Acid-base properties of brown seaweed biomass considered as a Donnan gel. A model reflecting electrostatic effects and chemical heterogeneity.

PubMed

Rey-Castro, Carlos; Lodeiro, Pablo; Herrero, Roberto; Sastre de Vicente, Manuel E

2003-11-15

Brown seaweeds are interesting materials to be used as biosorbents for heavy metals due to their high binding ability and low cost. The study of the passive biosorption of protons on this kind of materials and its dependency on pH, ionic strength, and medium composition is essential for the practical application of brown algae in wastewater treatment. This work reports the results of the study of the proton binding equilibria of dead biomass from the seaweeds Sargassum muticum, Cystoseira baccata, and Saccorhiza polyschides by potentiometric titration with a glass electrode in the pH range between 2 and 8. Two different salts, NaCl and KNO3, in concentrations ranging from 0.05 to 2 mol x L(-1), were used as background electrolytes. The influence of the ionic strength was accounted for by means of the Donnan model in combination with the master curve approach. Different empirical expressions to describe the swelling behavior of the biosorbent were tested. On the basis of the intrinsic affinity distribution analysis a unimodal Langmuir-Freundlich isotherm was selected to describe the proton binding properties. The results show very little influence of the type of salt. The ionic strength dependency of the proton binding is very similar for the three species, and average empirical expressions of the Donnan volume are proposed. The maximum proton binding capacities obtained ranged between 2.4 and 2.9 mol x kg(-1), with average intrinsic proton affinity constants between 3.1 and 3.3, and heterogeneity parameters of ca. 0.5 for S. muticum and C. baccata, and slightly higher (ca. 0.7) for S. polyschides. The combined Langmuir-Freundlich equation and Donnan model allowed a good description of the experimental charge vs pH curves obtained.

Kinetics of proton uptake and dye binding by photoactive yellow protein in wild type and in the E46Q and E46A mutants.

PubMed

Borucki, Berthold; Devanathan, Savitha; Otto, Harald; Cusanovich, Michael A; Tollin, Gordon; Heyn, Maarten P

2002-08-06

We studied the kinetics of proton uptake and release by photoactive yellow protein (PYP) from Ectothiorhodospira halophila in wild type and the E46Q and E46A mutants by transient absorption spectroscopy with the pH-indicator dyes bromocresol purple or cresol red in unbuffered solution. In parallel, we investigated the kinetics of chromophore protonation as monitored by the rise and decay of the blue-shifted state I(2) (lambda(max) = 355 nm). For wild type the proton uptake kinetics is synchronized with the fast phase of I(2) formation (tau = 500 micros at pH 6.2). The transient absorption signal from the dye also contains a slower component which is not due to dye deprotonation but is caused by dye binding to a hydrophobic patch that is transiently exposed in the structurally changed and partially unfolded I(2) intermediate. This conclusion is based on the wavelength, pH, and concentration dependence of the dye signal and on dye measurements in the presence of buffer. SVD analysis, moreover, indicates the presence of two components in the dye signal: protonation and dye binding. The dye binding has a rise time of about 4 ms and is coupled kinetically with a transition between two I(2) intermediates. In the mutant E46Q, which lacks the putative internal proton donor E46, the formation of I(2) is accelerated, but the proton uptake kinetics remains kinetically coupled to the fast phase of I(2) formation (tau = 100 micros at pH 6.3). For this mutant the protein conformational change, as monitored by the dye binding, occurs with about the same time constant as in wild type but with reduced amplitude. In the alkaline form of the mutant E46A the formation of the I(2)-like intermediate is even faster as is the proton uptake (tau = 20 micros at pH 8.3). No dye binding occurred in E46A, suggesting the absence of a conformational change. In all of the systems proton release is synchronized with the decay of I(2). Our results support mechanisms in which the chromophore of PYP is protonated directly from the external medium rather than by the internal donor E46.

Semi-empirical proton binding constants for natural organic matter

NASA Astrophysics Data System (ADS)

Matynia, Anthony; Lenoir, Thomas; Causse, Benjamin; Spadini, Lorenzo; Jacquet, Thierry; Manceau, Alain

2010-03-01

Average proton binding constants ( KH,i) for structure models of humic (HA) and fulvic (FA) acids were estimated semi-empirically by breaking down the macromolecules into reactive structural units (RSUs), and calculating KH,i values of the RSUs using linear free energy relationships (LFER) of Hammett. Predicted log KH,COOH and log KH,Ph-OH are 3.73 ± 0.13 and 9.83 ± 0.23 for HA, and 3.80 ± 0.20 and 9.87 ± 0.31 for FA. The predicted constants for phenolic-type sites (Ph-OH) are generally higher than those derived from potentiometric titrations, but the difference may not be significant in view of the considerable uncertainty of the acidity constants determined from acid-base measurements at high pH. The predicted constants for carboxylic-type sites agree well with titration data analyzed with Model VI (4.10 ± 0.16 for HA, 3.20 ± 0.13 for FA; Tipping, 1998), the Impermeable Sphere model (3.50-4.50 for HA; Avena et al., 1999), and the Stockholm Humic Model (4.10 ± 0.20 for HA, 3.50 ± 0.40 for FA; Gustafsson, 2001), but differ by about one log unit from those obtained by Milne et al. (2001) with the NICA-Donnan model (3.09 ± 0.51 for HA, 2.65 ± 0.43 for FA), and used to derive recommended generic values. To clarify this ambiguity, 10 high-quality titration data from Milne et al. (2001) were re-analyzed with the new predicted equilibrium constants. The data are described equally well with the previous and new sets of values ( R2 ⩾ 0.98), not necessarily because the NICA-Donnan model is overparametrized, but because titration lacks the sensitivity needed to quantify the full binding properties of humic substances. Correlations between NICA-Donnan parameters are discussed, but general progress is impeded by the unknown number of independent parameters that can be varied during regression of a model fit to titration data. The high consistency between predicted and experimental KH,COOH values, excluding those of Milne et al. (2001), gives faith in the proposed semi-empirical structural approach, and its usefulness to assess the plausibility of proton stability constants derived from simulations of titration data.

Involvement of histidine residues in the pH-dependent β-galactoside binding activity of human galectin-1.

PubMed

Hiramatsu, Hirotsugu; Takeuchi, Katsuyuki; Takeuchi, Hideo

2013-04-02

The pH dependence of the β-galactoside binding activity of human galectin-1 (hGal-1) was investigated by fluorescence spectroscopy using lactose as a ligand. The obtained binding constant Kb was 2.94 ± 0.10 mM(-1) at pH 7.5. The Kb value decreased at acidic pH with a midpoint of transition at pH 6.0 ± 0.1. To elucidate the molecular mechanism of the pH dependence, we investigated the structures of hGal-1 and its two His mutants (H44Q and H52Q) using fluorescence, circular dichroism, UV absorption, and UV resonance Raman spectroscopy. Analysis of the spectra has shown that the pKa values of His44 and His52 are 5.7 ± 0.2 and 6.3 ± 0.1, respectively. The protonation of His52 below pH 6.3 induces a small change in secondary structure and partly reduces the galactoside binding activity. On the other hand, the protonation of His44 below pH 5.7 exerts a cation-π interaction with Trp68 and largely diminishes the galactoside binding activity. With reference to the literature X-ray structures at pH 7.0 and 5.6, protonated His52 is proposed to move slightly away from the galactoside-binding region with a partial unfolding of the β-strand containing His52. On the other hand, protonated His44 becomes unable to form a hydrogen bond with galactoside and additionally induces a reorientation and/or displacement of Trp68 through cation-π interaction, leading to a loosening of the galactoside-binding pocket. These structural changes associated with His protonation are likely to be the origin of the pH dependence of the galactoside binding activity of hGal-1.

Protonation linked equilibria and apparent affinity constants: the thermodynamic profile of the alpha-chymotrypsin-proflavin interaction.

PubMed

Bruylants, Gilles; Wintjens, René; Looze, Yvan; Redfield, Christina; Bartik, Kristin

2007-12-01

Protonation/deprotonation equilibria are frequently linked to binding processes involving proteins. The presence of these thermodynamically linked equilibria affects the observable thermodynamic parameters of the interaction (K(obs), DeltaH(obs)(0) ). In order to try and elucidate the energetic factors that govern these binding processes, a complete thermodynamic characterisation of each intrinsic equilibrium linked to the complexation event is needed and should furthermore be correlated to structural information. We present here a detailed study, using NMR and ITC, of the interaction between alpha-chymotrypsin and one of its competitive inhibitors, proflavin. By performing proflavin titrations of the enzyme, at different pH values, we were able to highlight by NMR the effect of the complexation of the inhibitor on the ionisable residues of the catalytic triad of the enzyme. Using ITC we determined the intrinsic thermodynamic parameters of the different equilibria linked to the binding process. The possible driving forces of the interaction between alpha-chymotrypsin and proflavin are discussed in the light of the experimental data and on the basis of a model of the complex. This study emphasises the complementarities between ITC and NMR for the study of binding processes involving protonation/deprotonation equilibria.

Sequential Proton Loss Electron Transfer in Deactivation of Iron(IV) Binding Protein by Tyrosine Based Food Components.

