilution of lipid-ethanol solutions or phosphatidylcholine vesicles, prepared by sonication or extrusion and containing embedded glycolipids. High resolution x-ray structures of apo- and holo-GLTP indicate that major conformational alterations are not responsible for the glycolipid-induced GLTP signature response. Instead, glycolipid binding alters the local environment of Trp-96, which accounts for approximately 70% of total emission intensity of three Trp residues in GLTP and provides a stacking platform that aids formation of a hydrogen bond network with the ceramide-linked sugar of the glycolipid headgroup. The changes in Trp signal were used to quantitatively assess human GLTP binding affinity for various lipids including glycolipids containing different sugar headgroups and homogenous acyl chains. The presence of the glycolipid acyl chain and at least one sugar were essential for achieving a low-to-submicromolar dissociation constant that was only slightly altered by increased sugar headgroup complexity.

Hypnotic suggestibility, cognitive inhibition, and dissociation.

PubMed

Dienes, Zoltán; Brown, Elizabeth; Hutton, Sam; Kirsch, Irving; Mazzoni, Giuliana; Wright, Daniel B

2009-12-01

We examined two potential correlates of hypnotic suggestibility: dissociation and cognitive inhibition. Dissociation is the foundation of two of the major theories of hypnosis and other theories commonly postulate that hypnotic responding is a result of attentional abilities (including inhibition). Participants were administered the Waterloo-Stanford Group Scale of Hypnotic Susceptibility, Form C. Under the guise of an unrelated study, 180 of these participants also completed: a version of the Dissociative Experiences Scale that is normally distributed in non-clinical populations; a latent inhibition task, a spatial negative priming task, and a memory task designed to measure negative priming. The data ruled out even moderate correlations between hypnotic suggestibility and all the measures of dissociation and cognitive inhibition overall, though they also indicated gender differences. The results are a challenge for existing theories of hypnosis.

Dissociation, shame, complex PTSD, child maltreatment and intimate relationship self-concept in dissociative disorder, chronic PTSD and mixed psychiatric groups.

PubMed

Dorahy, Martin J; Middleton, Warwick; Seager, Lenaire; McGurrin, Patrick; Williams, Mary; Chambers, Ron

2015-02-01

Whilst a growing body of research has examined dissociation and other psychiatric symptoms in severe dissociative disorders (DDs), there has been no systematic examination of shame and sense of self in relationships in DDs. Chronic child abuse often associated with severe DDs, like dissociative identity disorder, is likely to heighten shame and relationship concerns. This study investigated complex posttraumatic stress disorder (PTSD), borderline and Schneiderian symptoms, dissociation, shame, child abuse, and various markers of self in relationships (e.g., relationship esteem, relationship depression, fear of relationships). Participants were assessed via clinical interview with psychometrically sound questionnaires. They fell into three diagnostic groups, dissociative disorder (n=39; primarily dissociative identity disorder), chronic PTSD (Chr-PTSD; n=13) or mixed psychiatric presentations (MP; n=21; primarily mood and anxiety disorders). All participants had a history of child abuse and/or neglect, and the groups did not differ on age and gender. The DD group was higher on nearly all measured variables than the MP group, and had more severe dissociative, borderline and Schneiderian symptoms than the Chr-PTSD sample. Shame and complex PTSD symptoms fell marginally short of predicting reductions in relationship esteem, pathological dissociative symptoms predicted increased relationship depression, and complex PTSD symptoms predicted fear of relationships. The representativeness of the samples was unknown. Severe psychiatric symptoms differentiate DDs from chronic PTSD, while dissociation and shame have a meaningful impact on specific markers of relationship functioning in psychiatric patients with a history of child abuse and neglect. Copyright © 2014 Elsevier B.V. All rights reserved.

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.

Trends in water monomer adsorption and dissociation on flat insulating surfaces.

PubMed

Hu, Xiao Liang; Carrasco, Javier; Klimeš, Jiří; Michaelides, Angelos

2011-07-21

The interaction of water with solid surfaces is key to a wide variety of industrial and natural processes. However, the basic principles that dictate how stable and in which state (intact or dissociated) water will be on a given surface are not fully understood. Towards this end, we have used density functional theory to examine water monomer adsorption on the (001) surfaces of a broad range of alkaline earth oxides, alkaline earth sulfides, alkali fluorides, and alkali chlorides. Some interesting general conclusions are arrived at: (i) on all the surfaces considered only a few specific adsorption structures are favoured; (ii) water becomes more stable upon descending the oxide and fluoride series but does not vary much upon going down the chloride and sulfide series; (iii) water is stabilised both by an increase in the lattice constant, which facilitates hydrogen bonding to the substrate, and by the flexibility of the substrate. These are also factors that favour water dissociation. We hope that this study is of some value in better understanding the surface science of water in general, and in assisting in the interpretation and design of future experiments. This journal is © the Owner Societies 2011

Prostaglandin E and F2 alpha receptors in human myometrium during the menstrual cycle and in pregnancy and labor

DOE Office of Scientific and Technical Information (OSTI.GOV)

Giannopoulos, G.; Jackson, K.; Kredentser, J.