PubMed

Tang, Ning; Skibsted, Leif H

2017-08-02

The iron(IV) binding protein ferrylmyoglobin, MbFe(IV)═O, was found to be reduced by tyrosine based food components in aqueous solution through a sequential proton loss electron transfer reaction mechanism without binding to the protein as confirmed by isothermal titration calorimetry. Dopamine and epinephrine are the most efficient food components reducing ferrylmyoglobin to oxymyoglobin, MbFe(II)O 2 , and metmyoglobin, MbFe(III), as revealed by multivariate curve resolution alternating least-squares with second order rate constants of 33.6 ± 2.3 L/mol/s (ΔH ⧧ of 19 ± 5 kJ/mol, ΔS ⧧ of -136 ± 18 J/mol K) and 228.9 ± 13.3 L/mol/s (ΔH ⧧ of 110 ± 7 kJ/mol, ΔS ⧧ of 131 ± 25 J/mol K), respectively, at pH 7.4 and 25 °C. The other tyrosine based food components were found to reduce ferrylmyoglobin to metmyoglobin with similar reduction rates at pH 7.4 and 25 °C. These reduction reactions were enhanced by protonation of ferrylmyoglobin and facilitated proton transfer at acidic conditions. Enthalpy-entropy compensation effects were observed for the activation parameters (ΔH ⧧ and ΔS ⧧ ), indicating the common reaction mechanism. Moreover, principal component analysis combined with heat map were performed to understand the relationship between density functional theory calculated molecular descriptors and kinetic data, which was further modeled by partial least squares for quantitative structure-activity relationship analysis. In addition, a three tyrosine residue containing protein, lysozyme, was also found to be able to reduce ferrylmyoglobin with a second order rate constant of 66 ± 28 L/mol/s as determined by a competitive kinetic method.

A spectroscopic study of phenylbutazone and aspirin bound to serum albumin in rheumatoid diseases

NASA Astrophysics Data System (ADS)

Maciążek-Jurczyk, M.; Sułkowska, A.; Bojko, B.; Równicka-Zubik, J.; Sułkowski, W. W.

2011-11-01

Interaction of phenylbutazone (PBZ) and aspirin (ASA), two drugs recommended in rheumatoid diseases (RDs), when binding to human (HSA) and bovine (BSA) serum albumins, has been studied by quenching of fluorescence and proton nuclear magnetic resonance ( 1HNMR) techniques. On the basis of spectrofluorescence measurements high affinity binding sites of PBZ and ASA on albumin as well as their interaction within the binding sites were described. A low affinity binding site has been studied by proton nuclear magnetic resonance spectroscopy. Using fluorescence spectroscopy the location of binding site in serum albumin (SA) for PBZ and ASA was found. Association constants Ka were determined for binary (i.e. PBZ-SA and ASA-SA) and ternary complexes (i.e. PBZ-[ASA]-SA and ASA-[PBZ]-SA). PBZ and ASA change the affinity of each other to the binding site in serum albumin (SA). The presence of ASA causes the increase of association constants KaI of PBZ-SA complex. Similarly, PBZ influences KaI of ASA-SA complex. This phenomenon shows that the strength of binding and the stability of the complexes increase in the presence of the second drug. The decrease of KaII values suggests that the competition between PBZ and ASA in binding to serum albumin in the second class of binding sites occurs. The analysis of 1HNMR spectral parameters i.e. changes of chemical shifts and relaxation times of the drug indicate that the presence of ASA weakens the interaction of PBZ with albumin. Similarly PBZ weakens the interaction of ASA with albumin. This conclusion points to the necessity of using a monitoring therapy owning to the possible increase of uncontrolled toxic effects.

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.

Residues in the H+ Translocation Site Define the pKa for Sugar Binding to LacY†

PubMed Central

Smirnova, Irina; Kasho, Vladimir; Sugihara, Junichi; Choe, Jun-Yong; Kaback, H. Ronald

2009-01-01

A remarkably high pKa of approximately 10.5 has been determined for sugar-binding affinity to the lactose permease of Escherichia coli (LacY), indicating that, under physiological conditions, substrate binds to fully protonated LacY. We have now systematically tested site-directed replacements for the residues involved in sugar binding, as well as H+ translocation and coupling, in order to determine which residues may be responsible for this alkaline pKa. Mutations in the sugar-binding site (Glu126, Trp151, Glu269) markedly decrease affinity for sugar but do not alter the pKa for binding. In contrast, replacements for residues involved in H+ translocation (Arg302, Tyr236, His322, Asp240, Glu325, Lys319) exhibit pKa values for sugar binding that are either shifted toward neutral pH or independent of pH. Values for the apparent dissociation constant for sugar binding (Kdapp) increase greatly for all mutants except neutral replacements for Glu325 or Lys319, which are characterized by remarkably high affinity sugar binding (i.e., low Kdapp) from pH 5.5 to pH 11. The pH dependence of the on- and off-rate constants for sugar binding measured directly by stopped-flow fluorometry implicates koff as a major factor for the affinity change at alkaline pH and confirms the effects of pH on Kdapp inferred from steady-state fluorometry. These results indicate that the high pKa for sugar binding by wild-type LacY cannot be ascribed to any single amino acid residue but appears to reside within a complex of residues involved in H+ translocation. There is structural evidence for water bound in this complex, and the water could be the site of protonation responsible for the pH dependence of sugar binding. PMID:19689129

Acid-base properties of humic substances from composted and thermally-dried sewage sludges and amended soils as determined by potentiometric titration and the NICA-Donnan model.

PubMed

Fernández, José M; Plaza, César; Senesi, Nicola; Polo, Alfredo

2007-09-01

The acid-base properties of humic acids (HAs) and fulvic acids (FAs) isolated from composted sewage sludge (CS), thermally-dried sewage sludge (TS), soils amended with either CS or TS at a rate of 80 t ha(-1)y(-1) for 3y and the corresponding unamended soil were investigated by use of potentiometric titrations. The non-ideal competitive adsorption (NICA)-Donnan model for a bimodal distribution of proton binding sites was fitted to titration data by use of a least-squares minimization method. The main fitting parameters of the NICA-Donnan model obtained for each HA and FA sample included site densities, median affinity constants and widths of affinity distributions for proton binding to low and high affinity sites, which were assumed to be, respectively, carboxylic- and phenolic-type groups. With respect to unamended soil HA and FA, the HAs and FAs from CS, and especially TS, were characterized by smaller acidic functional group contents, larger proton binding affinities of both carboxylic- and phenolic-type groups, and smaller heterogeneity of carboxylic and phenolic-type groups. Amendment with CS or TS led to a decrease of acidic functional group contents and a slight increase of proton binding affinities of carboxylic- and phenolic-type groups of soil HAs and FAs. These effects were more evident in the HA and FA fractions from CS-amended soil than in those from TS-amended soil.

The interaction of an ionizing ligand with enzymes having a single ionizing group. Implications for the reaction of folate analogues with dihydrofolate reductase.

PubMed

Stone, S R; Morrison, J F

1983-06-29

Binding theory has been developed for the reaction of an ionizing enzyme with an ionizing ligand. Consideration has been given to the most general scheme in which all possible reactions and interconversions occur as well as to schemes in which certain interactions do not take place. Equations have been derived in terms of the variation of the apparent dissociation constant (Kiapp) as a function of pH. These equations indicate that plots of pKiapp against pH can be wave-, half-bell- or bell-shaped according to the reactions involved. A wave is obtained whenever there is formation of the enzyme-ligand complexes, ionized enzyme . ionized ligand and protonated enzyme . protonated ligand. The additional formation of singly protonated enzyme-ligand complexes does not affect the wave form of the plot, but can influence the shape of the overall curve. The formation of either ionized enzyme . ionized ligand or protonated enzyme . protonated ligand, with or without singly protonated enzyme-ligand species, gives rise to a half-bell-shaped plot. If only singly protonated enzyme-ligand complexes are formed the plots are bell-shaped, but it is not possible to deduce the ionic forms of the reactants that participate in complex formation. Depending on the reaction pathways, true values for the ionization and dissociation constants may or may not be determined.

Eu(III) complexes as Anion-responsive Luminescent Sensors and PARACEST Agents

PubMed Central

Hammell, Jacob; Buttarazzi, Leandro; Huang, Ching-Hui; Morrow, Janet R.