The binding of prostaglandins E1 and F2 alpha has been studied in the human myometrium and cervix during the menstrual cycle and in the myometrium of pregnant patients at term before and during labor. Tritium-labeled prostaglandin E1 and F2 alpha binding was saturable and reversible. Scatchard analysis of tritium-labeled prostaglandin E1 binding was linear, which suggests a single class of high-affinity binding sites with an estimated apparent equilibrium dissociation constant of 2.5 to 5.4 nmol/L and inhibitor affinities of 0.9, 273, 273, and 217 nmol/L for prostaglandins E2, A1, B1, and F2 alpha, respectively. Scatchard analysis of tritium-labeled prostaglandin F2more » alpha, binding was also linear, but the affinity of these binding sites was much lower, with an average dissociation constant of 50 nmol/L and inhibitor affinities of 1.6, 2.2, and 11.2 nmol/L for prostaglandins E1, E2, and A1, respectively. In nonpregnant patients, the concentrations and affinities of tritium-labeled prostaglandin E1 binding sites were similar in the myometrium during the proliferative and secretory phases of the menstrual cycle, but the concentration of these sites was much lower in the cervix. The concentration of the tritium-labeled prostaglandin E1 binding sites was significantly lower in the myometrium of pregnant patients at term than in the myometrium of nonpregnant patients. The concentrations and affinities of tritium-labeled prostaglandin E1 binding sites were not significantly different in the upper and lower myometrium of pregnant patients at term or in the myometrium of such patients before and during labor. The concentrations of the tritium-labeled prostaglandin F2 alpha binding sites during the menstrual cycle and in pregnancy at term were similar to those of tritium-labeled prostaglandin E1 binding sites.« less

Thermodynamic stability of carbonic anhydrase: measurements of binding affinity and stoichiometry using ThermoFluor.

PubMed

Matulis, Daumantas; Kranz, James K; Salemme, F Raymond; Todd, Matthew J

2005-04-05

ThermoFluor (a miniaturized high-throughput protein stability assay) was used to analyze the linkage between protein thermal stability and ligand binding. Equilibrium binding ligands increase protein thermal stability by an amount proportional to the concentration and affinity of the ligand. Binding constants (K(b)) were measured by examining the systematic effect of ligand concentration on protein stability. The precise ligand effects depend on the thermodynamics of protein stability: in particular, the unfolding enthalpy. An extension of current theoretical treatments was developed for tight binding inhibitors, where ligand effect on T(m) can also reveal binding stoichiometry. A thermodynamic analysis of carbonic anhydrase by differential scanning calorimetry (DSC) enabled a dissection of the Gibbs free energy of stability into enthalpic and entropic components. Under certain conditions, thermal stability increased by over 30 degrees C; the heat capacity of protein unfolding was estimated from the dependence of calorimetric enthalpy on T(m). The binding affinity of six sulfonamide inhibitors to two isozymes (human type 1 and bovine type 2) was analyzed by both ThermoFluor and isothermal titration calorimetry (ITC), resulting in a good correlation in the rank ordering of ligand affinity. This combined investigation by ThermoFluor, ITC, and DSC provides a detailed picture of the linkage between ligand binding and protein stability. The systematic effect of ligands on stability is shown to be a general tool to measure affinity.

Quartz crystal microbalance for the cardiac markers/antibodies binding kinetic measurements in the plasma samples

NASA Astrophysics Data System (ADS)

Agafonova, L. E.; Shumyantseva, V. V.; Archakov, A. I.

2014-06-01

The quartz crystal microbalance (QCM) was exploited for cardiac markers detection and kinetic studies of immunochemical reaction of cardiac troponin I (cTnI) and human heart fatty acid binding protein (H-FABP) with the corresponding monoclonal antibodies in undiluted plasma (serum) and standard solutions. The QCM technique allowed to dynamically monitor the kinetic differences in specific interactions and nonspecific sorption, without multiple labeling procedures and separation steps. The affinity binding process was characterized by the association (ka) and the dissociation (kd) kinetic constants and the equilibrium association (K) constant, all of which were obtained from experimental data.

Development and Preliminary Psychometric Properties of an Instrument for the Measurement of Obsessional Dissociative Experiences: The Van Obsessional Dissociation Questionnaire (VOD-Q).

PubMed

Boysan, Murat; Yıldırım, Abdullah; Beşiroğlu, Lütfullah; Kefeli, Mehmet Celal; Kağan, Mücahit

2018-01-04

A growing body of research evidence documents the substantial associations between obsessive-compulsive disorder (OCD) and dissociation. This article describes the development and preliminary psychometric properties of the Van Obsessional Dissociation Questionnaire (VOD-Q). Obsessional dissociation is defined as a tendency to dissociate in reaction to distressing, unwanted and intrusive thoughts, images and impulses. The screening tool is conceptualized to tap obsessional dissociation across three dimensions: obsessional absorption, obsessional depersonalization/ derealization and obsessional amnesia. The VOD-Q, the Padua Inventory-Revised (PI-R), the Dissociative Experiences Scale (DES), and the Obsessive Belief Questionnaire (OBQ-44) were administered in this study. The results showed that the VOD-Q had excellent test-retest reliability (ranging from 0.73 to 0.90) and internal consistency (ranging from 0.90 to 0.97). The VOD-Q total and subscale scores were significantly associated with measures of obsessive-compulsive symptoms and dissociative experiences. OCD patients scored significantly higher on the VOD-Q than community participants. Based on the present findings, the VOD-Q appears to be a reliable and valid instrument for the measurement of obsessional dissociative experiences.

[Dissociative disorders: from Janet to DSM-IV].

PubMed

Nakatani, Y

2000-01-01

I reviewed the literature on dissociation and dissociative disorders from Pierre Janet to DSM-IV, and examined the current trends in research. Janet's theory on hysteria is multifaceted, and is based on three psychological models. Based on a hierarchical model, Janet related hysteric symptoms to the activities within the lower strata of mental hierarchy (automatisms psychologiques), which were demonstrably shown in somnambulism. A second model was based on the concept of a psychological system, which was hypothetically composed of ideas, images, feelings, sensations, and movements. According to this model, dissociation of psychological functions was fundamental to the mechanism of hysteria: loss of integration was thought to engender fixed ideas (ideas fixes) and to lead to the development of a system totally isolated from the whole personality system. Janet also attempted to explain various mental disorders using an economic model. He referred to a loss of equilibration between psychological force and psychological tension. Thus, an unexpected emotional experience was conceived to cause a consumption of reserved psychological force, which was in turn followed by exhaustion associated with hysteric symptoms. Whereas most current researchers regard Janet as the first to study psychological trauma as a principal cause of dissociation, I feel it is important to note that he also emphasized the role of stigmata, i.e., permanent traits of hysteric patients, which were represented as a suggestibility and a tendency toward a narrowing of the consciousness field. Discussion about dissociation and its relation to trauma all but disappeared after Janet. However, during the Second World War and post-war period, some psychiatrists began to pay attention to two emerging phenomena: a high incidence of dissociative symptoms such as fugue and amnesia among combatants, and traumatic neurosis frequently observed among ex-inmates of concentration camps. In the 1970s, interest in

Rate constants for chemical reactions in high-temperature nonequilibrium air

NASA Technical Reports Server (NTRS)

Jaffe, R. L.

1986-01-01

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

Biomolecular Interaction Analysis Using an Optical Surface Plasmon Resonance Biosensor: The Marquardt Algorithm vs Newton Iteration Algorithm

PubMed Central

Hu, Jiandong; Ma, Liuzheng; Wang, Shun; Yang, Jianming; Chang, Keke; Hu, Xinran; Sun, Xiaohui; Chen, Ruipeng; Jiang, Min; Zhu, Juanhua; Zhao, Yuanyuan

2015-01-01

Kinetic analysis of biomolecular interactions are powerfully used to quantify the binding kinetic constants for the determination of a complex formed or dissociated within a given time span. Surface plasmon resonance biosensors provide an essential approach in the analysis of the biomolecular interactions including the interaction process of antigen-antibody and receptors-ligand. The binding affinity of the antibody to the antigen (or the receptor to the ligand) reflects the biological activities of the control antibodies (or receptors) and the corresponding immune signal responses in the pathologic process. Moreover, both the association rate and dissociation rate of the receptor to ligand are the substantial parameters for the study of signal transmission between cells. A number of experimental data may lead to complicated real-time curves that do not fit well to the kinetic model. This paper presented an analysis approach of biomolecular interactions established by utilizing the Marquardt algorithm. This algorithm was intensively considered to implement in the homemade bioanalyzer to perform the nonlinear curve-fitting of the association and disassociation process of the receptor to ligand. Compared with the results from the Newton iteration algorithm, it shows that the Marquardt algorithm does not only reduce the dependence of the initial value to avoid the divergence but also can greatly reduce the iterative regression times. The association and dissociation rate constants, ka, kd and the affinity parameters for the biomolecular interaction, KA, KD, were experimentally obtained 6.969×105 mL·g-1·s-1, 0.00073 s-1, 9.5466×108 mL·g-1 and 1.0475×10-9 g·mL-1, respectively from the injection of the HBsAg solution with the concentration of 16ng·mL-1. The kinetic constants were evaluated distinctly by using the obtained data from the curve-fitting results. PMID:26147997

A rhodium(III) complex for high-affinity DNA base-pair mismatch recognition

PubMed Central

Junicke, Henrik; Hart, Jonathan R.; Kisko, Jennifer; Glebov, Oleg; Kirsch, Ilan R.; Barton, Jacqueline K.