2011-01-01

The Eu(III) complex of (1S,4S,7S,10S)-1,4,7,10-tetrakis(2-hydroxypropyl)-1,4,7,10-tetraazacyclododecane (S-THP) is studied as a sensor for biologically relevant anions. Anion interactions produce changes in the luminescence emission spectrum of the Eu(III) complex, in the 1H NMR spectrum, and correspondingly, in the PARACEST spectrum of the complex (PARACEST = paramagnetic chemical exchange saturation transfer). Direct excitation spectroscopy and luminescence lifetime studies of Eu(S-THP) give information about the speciation and nature of anion interactions including carbonate, acetate, lactate, citrate, phosphate and methylphosphate at pH 7.2. Data is consistent with the formation of both innersphere and outersphere complexes of Eu(S-THP) with acetate, lactate and carbonate. These anions have weak dissociation constants that range from 19–38 mM. Citrate binding to Eu(S-THP) is predominantly innersphere with a dissociation constant of 17 μM. Luminescence emission peak changes upon addition of anion to Eu(S-THP) show that there are two distinct binding events for phosphate and methylphosphate with dissociation constants of 0.3 mM and 3.0 mM for phosphate and 0.6 mM and 9.8 mM for methyl phosphate. Eu(THPC) contains an appended carbostyril derivative as an antenna to sensitize Eu(III) luminescence. Eu(THPC) binds phosphate and citrate with dissociation constants that are 10-fold less than that of the Eu(S-THP) parent, suggesting that functionalization through a pendent group disrupts the anion binding site. Eu(S-THP) functions as an anion responsive PARACEST agent through exchange of the alcohol protons with bulk water. The alcohol proton resonances of Eu(S-THP) shift downfield in the presence of acetate, lactate, citrate and methylphosphate, giving rise to distinct PARACEST peaks. In contrast, phosphate binds to Eu(S-THP) to suppress the PARACEST alcohol OH peak and carbonate does not markedly change the alcohol peak at 5 mM Eu(S-THP), 15 mM carbonate at pH 6.5 or 7.2. This work shows that the Eu(S-THP) complex has unique selectivity toward binding of biologically relevant anions and that anion binding results in changes in both the luminescence and PARACEST spectra of the complex. PMID:21548563

Eu(III) complexes as anion-responsive luminescent sensors and paramagnetic chemical exchange saturation transfer agents.

PubMed

Hammell, Jacob; Buttarazzi, Leandro; Huang, Ching-Hui; Morrow, Janet R

2011-06-06

The Eu(III) complex of (1S,4S,7S,10S)-1,4,7,10-tetrakis(2-hydroxypropyl)-1,4,7,10-tetraazacyclododecane (S-THP) is studied as a sensor for biologically relevant anions. Anion interactions produce changes in the luminescence emission spectrum of the Eu(III) complex, in the (1)H NMR spectrum, and correspondingly, in the PARACEST spectrum of the complex (PARACEST = paramagnetic chemical exchange saturation transfer). Direct excitation spectroscopy and luminescence lifetime studies of Eu(S-THP) give information about the speciation and nature of anion interactions including carbonate, acetate, lactate, citrate, phosphate, and methylphosphate at pH 7.2. Data is consistent with the formation of both innersphere and outersphere complexes of Eu(S-THP) with acetate, lactate, and carbonate. These anions have weak dissociation constants that range from 19 to 38 mM. Citrate binding to Eu(S-THP) is predominantly innersphere with a dissociation constant of 17 μM. Luminescence emission peak changes upon addition of anion to Eu(S-THP) show that there are two distinct binding events for phosphate and methylphosphate with dissociation constants of 0.3 mM and 3.0 mM for phosphate and 0.6 mM and 9.8 mM for methyl phosphate. Eu(THPC) contains an appended carbostyril derivative as an antenna to sensitize Eu(III) luminescence. Eu(THPC) binds phosphate and citrate with dissociation constants that are 10-fold less than that of the Eu(S-THP) parent, suggesting that functionalization through a pendent group disrupts the anion binding site. Eu(S-THP) functions as an anion responsive PARACEST agent through exchange of the alcohol protons with bulk water. The alcohol proton resonances of Eu(S-THP) shift downfield in the presence of acetate, lactate, citrate, and methylphosphate, giving rise to distinct PARACEST peaks. In contrast, phosphate binds to Eu(S-THP) to suppress the PARACEST alcohol OH peak and carbonate does not markedly change the alcohol peak at 5 mM Eu(S-THP), 15 mM carbonate at pH 6.5 or 7.2. This work shows that the Eu(S-THP) complex has unique selectivity toward binding of biologically relevant anions and that anion binding results in changes in both the luminescence and the PARACEST spectra of the complex. © 2011 American Chemical Society

Chemical modeling of acid-base properties of soluble biopolymers derived from municipal waste treatment materials.

PubMed

Tabasso, Silvia; Berto, Silvia; Rosato, Roberta; Marinos, Janeth Alicia Tafur; Ginepro, Marco; Zelano, Vincenzo; Daniele, Pier Giuseppe; Montoneri, Enzo

2015-02-04

This work reports a study of the proton-binding capacity of biopolymers obtained from different materials supplied by a municipal biowaste treatment plant located in Northern Italy. One material was the anaerobic fermentation digestate of the urban wastes organic humid fraction. The others were the compost of home and public gardening residues and the compost of the mix of the above residues, digestate and sewage sludge. These materials were hydrolyzed under alkaline conditions to yield the biopolymers by saponification. The biopolymers were characterized by 13C NMR spectroscopy, elemental analysis and potentiometric titration. The titration data were elaborated to attain chemical models for interpretation of the proton-binding capacity of the biopolymers obtaining the acidic sites concentrations and their protonation constants. The results obtained with the models and by NMR spectroscopy were elaborated together in order to better characterize the nature of the macromolecules. The chemical nature of the biopolymers was found dependent upon the nature of the sourcing materials.

Chemical Modeling of Acid-Base Properties of Soluble Biopolymers Derived from Municipal Waste Treatment Materials

PubMed Central

Tabasso, Silvia; Berto, Silvia; Rosato, Roberta; Tafur Marinos, Janeth Alicia; Ginepro, Marco; Zelano, Vincenzo; Daniele, Pier Giuseppe; Montoneri, Enzo

2015-01-01

This work reports a study of the proton-binding capacity of biopolymers obtained from different materials supplied by a municipal biowaste treatment plant located in Northern Italy. One material was the anaerobic fermentation digestate of the urban wastes organic humid fraction. The others were the compost of home and public gardening residues and the compost of the mix of the above residues, digestate and sewage sludge. These materials were hydrolyzed under alkaline conditions to yield the biopolymers by saponification. The biopolymers were characterized by 13C NMR spectroscopy, elemental analysis and potentiometric titration. The titration data were elaborated to attain chemical models for interpretation of the proton-binding capacity of the biopolymers obtaining the acidic sites concentrations and their protonation constants. The results obtained with the models and by NMR spectroscopy were elaborated together in order to better characterize the nature of the macromolecules. The chemical nature of the biopolymers was found dependent upon the nature of the sourcing materials. PMID:25658795

Mechanism of pH-dependent activation of the sodium-proton antiporter NhaA

NASA Astrophysics Data System (ADS)

Huang, Yandong; Chen, Wei; Dotson, David L.; Beckstein, Oliver; Shen, Jana

2016-10-01

Escherichia coli NhaA is a prototype sodium-proton antiporter, which has been extensively characterized by X-ray crystallography, biochemical and biophysical experiments. However, the identities of proton carriers and details of pH-regulated mechanism remain controversial. Here we report constant pH molecular dynamics data, which reveal that NhaA activation involves a net charge switch of a pH sensor at the entrance of the cytoplasmic funnel and opening of a hydrophobic gate at the end of the funnel. The latter is triggered by charging of Asp164, the first proton carrier. The second proton carrier Lys300 forms a salt bridge with Asp163 in the inactive state, and releases a proton when a sodium ion binds Asp163. These data reconcile current models and illustrate the power of state-of-the-art molecular dynamics simulations in providing atomic details of proton-coupled transport across membrane which is challenging to elucidate by experimental techniques.

Turn on ESPT: novel salicylaldehyde based sensor for biological important fluoride sensing.

PubMed

Liu, Kai; Zhao, Xiaojun; Liu, Qingxiang; Huo, Jianzhong; Fu, Huifang; Wang, Ying

2014-09-05

A novel and simple salicylaldehyde based anion fluorescent sensor 1 has been designed, which can selectively sense fluoride by 'turn on' excited-state intermolecular proton transfer (ESPT). The binding constant and the stoichiometry were obtained by non-linear least-square analysis of the titration curves. Copyright © 2014 Elsevier B.V. All rights reserved.

Striking Confinement Effect: AuCl[subscript 4][superscript -] Binding to Amines in a Nanocage Cavity

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

Henao, Juan D.; Suh, Young-Woong; Lee, Jeong-Kyu

2009-02-23

Binding of AuCl{sub 4}{sup -} to amine groups tethered to the interior of a 2 nm siloxane nanocage was determined in solutions containing various concentrations of acid. The mode of binding was inferred from EXAFS and UV-vis spectra to be by ligand exchange of amine for chloride, which implies that the amines remain unprotonated. Cyclic voltammetry confirmed that the Au complexes bind to the nanocage interior and established a 1:1 relationship between bound Au complex and amine groups. The results suggested a 5-7 pH unit shift in the protonation constant of the interior amines relative to free amines in solution.

Metal binding stoichiometry and isotherm choice in biosorption

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

Schiewer, S.; Wong, M.H.

1999-11-01

Seaweeds that possess a high metal binding capacity may be used as biosorbents for the removal of toxic heavy metals from wastewater. The binding of Cu and Ni by three brown algae (Sargassum, Colpomenia, Petalonia) and one green alga (Ulva) was investigated at pH 4.0 and pH 3.0. The greater binding strength of Cu is reflected in a binding constant that is about 10 times as high as that of Ni. The extent of metal binding followed the order Petalonia {approximately} Sargassum > Colpomenia > Ulva. This was caused by a decreasing number of binding sites and by much lowermore » metal binding constants for Ulva as compared to the brown algae. Three different stoichiometric assumptions are compared for describing the metal binding, which assume either that each metal ion M binds to one binding site B forming a BM complex or that a divalent metal ion M binds to two monovalent sites B forming BM{sub 0.5} or B{sub 2}M complexes, respectively. Stoichiometry plots are proposed as tools to discern the relevant binding stoichiometry. The pH effect in metal binding and the change in proton binding were well predicted for the B{sub 2}M or BM{sub 0.5} stoichiometries with the former being better for Cu and the latter preferable for Ni. Overall, the BM{sub 0.5} model is recommended because it avoids iterations.« less

Number of independent parameters in the potentiometric titration of humic substances.