2003-01-01

A rhodium(III) complex, rac-[Rh(bpy)2phzi]3+ (bpy, 2,2′-bipyridine; phzi, benzo[a]phenazine-5,6-quinone diimine) has been designed as a sterically demanding intercalator targeted to destabilized mismatched sites in double-helical DNA. The complex is readily synthesized by condensation of the phenazine quinone with the corresponding diammine complex. Upon photoactivation, the complex promotes direct strand scission at single-base mismatch sites within the DNA duplex. As with the parent mismatch-specific reagent, [Rh(bpy)2(chrysi)]3+ [chrysene-5,6-quinone diimine (chrysi)], mismatch selectivity depends on the helix destabilization associated with mispairing. Unlike the parent chrysi complex, the phzi analogue binds and cleaves with high affinity and efficiency. The specific binding constants for CA, CC, and CT mismatches within a 31-mer oligonucleotide duplex are 0.3, 1, and 6 × 107 M−1, respectively; site-specific photocleavage is evident at nanomolar concentrations. Moreover, the specificity, defined as the ratio in binding affinities for mispaired vs. well paired sites, is maintained. The increase in affinity is attributed to greater stability in the mismatched site associated with stacking by the heterocyclic aromatic ligand. The high-affinity complex is also applied in the differential cleavage of DNA obtained from cell lines deficient in mismatch repair vs. those proficient in mismatch repair. Agreement is found between photocleavage by the mismatch-specific probes and deficiency in mismatch repair. This mismatch-specific targeting, therefore, offers a potential strategy for new chemotherapeutic design. PMID:12610209

Influence of Coulombic repulsion on the dissociation pathways and energetics of multiprotein complexes in the gas phase.

PubMed

Sinelnikov, Igor; Kitova, Elena N; Klassen, John S

2007-04-01

Thermal dissociation experiments, implemented with blackbody infrared radiative dissociation and Fourier-transform ion cyclotron resonance mass spectrometry, are performed on gaseous protonated and deprotonated ions of the homopentameric B subunits of Shiga toxin 1 (Stx1 B5) and Shiga toxin 2 (Stx2 B5) and the homotetramer streptavidin (S4). Dissociation of the gaseous, multisubunit complexes proceeds predominantly by the loss of a single subunit. Notably, the fractional partitioning of charge between the product ions, i.e., the leaving subunit and the resulting multimer, for a given complex is, within error, constant over the range of charge states investigated. The Arrhenius activation parameters (E(a), A) measured for the loss of subunit decrease with increasing charge state of the complex. However, the parameters for the protonated and deprotonated ions, with the same number of charges, are indistinguishable. The influence of the complex charge state on the dissociation pathways and the magnitude of the dissociation E(a) are modeled theoretically with the discrete charge droplet model (DCDM) and the protein structure model (PSM), wherein the structure of the subunits is considered. Importantly, the major subunit charge states observed experimentally for the Stx1 B5(n+/-) ions correspond to the minimum energy charge distribution predicted by DCDM and PSM assuming a late dissociative transition-state (TS); while for structurally-related Stx2 B5(n+) ions, the experimental charge distribution corresponds to an early TS. It is proposed that the lateness of the TS is related, in part, to the degree of unfolding of the leaving subunit, with Stx1 B being more unfolded than Stx2 B. PSM, incorporating significant subunit unfolding is necessary to account for the product ions observed for the S4(n+) ions. The contribution of Coulombic repulsion to the dissociation E(a) is quantified and the intrinsic activation energy is estimated for the first time.

A latent class analysis of dissociation and posttraumatic stress disorder: evidence for a dissociative subtype.

PubMed

Wolf, Erika J; Miller, Mark W; Reardon, Annemarie F; Ryabchenko, Karen A; Castillo, Diane; Freund, Rachel

2012-07-01

The nature of the relationship of dissociation to posttraumatic stress disorder (PTSD) is controversial and of considerable clinical and nosologic importance. To examine evidence for a dissociative subtype of PTSD and to examine its association with different types of trauma. A latent profile analysis of cross-sectional data from structured clinical interviews indexing DSM-IV symptoms of current PTSD and dissociation. The VA Boston Healthcare System and the New Mexico VA Health Care System. A total of 492 veterans and their intimate partners, all of whom had a history of trauma. Participants reported exposure to a variety of traumatic events, including combat, childhood physical and sexual abuse, partner abuse, motor vehicle accidents, and natural disasters, with most participants reporting exposure to multiple types of traumatic events. Forty-two percent of the sample met the criteria for a current diagnosis of PTSD. Item-level scores on the Clinician-Administered PTSD Scale. A latent profile analysis suggested a 3-class solution: a low PTSD severity subgroup, a high PTSD severity subgroup characterized by elevations across the 17 core symptoms of the disorder, and a small but distinctly dissociative subgroup that composed 12% of individuals with a current diagnosis of PTSD. The latter group was characterized by severe PTSD symptoms combined with marked elevations on items assessing flashbacks, derealization, and depersonalization. Individuals in this subgroup also endorsed greater exposure to childhood and adult sexual trauma compared with the other 2 groups, suggesting a possible etiologic link with the experience of repeated sexual trauma. These results support the subtype hypothesis of the association between PTSD and dissociation and suggest that dissociation is a highly salient facet of posttraumatic psychopathology in a subset of individuals with the disorder.

Photolabeling of Tonoplast from Sugar Beet Cell Suspensions by [3H]5-(N-Methyl-N-Isobutyl)-Amiloride, an Inhibitor of the Vacuolar Na+/H+ Antiport 1

PubMed Central

Barkla, Bronwyn J.; Charuk, Jeffrey H. M.; Cragoe, Edward J.; Blumwald, Eduardo

1990-01-01

The effects of 5-(N-methyl-N-isobutyl)-amiloride (MIA), an amiloride analog, was tested on the Na+/H+ antiport activity of intact vacuoles and tonoplast vesicles isolated from sugar beet (Beta vulgaris L.) cell suspension cultures. MIA inhibited Na+/H+ exchange in a competitive manner with a Ki of 2.5 and 5.9 micromolar for ΔpH-dependent 22Na+ influx in tonoplast vesicles and Na+-dependent H+ efflux in intact vacuoles, respectively. Scatchard analysis of the binding of [3H]MIA to tonoplast membranes revealed a high affinity binding component with a Kd of 1.3 micromolar. The close relationship between the dissociation constant value obtained and the constants of inhibition for MIA obtained by fluorescence quenching and isotope exchange suggests that the high affinity component represents a class of sites associated with the tonoplast Na+/H+ antiport. Photolabeling of the tonoplast with [3H]MIA revealed two sets of polypeptides with a different affinity to amiloride and its analog. Images Figure 7 PMID:16667602

Analysis of free drug fractions in serum by ultrafast affinity extraction and two-dimensional affinity chromatography using α1-acid glycoprotein microcolumns.

PubMed

Bi, Cong; Zheng, Xiwei; Hage, David S

2016-02-05

In the circulatory system, many drugs are reversibly bound to serum proteins such as human serum albumin (HSA) and alpha1-acid glycoprotein (AGP), resulting in both free and protein-bound fractions for these drugs. This report examined the use of microcolumns containing immobilized AGP for the measurement of free drug fractions by ultrafast affinity extraction and a two-dimensional affinity system. Several drugs known to bind AGP were used as models to develop and evaluate this approach. Factors considered during the creation of this method included the retention of the drugs on the microcolumns, the injection flow rate, the microcolumn size, and the times at which a second AGP column was placed on-line with the microcolumn. The final system had residence times of only 110-830ms during sample passage through the AGP microcolumns and allowed free drug fractions to be determined within 10-20min when using only 3-10μL of sample per injection. This method was used to measure the free fractions of the model drugs at typical therapeutic levels in serum, giving good agreement with the results obtained by ultrafiltration. This approach was also used to estimate the binding constants for each drug with AGP in serum, even for drugs that had significant interactions with both AGP and HSA in such samples. These results indicated that AGP microcolumns could be used with ultrafast affinity extraction to measure free drug fractions in a label-free manner and to study the binding of drugs with AGP in complex samples such as serum. Copyright © 2015 Elsevier B.V. All rights reserved.

Mobility-based correction for accurate determination of binding constants by capillary electrophoresis-frontal analysis.

PubMed

Qian, Cheng; Kovalchik, Kevin A; MacLennan, Matthew S; Huang, Xiaohua; Chen, David D Y

2017-06-01

Capillary electrophoresis frontal analysis (CE-FA) can be used to determine binding affinity of molecular interactions. However, its current data processing method mandate specific requirement on the mobilities of the binding pair in order to obtain accurate binding constants. This work shows that significant errors are resulted when the mobilities of the interacting species do not meet these requirements. Therefore, the applicability of CE-FA in many real word applications becomes questionable. An electrophoretic mobility-based correction method is developed in this work based on the flux of each species. A simulation program and a pair of model compounds are used to verify the new equations and evaluate the effectiveness of this method. Ibuprofen and hydroxypropyl-β-cyclodextrinare used to demonstrate the differences in the obtained binding constant by CE-FA when different calculation methods are used, and the results are compared with those obtained by affinity capillary electrophoresis (ACE). The results suggest that CE-FA, with the mobility-based correction method, can be a generally applicable method for a much wider range of applications. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Respiratory properties of blood and arterial blood gases in the tegu lizard: effects of temperature and hypercapnia.