PubMed

Lenoir, Thomas; Manceau, Alain

2010-03-16

With the advent of high-precision automatic titrators operating in pH stat mode, measuring the mass balance of protons in solid-solution mixtures against the pH of natural and synthetic polyelectrolytes is now routine. However, titration curves of complex molecules typically lack obvious inflection points, which complicates their analysis despite the high-precision measurements. The calculation of site densities and median proton affinity constants (pK) from such data can lead to considerable covariance between fit parameters. Knowing the number of independent parameters that can be freely varied during the least-squares minimization of a model fit to titration data is necessary to improve the model's applicability. This number was calculated for natural organic matter by applying principal component analysis (PCA) to a reference data set of 47 independent titration curves from fulvic and humic acids measured at I = 0.1 M. The complete data set was reconstructed statistically from pH 3.5 to 9.8 with only six parameters, compared to seven or eight generally adjusted with common semi-empirical speciation models for organic matter, and explains correlations that occur with the higher number of parameters. Existing proton-binding models are not necessarily overparametrized, but instead titration data lack the sensitivity needed to quantify the full set of binding properties of humic materials. Model-independent conditional pK values can be obtained directly from the derivative of titration data, and this approach is the most conservative. The apparent proton-binding constants of the 23 fulvic acids (FA) and 24 humic acids (HA) derived from a high-quality polynomial parametrization of the data set are pK(H,COOH)(FA) = 4.18 +/- 0.21, pK(H,Ph-OH)(FA) = 9.29 +/- 0.33, pK(H,COOH)(HA) = 4.49 +/- 0.18, and pK(H,Ph-OH)(HA) = 9.29 +/- 0.38. Their values at other ionic strengths are more reliably calculated with the empirical Davies equation than any existing model fit.

Proton transfer from C-6 of uridine 5'-monophosphate catalyzed by orotidine 5'-monophosphate decarboxylase: formation and stability of a vinyl carbanion intermediate and the effect of a 5-fluoro substituent.

PubMed

Tsang, Wing-Yin; Wood, B McKay; Wong, Freeman M; Wu, Weiming; Gerlt, John A; Amyes, Tina L; Richard, John P

2012-09-05

The exchange for deuterium of the C-6 protons of uridine 5'-monophosphate (UMP) and 5-fluorouridine 5'-monophosphate (F-UMP) catalyzed by yeast orotidine 5'-monophosphate decarboxylase (ScOMPDC) at pD 6.5-9.3 and 25 °C was monitored by (1)H NMR spectroscopy. Deuterium exchange proceeds by proton transfer from C-6 of the bound nucleotide to the deprotonated side chain of Lys-93 to give the enzyme-bound vinyl carbanion. The pD-rate profiles for k(cat) give turnover numbers for deuterium exchange into enzyme-bound UMP and F-UMP of 1.2 × 10(-5) and 0.041 s(-1), respectively, so that the 5-fluoro substituent results in a 3400-fold increase in the first-order rate constant for deuterium exchange. The binding of UMP and F-UMP to ScOMPDC results in 0.5 and 1.4 unit decreases, respectively, in the pK(a) of the side chain of the catalytic base Lys-93, showing that these nucleotides bind preferentially to the deprotonated enzyme. We also report the first carbon acid pK(a) values for proton transfer from C-6 of uridine (pK(CH) = 28.8) and 5-fluorouridine (pK(CH) = 25.1) in aqueous solution. The stabilizing effects of the 5-fluoro substituent on C-6 carbanion formation in solution (5 kcal/mol) and at ScOMPDC (6 kcal/mol) are similar. The binding of UMP and F-UMP to ScOMPDC results in a greater than 5 × 10(9)-fold increase in the equilibrium constant for proton transfer from C-6, so that ScOMPDC stabilizes the bound vinyl carbanions, relative to the bound nucleotides, by at least 13 kcal/mol. The pD-rate profile for k(cat)/K(m) for deuterium exchange into F-UMP gives the intrinsic second-order rate constant for exchange catalyzed by the deprotonated enzyme as 2300 M(-1) s(-1). This was used to calculate a total rate acceleration for ScOMPDC-catalyzed deuterium exchange of 3 × 10(10) M(-1), which corresponds to a transition-state stabilization for deuterium exchange of 14 kcal/mol. We conclude that a large portion of the total transition-state stabilization for the decarboxylation of orotidine 5'-monophosphate can be accounted for by stabilization of the enzyme-bound vinyl carbanion intermediate of the stepwise reaction.

Proton Transfer from C-6 of Uridine 5′-Monophosphate Catalyzed by Orotidine 5′-Monophosphate Decarboxylase: Formation and Stability of a Vinyl Carbanion Intermediate and the Effect of a 5-Fluoro Substituent

PubMed Central

Tsang, Wing-Yin; Wood, B. McKay; Wong, Freeman M.; Wu, Weiming; Gerlt, John A.; Amyes, Tina L.; Richard, John P.

2012-01-01

The exchange for deuterium of the C-6 protons of uridine 5′-monophosphate (UMP) and 5-fluorouridine 5′-monophosphate (F-UMP) catalyzed by yeast orotidine 5′-monophosphate decarboxylase (ScOMPDC) at pD 6.5 – 9.3 and 25 °C was monitored by 1H NMR spectroscopy. Deuterium exchange proceeds by proton transfer from C-6 of the bound nucleotide to the deprotonated side chain of Lys-93 to give the enzyme-bound vinyl carbanion. The pD-rate profiles for kcat give turnover numbers for deuterium exchange into enzyme-bound UMP and F-UMP of 1.2 × 10−5 and 0.041 s−1, respectively, so that the 5-fluoro substituent results in a 3400-fold increase in the first-order rate constant for deuterium exchange. The binding of UMP and F-UMP to ScOMPDC results in 0.5 and 1.4 unit decreases, respectively, in the pKa of the side chain of the catalytic base Lys-93, showing that these nucleotides bind preferentially to the deprotonated enzyme. We also report the first carbon acid pKas for proton transfer from C-6 of uridine (pKCH = 28.8) and 5-fluorouridine (pKCH = 25.1) in aqueous solution. The stabilizing effects of the 5-fluoro substituent on C-6 carbanion formation in solution (5 kcal/mol) and at ScOMPDC (6 kcal/mol) are similar. The binding of UMP and F-UMP to ScOMPDC results in a greater than 5 × 109-fold increase in the equilibrium constant for proton transfer from C-6 so that ScOMPDC stabilizes the bound vinyl carbanions, relative to the bound nucleotides, by at least 13 kcal/mol. The pD-rate profile for kcat/Km for deuterium exchange into F-UMP gives the intrinsic second-order rate constant for exchange catalyzed by the deprotonated enzyme as 2300 M−1 s−1. This was used to calculate a total rate acceleration for ScOMPDC-catalyzed deuterium exchange of 3 × 1010 M−1, which corresponds to a transition state stabilization for deuterium exchange of 14 kcal/mol. We conclude that a large portion of the total transition state stabilization for the decarboxylation of orotidine 5′-monophosphate can be accounted for by stabilization of the enzyme-bound vinyl carbanion intermediate of the stepwise reaction. PMID:22812629

Rates of proton transfer to Fe-S-based clusters: comparison of clusters containing {MFe(mu(2)-S)(2)}n+ and {MFe(3)(mu(3)-S)(4)}n+ (M = Fe, Mo, or W) cores.

PubMed

Bates, Katie; Garrett, Brendan; Henderson, Richard A

2007-12-24

The rates of proton transfer from [pyrH]+ (pyr = pyrrolidine) to the binuclear complexes [Fe2S2Cl4]2- and [S2MS2FeCl2]2- (M = Mo or W) are reported. The reactions were studied using stopped-flow spectrophotometry, and the rate constants for proton transfer were determined from analysis of the kinetics of the substitution reactions of these clusters with the nucleophiles Br- or PhS- in the presence of [pyrH]+. In general, Br- is a poor nucleophile for these clusters, and proton transfer occurs before Br- binds, allowing direct measure of the rate of proton transfer from [pyrH]+ to the cluster. In contrast, PhS- is a better nucleophile, and a pathway in which PhS- binds preferentially to the cluster prior to proton transfer from [pyrH]+ usually operates. For the reaction of [Fe2S2Cl4]2- with PhS- in the presence of [pyrH]+ both pathways are observed. Comparison of the results presented in this paper with analogous studies reported earlier on cuboidal Fe-S-based clusters allows discussion of the factors which affect the rates of proton transfer in synthetic clusters including the nuclearity of the cluster core, the metal composition, and the nature of the terminal ligands. The possible relevance of these findings to the protonation sites of natural Fe-S-based clusters, including FeMo-cofactor from nitrogenase, are presented.

Comparison of cation adsorption by isostructural rutile and cassiterite.

PubMed

Machesky, Michael; Wesolowski, David; Rosenqvist, Jörgen; Předota, Milan; Vlcek, Lukas; Ridley, Moira; Kohli, Vaibhav; Zhang, Zhan; Fenter, Paul; Cummings, Peter; Lvov, Serguei; Fedkin, Mark; Rodriguez-Santiago, Victor; Kubicki, James; Bandura, Andrei

2011-04-19

Macroscopic net proton charging curves for powdered rutile and cassiterite specimens with the (110) crystal face predominant, as a function of pH in RbCl and NaCl solutions, trace SrCl(2) in NaCl, and trace ZnCl(2) in NaCl and Na Triflate solutions, are compared to corresponding molecular-level information obtained from static DFT optimizations and classical MD simulations, as well as synchrotron X-ray methods. The similarities and differences in the macroscopic charging behavior of rutile and cassiterite largely reflect the cation binding modes observed at the molecular level. Cation adsorption is primarily inner-sphere on both isostructural (110) surfaces, despite predictions that outer-sphere binding should predominate on low bulk dielectric constant oxides such as cassiterite (ε(bulk) ≈ 11). Inner-sphere adsorption is also significant for Rb(+) and Na(+) on neutral surfaces, whereas Cl(-) binding is predominately outer-sphere. As negative surface charge increases, relatively more Rb(+), Na(+), and especially Sr(2+) are bound in highly desolvated tetradentate fashion on the rutile (110) surface, largely accounting for enhanced negative charge development relative to cassiterite. Charging curves in the presence of Zn(2+) are very steep but similar for both oxides, reflective of Zn(2+) hydrolysis (and accompanying proton release) during the adsorption process, and the similar binding modes for ZnOH(+) on both surfaces. These results suggest that differences in cation adsorption between high and low bulk dielectric constant oxides are more subtly related to the relative degree of cation desolvation accompanying inner-sphere binding (i.e., more tetradentate binding on rutile), rather than distinct inner- and outer-sphere adsorption modes. Cation desolvation may be favored at the rutile (110) surface in part because inner-sphere water molecules are bound further from and less tightly than on the cassiterite (110) surface. Hence, their removal upon inner-sphere cation binding is relatively more favorable. © 2011 American Chemical Society

Potentiometric Determination of Phytic Acid and Investigations of Phytate Interactions with Some Metal Ions.

PubMed

Marolt, Gregor; Pihlar, Boris

2015-01-01

Determination of correct amount (concentration) of phytic acid is of vital importance when dealing with protonation and/or metal complexation equilibria. A novel approach for precise and reliable assay of phytic acid, based on the difference between end points by potentiometric titration, has been presented. Twelve phytic acid protons are classified into three groups of acidity, which enables detection of 2 to 3 distinct equivalent points (EPs) depending on experimental conditions, e.g. counter-ion concentration. Using the differences between individual EPs enables correct phytate determination as well as identification of potential contamination and/or determination of initial protonation degree. Impact of uncertainty of phytate amount on the calculation of protonation constants has been evaluated using computer simulation program (Hyperquad2013). With the analysis of titration curves different binding sites on phytate ligand have been proposed for complexation of Ca2+ and Fe3+ ions.

Interactions of poly(amidoamine) dendrimers with human serum albumin: binding constants and mechanisms.

PubMed

Giri, Jyotsnendu; Diallo, Mamadou S; Simpson, André J; Liu, Yi; Goddard, William A; Kumar, Rajeev; Woods, Gwen C

2011-05-24

The interactions of nanomaterials with plasma proteins have a significant impact on their in vivo transport and fate in biological fluids. This article discusses the binding of human serum albumin (HSA) to poly(amidoamine) [PAMAM] dendrimers. We use protein-coated silica particles to measure the HSA binding constants (K(b)) of a homologous series of 19 PAMAM dendrimers in aqueous solutions at physiological pH (7.4) as a function of dendrimer generation, terminal group, and core chemistry. To gain insight into the mechanisms of HSA binding to PAMAM dendrimers, we combined (1)H NMR, saturation transfer difference (STD) NMR, and NMR diffusion ordered spectroscopy (DOSY) of dendrimer-HSA complexes with atomistic molecular dynamics (MD) simulations of dendrimer conformation in aqueous solutions. The binding measurements show that the HSA binding constants (K(b)) of PAMAM dendrimers depend on dendrimer size and terminal group chemistry. The NMR (1)H and DOSY experiments indicate that the interactions between HSA and PAMAM dendrimers are relatively weak. The (1)H NMR STD experiments and MD simulations suggest that the inner shell protons of the dendrimers groups interact more strongly with HSA proteins. These interactions, which are consistently observed for different dendrimer generations (G0-NH(2)vs G4-NH(2)) and terminal groups (G4-NH(2)vs G4-OH with amidoethanol groups), suggest that PAMAM dendrimers adopt backfolded configurations as they form weak complexes with HSA proteins in aqueous solutions at physiological pH (7.4).

Thermodynamic and Spectroscopic Studies of Trivalent f -element Complexation with Ethylenediamine- N,N '-di(acetylglycine)- N,N '-diacetic Acid

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

Heathman, Colt R.; Grimes, Travis S.; Zalupski, Peter R.

In this study, the coordination behavior and thermodynamic features of complexation of trivalent lanthanides and americium by ethylenediamine- N,N'-di(acetylglycine)- N,N'-diacetic acid (EDDAG-DA) (bisamide-substituted-EDTA) were investigated by potentiometric and spectroscopic techniques. Acid dissociation constants (K a) and complexation constants (β) of lanthanides (except Pm) were determined by potentiometric analysis. Absorption spectroscopy was used to determine stability constants for the binding of trivalent americium and neodymium by EDDAG-DA under similar conditions. The potentiometry revealed 5 discernible protonation constants and 3 distinct metal–ligand complexes (identified as ML –, MHL, and MH 2L +). Time-resolved fluorescence studies of Eu-(EDDAG-DA) solutions (at varying pH) identifiedmore » a constant inner-sphere hydration number of 3, suggesting that glycine functionalities contained in the amide pendant arms are not involved in metal complexation and are protonated under more acidic conditions. The thermodynamic studies identified that f-element coordination by EDDAG-DA is similar to that observed for ethylenediamine- N,N,N',N'-tetraacetic acid (EDTA). However, coordination via two amidic oxygens of EDDAG-DA lowers its trivalent f-element complex stability by roughly 3 orders of magnitude relative to EDTA.« less

Thermodynamic and Spectroscopic Studies of Trivalent f -element Complexation with Ethylenediamine- N,N '-di(acetylglycine)- N,N '-diacetic Acid

DOE PAGES

Heathman, Colt R.; Grimes, Travis S.; Zalupski, Peter R.

2016-03-21

In this study, the coordination behavior and thermodynamic features of complexation of trivalent lanthanides and americium by ethylenediamine- N,N'-di(acetylglycine)- N,N'-diacetic acid (EDDAG-DA) (bisamide-substituted-EDTA) were investigated by potentiometric and spectroscopic techniques. Acid dissociation constants (K a) and complexation constants (β) of lanthanides (except Pm) were determined by potentiometric analysis. Absorption spectroscopy was used to determine stability constants for the binding of trivalent americium and neodymium by EDDAG-DA under similar conditions. The potentiometry revealed 5 discernible protonation constants and 3 distinct metal–ligand complexes (identified as ML –, MHL, and MH 2L +). Time-resolved fluorescence studies of Eu-(EDDAG-DA) solutions (at varying pH) identifiedmore » a constant inner-sphere hydration number of 3, suggesting that glycine functionalities contained in the amide pendant arms are not involved in metal complexation and are protonated under more acidic conditions. The thermodynamic studies identified that f-element coordination by EDDAG-DA is similar to that observed for ethylenediamine- N,N,N',N'-tetraacetic acid (EDTA). However, coordination via two amidic oxygens of EDDAG-DA lowers its trivalent f-element complex stability by roughly 3 orders of magnitude relative to EDTA.« less

Proton and metal ion binding to natural organic polyelectrolytes-I. Studies with synthetic model compounds

USGS Publications Warehouse

Marinsky, J.A.; Reddy, M.M.

1984-01-01

A unified physico-chemical model, based on a modified Henderson-Hasselbalch equation, for the analysis of ion complexation reactions involving charged polymeric systems is presented and verified. In this model pH = pKa+p(??Ka) + log(??/1 - ??) where Ka is the intrinsic acid dissociation constant of the ionizable functional groups on the polymer, ??Ka is the deviation of the intrinsic constant due to electrostatic interaction between the hydrogen ion and the polyanion, and alpha (??) is the polyacid degree of ionization. Using this approach pKa values for repeating acidic units of polyacrylic (PAA) and polymethacrylic (PMA) acids were found to be 4.25 ?? 0.03 and 4.8 ?? 0.1, respectively. The polyion electrostatic deviation term derived from the potentiometric titration data (i.e. p(??Ka)) is used to calculate metal ion concentration at the complexation site on the surface of the polyanion. Intrinsic cobalt-polycarboxylate binding constants (7.5 for PAA and 5.6 for PMA), obtained using this procedure, are consistent with the range of published binding constants for cobalt-monomer carboxylate complexes. In two phase systems incorporation of a Donnan membrane potential term allows determination of the intrinsic pKa of a cross-linked PMA gel, pKa = 4.83, in excellent agreement with the value obtained for the linear polyelectrolyte and the monomer. Similarly, the intrinsic stability constant for cobalt ion binding to a PMA-gel (??CoPMA+ = 11) was found to be in agreement with the linear polyelectrolyte analogue and the published data for cobalt-carboxylate monodentate complexes. ?? 1984.