PubMed

Wood, S C; Glass, M L; Andersen, N A; Heisler, N

1987-01-01

The effects of body temperature and hypercapnia (7% inspired CO2) on arterial blood gases, plasma pH, and the characteristics of the blood oxygen dissociation curve were determined in Tegu lizards (Tupinambis nigropunctatus). Arterial pH fell from 7.59 to 7.50 when body temperature was increased from 25 to 35 degrees C. The pH/temperature coefficient (delta pH/delta t = -0.009 U/degrees C) was half of that predicted on the basis of 'constant relative alkalinity' and the alphastat hypothesis. The fall in plasma pH resulted from a decrease in plasma [HCO3-], and a rise in plasma Pco2. The O2 affinity of Tegu blood, expressed by the partial pressure at half saturation (P50), decreased with temperature in vitro from 42.3 to 49.6 torr at pH 7.4. The apparent enthalpy (delta H = -3.1 kcal/mol) is about 1/4 of that of human blood. In vivo, the arterial blood oxygen saturation decreased from 89% at 25 degrees to 82% at 35 degrees C. Arterial Po2 increased from 61 to 71 torr as expected from the right-shift of the oxygen dissociation curve. During environmental hypercapnia (7% CO2, 21% O2, 72% N2 inspired concentrations), arterial pH decreased to 7.28. Arterial O2 saturation remained constant and arterial Po2 increased from 61 to 85 torr due to the right-shift of the oxygen dissociation curve. The comparatively small effect of changes in temperature on the oxygen affinity of Tegu blood (directly according to the delta H value, and indirectly via changes in blood pH) results in a relatively small right shift of the oxygen dissociation curve, and accordingly in relatively high arterial and tissue Po2 values also at higher temperatures.

Tubulin Dimer Reversible Dissociation

PubMed Central

Schuck, Peter; Sackett, Dan L.

2016-01-01

Tubulins are evolutionarily conserved proteins that reversibly polymerize and direct intracellular traffic. Of the tubulin family only αβ-tubulin forms stable dimers. We investigated the monomer-dimer equilibrium of rat brain αβ-tubulin using analytical ultracentrifugation and fluorescence anisotropy, observing tubulin in virtually fully monomeric and dimeric states. Monomeric tubulin was stable for a few hours and exchanged into preformed dimers, demonstrating reversibility of dimer dissociation. Global analysis combining sedimentation velocity and fluorescence anisotropy yielded Kd = 84 (54–123) nm. Dimer dissociation kinetics were measured by analyzing the shape of the sedimentation boundary and by the relaxation of fluorescence anisotropy following rapid dilution of labeled tubulin, yielding koff in the range 10−3–10−2 s−1. Thus, tubulin dimers reversibly dissociate with moderately fast kinetics. Monomer-monomer association is much less sensitive than dimer-dimer association to solution changes (GTP/GDP, urea, and trimethylamine oxide). PMID:26934918

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.

Selection and identification of a DNA aptamer targeted to Vibrio parahemolyticus.

PubMed

Duan, Nuo; Wu, Shijia; Chen, Xiujuan; Huang, Yukun; Wang, Zhouping

2012-04-25

A whole-bacterium systemic evolution of ligands by exponential enrichment (SELEX) method was applied to a combinatorial library of FAM-labeled single-stranded DNA molecules to identify DNA aptamers demonstrating specific binding to Vibrio parahemolyticus . FAM-labeled aptamer sequences with high binding affinity to V. parahemolyticus were identified by flow cytometric analysis. Aptamer A3P, which showed a particularly high binding affinity in preliminary studies, was chosen for further characterization. This aptamer displayed a dissociation constant (K(d)) of 16.88 ± 1.92 nM. Binding assays to assess the specificity of aptamer A3P showed a high binding affinity (76%) for V. parahemolyticus and a low apparent binding affinity (4%) for other bacteria. Whole-bacterium SELEX is a promising technique for the design of aptamer-based molecular probes for microbial pathogens that does not require the labor-intensive steps of isolating and purifying complex markers or targets.

Aspects of Rhodobacter sphaeroides ChrR required for stimuli to promote dissociation of σE/ChrR complexes

PubMed Central

Greenwell, Roger; Nam, Tae-Wook; Donohue, Timothy J.

2011-01-01

In the photosynthetic bacterium Rhodobacter sphaeroides, a transcriptional response to the reactive oxygen species singlet oxygen (1O2) is mediated by ChrR, a zinc metalloprotein that binds to and inhibits activity of the alternative sigma factor, σE. We provide evidence that 1O2 promotes dissociation of σE from ChrR to activate transcription in vivo. To identify what is required for 1O2 to promote dissociation of σE/ChrR complexes, we analyzed the in vivo properties of variant ChrR proteins with amino acid changes in conserved residues of the C-terminal cupin-like domain (ChrR-CLD). We found that 1O2 was unable to promote detectable dissociation of σE/ChrR complexes when the ChrR-CLD zinc ligands (His141, His143, Glu147, and His177) were substituted with alanine, even though individual substitutions caused a 2- to 10-fold decrease in zinc affinity for this domain relative to that of wild-type ChrR (Kd ∼4.6 × 10−10 M). We conclude that the side chains of these invariant residues play a crucial role in the response to 1O2. Additionally, we found that cells containing variant ChrR proteins with single amino acid substitutions at Cys187 or Cys189 exhibited σE activity similar to those containing wild-type ChrR when exposed to 1O2, suggesting that these thiol side chains are not required for 1O2 to induce σE activity in vivo. Finally, we found that the same aspects of R. sphaeroides ChrR needed for a response to 1O2 are required for dissociation of σE/ChrR in the presence of the organic hydroperoxide, tert-butyl hydroperoxide (t-BOOH). PMID:21295582

Dye-ligand affinity systems.