DNA binding of a proflavine derivative bearing a platinum hanging residue.

PubMed

Biagini, Silvia; Bianchi, Antonio; Biver, Tarita; Boggioni, Alessia; Nikolayenko, Igor V; Secco, Fernando; Venturini, Marcella

2011-04-01

New platinum(II) complex of 3,6-diamine-9-[6,6-bis(2-aminohethyl)-1,6-diaminohexyl]acridine, AzaPt, has been synthesised and characterised. Behaviour of AzaPt in solution (protonation and possible self-aggregation phenomena) has been investigated by spectral methods (absorbance and fluorescence) at I=0.1M and 25°C, and the equilibrium parameters of binding to calf thymus DNA have been established. Two different modes of DNA binding by the complex were detected, which depend on the polymer to dye molar ratio (P/D). At relatively low P/D values the mode was interpreted as binding by the polyamine residue external to the base pairs, while at high P/D values the binding corresponds to intercalation of the proflavine residue. Such interpretation is supported by the observed salt effect on binding and the temperature variation of the binding constants, which allowed estimating the ΔH and ΔS values contributions. Spectrophotometric analysis of the long time range binding revealed that AzaPt is involved in a slow reaction, interpreted as an attack by the platinum ion on the nucleobases. The time constant for such interaction was calculated and found to be the same order of magnitude as for processes responsible for the action of anti-tumour drugs that do covalently bind to polynucleotides. Copyright © 2010 Elsevier Inc. All rights reserved.

Role of protons in the pump cycle of KdpFABC investigated by time-resolved kinetic experiments.

PubMed

Damnjanovic, Bojana; Apell, Hans-Jürgen

2014-05-20

The time-resolved kinetics of the KdpFABC complex solubilized in Aminoxide WS-35 was investigated by ATP concentration jump experiments. ATP was photoreleased from its inactive precursor, caged ATP, and charge movements in the membrane domain of the KdpFABC were detected by the electrochromic dye RH421. At low ATP concentrations, the ATP binding step became rate-limiting with an apparent, pH-independent ATP binding affinity of ~70 μM. At saturating ATP concentrations, the rate-limiting step is the conformational transition (E1-P → P-E2) with a rate constant of ~1.7 s(-1) at 20 °C that was independent of K(+) concentration. This observation together with the detected fluorescence decrease indicates that K(+) (or another positive ion) is bound in the membrane domain after enzyme phosphorylation and the conformational transition to the P-E2 state. pH dependence experiments revealed different roles of H(+) in the transport mechanism. Two different functions of protons for the ion pump must be distinguished. On one hand, there are electrogenically bound "functional" protons, which are not transported but prerequisite for the performance of the ATP-driven half-cycle. On the other hand, protons bind to the transport sites, acting as weak congeners of K(+). There possibly are noncompetitively bound protons, affecting the enzyme activity and/or coupling between KdpA and KdpB subunits. Finally, the recently proposed Post-Albers model for the KdpFABC complex was supplemented with stoichiometry factors of 2 for K(+) and 3 for H(+), and additional inhibitory side reactions controlled by H(+) were introduced, which are relevant at pH <6.5 and/or in the absence of K(+).

A calorimetric investigation of the interaction of the lac repressor with inducer.

PubMed

Donnér, J; Caruthers, M H; Gill, S J

1982-12-25

A calorimetric study has been made of the interaction between the lac repressor and isopropyl-1-thio-beta-D-galactopyranoside (IPTG). The buffer-corrected enthalpy of reaction at 25 degrees C was found to be -15.6, -24.7, -4.6 kJ/mol of bound IPTG at pH 7.0, pH 8.1, and pH 9.0, respectively. This large range of enthalpy values is in contrast to a maximum difference in the free energy of the reaction of only 1.5 kJ/mol of bound IPTG between these pH values. The reaction was found by calorimetric measurements in different buffers to be accompanied by an uptake of 0.29 mol of protons/mol of bound IPTG at pH 8.1. The pH dependency of the reaction enthalpy suggests differences in the extent of protonation of the binding site and the involvement of H bonding with IPTG. The lack of strong hydrophobic contributions in the IPTG binding process is revealed by the absence of any determinable heat capacity change for the reaction at pH 7.0. The presence of phosphate buffer significantly alters the enthalpy of IPTG binding at higher pH values, but has little effect upon the binding constant. This implies that highly negative phosphate species change the nature of the IPTG binding site without any displacement of phosphate upon IPTG binding.

PARACEST Properties of a Dinuclear Neodymium(III) Complex Bound to DNA or Carbonate

PubMed Central

Nwe, Kido; Andolina, Christopher M.; Huang, Ching-Hui; Morrow, Janet R.

2009-01-01

A dinuclear Nd(III) macrocyclic complex of 1 (1,4-bis[1-(4,7,10-tris(carbamoylmethyl)-1,4,7,10-tetraazacyclododecane]-p-xylene) and mononuclear complexes of 1,4,7-tris-1,4,7,10-tetraazacyclododecane 2, and 1,4,7-tris[(N-N-diethyl)carbamoylmethyl]-1,4,7,10-tetraazacyclododecane, 3, are prepared. Complexes of 1 and 2 give rise to a PARACEST (paramagnetic chemical exchange saturation transfer) peak from exchangeable amide protons that resonate approximately 12 ppm downfield from the bulk water proton resonance. The dinuclear Nd(III) complex is promising as a PARACEST contrast agent for MRI applications because it has an optimal pH of 7.5 and the rate constant for amide proton exchange (2700 s−1) is nearly as large as it can be within slow exchange conditions with bulk water. Dinuclear Ln2(1) complexes (Ln(III) = Nd(III), Eu(III)) bind tightly to anionic ligands including carbonate, diethylphosphate and DNA. The CEST amide peak of Nd2(1) is enhanced by certain DNA sequences that contain hairpin loops, but decreases in the presence of diethyl phosphate or carbonate. Direct excitation luminescence studies of Eu2(1) show that double-stranded and hairpin loop DNA sequences displace one water ligand on each Eu(III) center. DNA displaces carbonate ion despite the low dissociation constant for the Eu2(1) carbonate complex (Kd = 15 µM). Enhancement of the CEST effect of a lanthanide complex by binding to DNA is a promising step toward the preparation of PARACEST agents containing DNA scaffolds. PMID:19555071

PARACEST properties of a dinuclear neodymium(III) complex bound to DNA or carbonate.

PubMed

Nwe, Kido; Andolina, Christopher M; Huang, Ching-Hui; Morrow, Janet R

2009-07-01

A dinuclear Nd(III) macrocyclic complex of 1 (1,4-bis[1-(4,7,10-tris(carbamoylmethyl)-1,4,7,10-tetraazacyclododecane]-p-xylene) and mononuclear complexes of 1,4,7-tris-1,4,7,10-tetraazacyclododecane, 2, and 1,4,7-tris[(N-N-diethyl)carbamoylmethyl]-1,4,7,10-tetraazacyclododecane, 3, are prepared. Complexes of 1 and 2 give rise to a PARACEST (paramagnetic chemical exchange saturation transfer) peak from exchangeable amide protons that resonate approximately 12 ppm downfield from the bulk water proton resonance. The dinuclear Nd(III) complex is promising as a PARACEST contrast agent for MRI applications, because it has an optimal pH of 7.5 and the rate constant for amide proton exchange (2700 s(-1)) is nearly as large as it can be within slow exchange conditions with bulk water. Dinuclear Ln(2)(1) complexes (Ln(III) = Nd(III), Eu(III)) bind tightly to anionic ligands including carbonate, diethyl phosphate, and DNA. The CEST amide peak of Nd(2)(1) is enhanced by certain DNA sequences that contain hairpin loops, but decreases in the presence of diethyl phosphate or carbonate. Direct excitation luminescence studies of Eu(2)(1) show that double-stranded and hairpin-loop DNA sequences displace one water ligand on each Eu(III) center. DNA displaces carbonate ion despite the low dissociation constant for the Eu(2)(1) carbonate complex (K(d) = 15 microM). Enhancement of the CEST effect of a lanthanide complex by binding to DNA is a promising step toward the preparation of PARACEST agents containing DNA scaffolds.

Enzymatic function of loop movement in enolase: preparation and some properties of H159N, H159A, H159F, and N207A enolases.

PubMed

Brewer, John M; Glover, Claiborne V C; Holland, Michael J; Lebioda, Lukasz

2003-05-01

The hypothesis that His159 in yeast enolase moves on a polypeptide loop to protonate the phosphoryl of 2-phosphoglycerate to initiate its conversion to phosphoenolpyruvate was tested by preparing H159N, H159A, and H159F enolases. These have 0.07%-0.25% of the native activity under standard assay conditions and the pH dependence of maximum velocities of H159A and H159N mutants is markedly altered. Activation by Mg2+ is biphasic, with the smaller Mg2+ activation constant closer to that of the "catalytic" Mg2+ binding site of native enolase and the larger in the mM range in which native enolase is inhibited. A third Mg2+ may bind to the phosphoryl, functionally replacing proton donation by His159. N207A enolase lacks an intersubunit interaction that stabilizes the closed loop(s) conformation when 2-phosphoglycerate binds. It has 21% of the native activity, also exhibits biphasic Mg2+ activation, and its reaction with the aldehyde analogue of the substrate is more strongly inhibited than is its normal enzymatic reaction. Polypeptide loop(s) closure may keep a proton from His159 interacting with the substrate phosphoryl oxygen long enough to stabilize a carbanion intermediate.

Nonadiabatic rate constants for proton transfer and proton-coupled electron transfer reactions in solution: Effects of quadratic term in the vibronic coupling expansion

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

Soudackov, Alexander V.; Hammes-Schiffer, Sharon

2015-11-21

Rate constant expressions for vibronically nonadiabatic proton transfer and proton-coupled electron transfer reactions are presented and analyzed. The regimes covered include electronically adiabatic and nonadiabatic reactions, as well as high-frequency and low-frequency proton donor-acceptor vibrational modes. These rate constants differ from previous rate constants derived with the cumulant expansion approach in that the logarithmic expansion of the vibronic coupling in terms of the proton donor-acceptor distance includes a quadratic as well as a linear term. The analysis illustrates that inclusion of this quadratic term in the framework of the cumulant expansion framework may significantly impact the rate constants at highmore » temperatures for proton transfer interfaces with soft proton donor-acceptor modes that are associated with small force constants and weak hydrogen bonds. The effects of the quadratic term may also become significant in these regimes when using the vibronic coupling expansion in conjunction with a thermal averaging procedure for calculating the rate constant. In this case, however, the expansion of the coupling can be avoided entirely by calculating the couplings explicitly for the range of proton donor-acceptor distances sampled. The effects of the quadratic term for weak hydrogen-bonding systems are less significant for more physically realistic models that prevent the sampling of unphysical short proton donor-acceptor distances. Additionally, the rigorous relation between the cumulant expansion and thermal averaging approaches is clarified. In particular, the cumulant expansion rate constant includes effects from dynamical interference between the proton donor-acceptor and solvent motions and becomes equivalent to the thermally averaged rate constant when these dynamical effects are neglected. This analysis identifies the regimes in which each rate constant expression is valid and thus will be important for future applications to proton transfer and proton-coupled electron transfer in chemical and biological processes.« less

Binding and Energetics of Electron Transfer between an Artificial Four-Helix Mn-Protein and Reaction Centers from Rhodobacter sphaeroides.

PubMed

Espiritu, Eduardo; Olson, Tien L; Williams, JoAnn C; Allen, James P

2017-12-12

The ability of an artificial four-helix bundle Mn-protein, P1, to bind and transfer an electron to photosynthetic reaction centers from the purple bacterium Rhodobacter sphaeroides was characterized using optical spectroscopy. Upon illumination of reaction centers, an electron is transferred from P, the bacteriochlorophyll dimer, to Q A , the primary electron acceptor. The P1 Mn-protein can bind to the reaction center and reduce the oxidized bacteriochlorophyll dimer, P + , with a dissociation constant of 1.2 μM at pH 9.4, comparable to the binding constant of c-type cytochromes. Amino acid substitutions of surface residues on the Mn-protein resulted in increases in the dissociation constant to 8.3 μM. The extent of reduction of P + by the P1 Mn-protein was dependent on the P/P + midpoint potential and the pH. Analysis of the free energy difference yielded a midpoint potential of approximately 635 mV at pH 9.4 for the Mn cofactor of the P1 Mn-protein, a value similar to those found for other Mn cofactors in proteins. The linear dependence of -56 mV/pH is consistent with one proton being released upon Mn oxidation, allowing the complex to maintain overall charge neutrality. These outcomes demonstrate the feasibility of designing four-helix bundles and other artificial metalloproteins to bind and transfer electrons to bacterial reaction centers and establish the usefulness of this system as a platform for designing sites to bind novel metal cofactors capable of performing complex oxidation-reduction reactions.

Mechanism of host-guest complexation by cucurbituril.

PubMed

Márquez, César; Hudgins, Robert R; Nau, Werner M

2004-05-12

The factors affecting host-guest complexation between the molecular container compound cucurbit[6]uril (CB6) and various guests in aqueous solution are studied, and a detailed complexation mechanism in the presence of cations is derived. The formation of the supramolecular complex is studied in detail for cyclohexylmethylammonium ion as guest. The kinetics and thermodynamics of complexation is monitored by NMR as a function of temperature, salt concentration, and cation size. The binding constants and the ingression rate constants decrease with increasing salt concentration and cation-binding constant, in agreement with a competitive binding of the ammonium site of the guest and the metal cation with the ureido carbonyl portals of CB6. Studies as a function of guest size indicate that the effective container volume of the CB6 cavity is approximately 105 A(3). It is suggested that larger guests are excluded for two reasons: a high activation barrier for ingression imposed by the tight CB6 portals and a destabilization of the complex due to steric repulsion inside. For example, in the case of the nearly spherical azoalkane homologues 2,3-diazabicyclo[2.2.1]hept-2-ene (DBH, volume ca. 96 A(3)) and 2,3-diazabicyclo[2.2.2]oct-2-ene (DBO, volume ca. 110 A(3)), the former forms the CB6 complex promptly with a sizable binding constant (1300 M(-1)), while the latter does not form a complex even after several months at optimized complexation conditions. Molecular mechanics calculations are performed for several CB6/guest complexes. A qualitative agreement is found between experimental and calculated activation energies for ingression as a function of both guest size and state of protonation. The potential role of constrictive binding by CB6 is discussed.

Computation of pH-Dependent Binding Free Energies

PubMed Central

Kim, M. Olivia; McCammon, J. Andrew

2015-01-01

Protein-ligand binding accompanies changes in the surrounding electrostatic environments of the two binding partners and may lead to changes in protonation upon binding. In cases where the complex formation results in a net transfer of protons, the binding process is pH-dependent. However, conventional free energy computations or molecular docking protocols typically employ fixed protonation states for the titratable groups in both binding partners set a priori, which are identical for the free and bound states. In this review, we draw attention to these important yet largely ignored binding-induced protonation changes in protein-ligand association by outlining physical origins and prevalence of the protonation changes upon binding. Following a summary of various theoretical methods for pKa prediction, we discuss the theoretical framework to examine the pH dependence of protein-ligand binding processes. PMID:26202905

Specific Binding of Adamantane Drugs and Direction of their Polar Amines in the Pore of the Influenza M2 Transmembrane Domain in Lipid Bilayers and Dodecylphosphocholine Micelles Determined by NMR Spectroscopy

PubMed Central

Cady, Sarah D.; Wang, Jun; Wu, Yibing; DeGrado, William F.; Hong, Mei

2011-01-01

The transmembrane domain of the influenza M2 protein (M2TM) forms a tetrameric proton channel important for the virus lifecycle. The proton-channel activity is inhibited by amine-containing adamantyl drugs amantadine and rimantadine, which have been shown to bind specifically to the pore of M2TM near Ser31. However, whether the polar amine points to the N- or C-terminus of the channel has not yet been determined. Elucidating the polar group direction will shed light on the mechanism by which drug binding inhibits this proton channel and will facilitate rational design of new inhibitors. In this study, we determine the polar amine direction using M2TM reconstituted in lipid bilayers as well as DPC micelles. 13C-2H rotational-echo double-resonance NMR experiments of 13C-labeled M2TM and methyl-deuterated rimantadine in lipid bilayers showed that the polar amine pointed to the C-terminus of the channel, with the methyl group close to Gly34. Solution NMR experiments of M2TM in dodecylphosphocholine (DPC) micelles indicate that drug binding causes significant chemical shift perturbations of the protein that are very similar to those seen for M2TM and M2(18–60) bound to lipid bilayers. Specific 2H-labeling of the drugs permitted the assignment of drug-protein cross peaks, which indicate that amantadine and rimantadine bind to the pore in the same fashion as for bilayer-bound M2TM. These results strongly suggest that adamantyl inhibition of M2TM is achieved not only by direct physical occlusion of the pore but also by perturbing the equilibrium constant of the proton-sensing residue His37. The reproduction of the pharmacologically relevant specific pore-binding site in DPC micelles, which was not observed with a different detergent, DHPC, underscores the significant influence of the detergent environment on the functional structure of membrane proteins. PMID:21381693

Proton NMR spectroscopic characterization of binary and ternary complexes of cobalt(II) carboxypeptidase A with inhibitors

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

Bertini, I.; Luchinat, C.; Messori, L.