PubMed

Denizli, A; Pişkin, E

2001-10-30

Dye-ligands have been considered as one of the important alternatives to natural counterparts for specific affinity chromatography. Dye-ligands are able to bind most types of proteins, in some cases in a remarkably specific manner. They are commercially available, inexpensive, and can easily be immobilized, especially on matrices bearing hydroxyl groups. Although dyes are all synthetic in nature, they are still classified as affinity ligands because they interact with the active sites of many proteins mimicking the structure of the substrates, cofactors, or binding agents for those proteins. A number of textile dyes, known as reactive dyes, have been used for protein purification. Most of these reactive dyes consist of a chromophore (either azo dyes, anthraquinone, or phathalocyanine), linked to a reactive group (often a mono- or dichlorotriazine ring). The interaction between the dye ligand and proteins can be by complex combination of electrostatic, hydrophobic, hydrogen bonding. Selection of the supporting matrix is the first important consideration in dye-affinity systems. There are several methods for immobilization of dye molecules onto the support matrix, in which usually several intermediate steps are followed. Both the adsorption and elution steps should carefully be optimized/designed for a successful separation. Dye-affinity systems in the form of spherical sorbents or as affinity membranes have been used in protein separation.

Assessment of dissociation in Bosnian treatment-seeking refugees in Denmark.

PubMed

Palic, Sabina; Carlsson, Jessica; Armour, Cherie; Elklit, Ask

2015-05-01

Dissociative experiences are common in traumatized individuals, and can sometimes be mistaken for psychosis. It is difficult to identify pathological dissociation in the treatment of traumatized refugees, because there is a lack of systematic clinical descriptions of dissociative phenomena in refugees. Furthermore, we are currently unaware of how dissociation measures perform in this clinical group. To describe the phenomenology of dissociative symptoms in Bosnian treatment-seeking refugees in Denmark. As a part of a larger study, dissociation was assessed systematically in 86 Bosnian treatment-seeking refugees using a semi-structured clinical interview (Structured Interview for Disorders of Extreme Stress-dissociation subscale; SIDES-D) and a self-report scale (Dissociative Experiences Scale; DES). The SIDES-D indicated twice as high prevalence of pathological dissociation as the DES. According to the DES, 30% of the refugees had pathological dissociation 15 years after their resettlement. On the SIDES-D, depersonalization and derealization experiences were the most common. Also, questions about depersonalization and derealization at times elicited reporting of visual and perceptual hallucinations, which were unrelated to traumatic re-experiencing. Questions about personality alteration elicited spontaneous reports of a phenomenon of "split" pre- and post-war identity in the refugee group. Whether this in fact is a dissociative phenomenon, characteristic of severe traumatization in adulthood, needs further examination. Knowledge of dissociative symptoms in traumatized refugees is important in clinical settings to prevent misclassification and to better target psychotherapeutic interventions. Much development in the measurement of dissociation in refugees is needed.

Cobalt porphyrin-mediated oxygen transport in a polymer membrane. Effect of the cobalt porphyrin structure on the oxygen-binding reaction, oxygen-diffusion constants, and oxygen-transport efficiency

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nishide, Hiroyuki; Suzuki, Takayuki; Kawakami, Hiroyoshi

1994-05-12

New derivatives of (meso-[alpha],[alpha],[alpha],[alpha]-tetrakis(o-pivalamidophenyl)porphinato)cobalt (CoPs) were characterized by oxygen-binding equilibrium and rate constants of the cobalt centered in the porphyrins. They depended on the structure of the porphyrin; for example, the rate constants of oxygen binding and dissociation (k[sub on] and k[sub off]) for [alpha][sup 3][beta]-CoP[sub 4]P were 3 and 20 times as large as those for [alpha][sup 4]-CoB[sub 4]P, respectively. Oxygen transport through the polymer membranes containing CoPs as the fixed oxygen carriers was facilitated and was affected by the oxygen-binding character or the structure of CoPs. The logarithmically linear correlation of the oxygen-dissociation rate constant of CoPs (k[submore » off] = (3-66) x 10[sup 3] S[sup [minus]1]) with the diffusion constant of oxygen via CoPs fixed in the membranes (D[sub cc] = (3-140) x 10[sup [minus]9] cm[sup 2] s[sup [minus]1]) was given for those six CoP derivatives. 26 refs., 5 figs., 2 tabs.« less

People reporting experiences of mediumship have higher dissociation symptom scores than non-mediums, but below thresholds for pathological dissociation

PubMed Central

Wahbeh, Helané; Radin, Dean

2018-01-01

Background: Dissociative states exist on a continuum from nonpathological forms, such as highway hypnosis and day-dreaming, to pathological states of derealization and depersonalization. Claims of communication with deceased individuals, known as mediumship, were once regarded as a pathological form of dissociation, but current definitions recognize the continuum and include distress and functional disability as symptoms of pathology. This study examined the relationship between dissociative symptoms and mediumship in a large convenience sample. Methods: Secondary analyses of cross-sectional survey data were conducted. The survey included demographics, the Dissociation Experience Scale Taxon (DES-T, score range 0-100), as well as questions about instances of mediumship experiences. Summary statistics and linear and logistic regressions explored the relationship between dissociative symptoms and mediumship endorsement. Results: 3,023 participants were included and were mostly middle-aged (51 years ± 16; range 17-96), female (70%), Caucasian (85%), college educated (88%), had an annual income over $50,000 (55%), and were raised Christian (71%) but were presently described as Spiritual but not Religious (60%). Mediumship experiences were endorsed by 42% of participants, the experiences usually began in childhood (81%), and 53% had family members who reported similar experiences. The mean DES-T score across all participants was 14.4 ± 17.3, with a mean of 18.2 ± 19.3 for those claiming mediumship experiences and 11.8 ± 15.2 for those who did not (t = -10.3, p < 0.0005). The DES-T threshold score for pathological dissociation is 30. Conclusions: On average, individuals claiming mediumship experiences had higher dissociation scores than non-claimants, but neither group exceeded the DES-T threshold for pathology. Future studies exploring dissociative differences between these groups may benefit from using more comprehensive measures of dissociative symptoms as well as

Toxic metals (Ni2+, Pb2+, Hg2+) binding affinity of dissolved organic matter (DOM) derived from different ages municipal landfill leachate

NASA Astrophysics Data System (ADS)

Rikta, S. Y.; Tareq, Shafi M.; Uddin, M. Khabir

2018-03-01

Solid waste production is rapidly increasing in Bangladesh and landfill leachate is the consequence of the decomposition of this waste. These leachates contain heavy metals and significant amount of dissolved organic matter (DOM). DOM is known to have considerable role in heavy metals speciation. Hence, it is important to characterize DOM/leachate and evaluate toxic metals binding affinity of DOM. The objectives of this study were to characterize the DOM in landfill leachate through physico-chemical and optical analyses and to investigate the toxic metals (Ni2+, Pb2+ and Hg2+) binding affinity of three different ages (fresh sample L-1, young sample L-2 and mature sample L-3) DOM samples. Results suggested that leachate is a potential pollutant which contained very high organic pollutant load. Conditional stability constant (Log K) and percentages of fluorophores that correspond to metal binding (% f) values indicated that young DOM sample (L-2) had the highest binding affinity to all the three metals ions. In general, DOM samples showed the following order affinity to the metal ions; Ni2+ binding affinity: L-2 > L-3 > L-1, Pb2+ binding affinity: L-2 > L-3 > L-1 and Hg2+ binding affinity: L-2 > L-1 > L-3.

The Parenting Experiences of Mothers with Dissociative Disorders.

ERIC Educational Resources Information Center

Benjamin, Lynn R.; Benjamin, Robert; Rind, Bruce

1998-01-01

Presents a qualitative analysis of the experience of parenting of mothers with dissociative disorders. Using the mothers' words, describes how the five symptom areas of dissociation impeded their parenting efforts. Discusses the necessity of addressing parenting in the treatment of client-mothers with dissociative disorders. (Author/MKA)

Childhood Trauma and Dissociative Intimate Partner Violence.

PubMed

Webermann, Aliya R; Murphy, Christopher M

2018-04-01

The present study assesses childhood abuse/neglect as a predictor of dissociative intimate partner violence (IPV) among 118 partner-abusive men. One third (36%) endorsed dissociative IPV, most commonly losing control (18%), surroundings seeming unreal (16%), feeling someone other than oneself is aggressing (16%), and seeing oneself from a distance aggressing (10%). Childhood physical abuse/neglect predicted IPV-specific derealization/depersonalization, aggressive self-states, and flashbacks to past violence. Childhood emotional abuse/neglect predicted derealization/depersonalization, blackouts, and flashbacks. Childhood sexual abuse uniquely predicted amnesia. Other potential traumas did not predict dissociative IPV, suggesting dissociative IPV is influenced by trauma-based emotion dysregulation wherein childhood abuse/neglect survivors disconnect from their abusive behavior.

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

Affinity-aware checkpoint restart

DOE Office of Scientific and Technical Information (OSTI.GOV)

Saini, Ajay; Rezaei, Arash; Mueller, Frank