The binding of L- and D-phenylalanine and carboxylate inhibitors to cobalt(II)-substituted carboxypeptidase A, Co(II)CPD (E), in the presence and absence of pseudohalogens (X = N/sub 3//sup -/, NCO/sup -/, and NCS/sup -/) has been studied by /sup 1/H NMR spectroscopy. This technique monitors the proton signals of histidine residues bound to cobalt(II) and is therefore sensitive to the interactions of inhibitors that perturb the coordination sphere of the metal. Enzyme-inhibitor complexes, E/times/I, E/times/I/sub 2/, and E/times/I/times/X, each with characteristic NMR features, have been identified. The NMR data suggest that when the carboxylate group of a substrate of inhibitor binds atmore » the active site, a conformational change occurs that allows a second ligand molecule to bind to the metal ion, altering its coordination sphere and thereby attenuating the bidentate behavior of Glu-72. The /sup 1/H NMR signals also reflect alterations in the histidine interactions with the metal upon inhibitor binding. Isotropic shifts in the signals for the C-4 (c) and N protons (a) of one of the histidine ligands are readily observed in all of these complexes. These signals are relatively constant for all E/times/I and E/times/I/times/X complexes, indicating that this ligand is in a relatively fixed or buried conformation. However in the 2:1 carboxylate inhibitor (E/times/I/sub 2/) complexes, both signals are shifted upfield, suggesting a disturbance in the interaction of this histidine with the metal.« less

Proton transfer pathways, energy landscape, and kinetics in creatine-water systems.

PubMed

Ivchenko, Olga; Whittleston, Chris S; Carr, Joanne M; Imhof, Petra; Goerke, Steffen; Bachert, Peter; Wales, David J

2014-02-27

We study the exchange processes of the metabolite creatine, which is present in both tumorous and normal tissues and has NH2 and NH groups that can transfer protons to water. Creatine produces chemical exchange saturation transfer (CEST) contrast in magnetic resonance imaging (MRI). The proton transfer pathway from zwitterionic creatine to water is examined using a kinetic transition network constructed from the discrete path sampling approach and an approximate quantum-chemical energy function, employing the self-consistent-charge density-functional tight-binding (SCC-DFTB) method. The resulting potential energy surface is visualized by constructing disconnectivity graphs. The energy landscape consists of two distinct regions corresponding to the zwitterionic creatine structures and deprotonated creatine. The activation energy that characterizes the proton transfer from the creatine NH2 group to water was determined from an Arrhenius fit of rate constants as a function of temperature, obtained from harmonic transition state theory. The result is in reasonable agreement with values obtained in water exchange spectroscopy (WEX) experiments.

Unraveling HIV protease flaps dynamics by Constant pH Molecular Dynamics simulations.

PubMed

Soares, Rosemberg O; Torres, Pedro H M; da Silva, Manuela L; Pascutti, Pedro G

2016-08-01

The active site of HIV protease (HIV-PR) is covered by two flaps. These flaps are known to be essential for the catalytic activity of the HIV-PR, but their exact conformations at the different stages of the enzymatic pathway remain subject to debate. Understanding the correct functional dynamics of the flaps might aid the development of new HIV-PR inhibitors. It is known that, the HIV-PR catalytic efficiency is pH-dependent, likely due to the influence of processes such as charge transfer and protonation/deprotonation of ionizable residues. Several Molecular Dynamics (MD) simulations have reported information about the HIV-PR flaps. However, in MD simulations the protonation of a residue is fixed and thus it is not possible to study the correlation between conformation and protonation state. To address this shortcoming, this work attempts to capture, through Constant pH Molecular Dynamics (CpHMD), the conformations of the apo, substrate-bound and inhibitor-bound HIV-PR, which differ drastically in their flap arrangements. The results show that the HIV-PR flaps conformations are defined by the protonation of the catalytic residues Asp25/Asp25' and that these residues are sensitive to pH changes. This study suggests that the catalytic aspartates can modulate the opening of the active site and substrate binding. Copyright © 2016 Elsevier Inc. All rights reserved.

Copper removal by algal biomass: biosorbents characterization and equilibrium modelling.

PubMed

Vilar, Vítor J P; Botelho, Cidália M S; Pinheiro, José P S; Domingos, Rute F; Boaventura, Rui A R

2009-04-30

The general principles of Cu(II) binding to algal waste from agar extraction, composite material and algae Gelidium, and different modelling approaches, are discussed. FTIR analyses provided a detailed description of the possible binding groups present in the biosorbents, as carboxylic groups (D-glucuronic and pyruvic acids), hydroxyl groups (cellulose, agar and floridean starch) and sulfonate groups (sulphated galactans). Potentiometric acid-base titrations showed a heterogeneous distribution of two major binding groups, carboxyl and hydroxyl, following the quasi-Gaussian affinity constant distribution suggested by Sips, which permitted to estimate the maximum amount of acid functional groups (0.36, 0.25 and 0.1 mmol g(-1)) and proton binding parameters (pK(H)=5.0, 5.3 and 4.4; m(H)=0.43, 0.37, 0.33), respectively for algae Gelidium, algal waste and composite material. A non-ideal, semi-empirical, thermodynamically consistent (NICCA) isotherm fitted better the experimental ion binding data for different pH values and copper concentrations, considering only the acid functional groups, than the discrete model. Values of pK(M) (3.2; 3.6 and 3.3), n(M) (0.98, 0.91, 1.0) and p (0.67, 0.53 and 0.43) were obtained, respectively for algae Gelidium, algal waste and composite material. NICCA model reflects the complex macromolecular systems that take part in biosorption considering the heterogeneity of the biosorbent, the competition between protons and metals ions to the binding sites and the stoichiometry for different ions.

DNA binding by a new metallointercalator that contains a proflavine group bearing a hanging chelating unit.

PubMed

Bazzicalupi, Carla; Bencini, Andrea; Bianchi, Antonio; Biver, Tarita; Boggioni, Alessia; Bonacchi, Sara; Danesi, Andrea; Giorgi, Claudia; Gratteri, Paola; Ingraín, Antonio Marchal; Secco, Fernando; Sissi, Claudia; Valtancoli, Barbara; Venturini, Marcella

2008-01-01

The new bifunctional molecule 3,6-diamine-9-[6,6-bis(2-aminoethyl)-1,6-diaminohexyl]acridine (D), which is characterised by both an aromatic moiety and a separate metal-complexing polyamine centre, has been synthesised. The characteristics of D and its ZnII complex ([ZnD]) (protonation and metal-complexing constants, optical properties and self-aggregation phenomena) have been analysed by means of NMR spectroscopy, potentiometric, spectrophotometric and spectrofluorimetric techniques. The equilibria and kinetics of the binding process of D and [ZnD] to calf thymus DNA have been investigated at I=0.11 M (NaCl) and 298.1 K by using spectroscopic methods and the stopped-flow technique. Static measurements show biphasic behaviour for both D-DNA and [ZnD]-DNA systems; this reveals the occurrence of two different binding processes depending on the polymer-to-dye molar ratio (P/D). The binding mode that occurs at low P/D values is interpreted in terms of external binding with a notable contribution from the polyamine residue. The binding mode at high P/D values corresponds to intercalation of the proflavine residue. Stopped-flow, circular dichroism and supercoiled-DNA unwinding experiments corroborate the proposed mechanism. Molecular-modelling studies support the intercalative process and evidence the influence of NH+...O interactions between the protonated acridine nitrogen atom and the oxygen atoms of the polyanion; these interactions play a key role in determining the conformation of DNA adducts.

Proton-bound dimers of nitrogen heterocyclic molecules: Substituent effects on the structures and binding energies of homodimers of diazine, triazine, and fluoropyridine

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

Attah, Isaac K.; Platt, Sean P.; Meot-Ner, Michael

2014-03-21

The bonding energies of proton-bound homodimers BH{sup +}B were measured by ion mobility equilibrium studies and calculated at the DFT B3LYP/6-311++G{sup **} level, for a series of nitrogen heterocyclic molecules (B) with electron-withdrawing in-ring N and on-ring F substituents. The binding energies (ΔH°{sub dissoc}) of the proton-bound dimers (BH{sup +}B) vary significantly, from 29.7 to 18.1 kcal/mol, decreasing linearly with decreasing the proton affinity of the monomer (B). This trend differs significantly from the constant binding energies of most homodimers of other organic nitrogen and oxygen bases. The experimentally measured ΔH°{sub dissoc} for (1,3-diazine){sub 2}H{sup +}, i.e., (pyrimidine){sub 2}H{sup +}more » and (3-F-pyridine){sub 2}H{sup +} are 22.7 and 23.0 kcal/mol, respectively. The measured ΔH°{sub dissoc} for the pyrimidine{sup ·+}(3-F-pyridine) radical cation dimer (19.2 kcal/mol) is signifcantly lower than that of the proton-bound homodimers of pyrimidine and 3-F-pyridine, reflecting the stronger interaction in the ionic H-bond of the protonated dimers. The calculated binding energies for (1,2-diazine){sub 2}H{sup +}, (pyridine){sub 2}H{sup +}, (2-F-pyridine){sub 2}H{sup +}, (3-F-pyridine){sub 2}H{sup +}, (2,6-di-F-pyridine){sub 2}H{sup +}, (4-F-pyridine){sub 2}H{sup +}, (1,3-diazine){sub 2}H{sup +}, (1,4-diazine){sub 2}H{sup +}, (1,3,5-triazine){sub 2}H{sup +}, and (pentafluoropyridine){sub 2}H{sup +} are 29.7, 24.9, 24.8, 23.3, 23.2, 23.0, 22.4, 21.9, 19.3, and 18.1 kcal/mol, respectively. The electron-withdrawing substituents form internal dipoles whose electrostatic interactions contribute to both the decreased proton affinities of (B) and the decreased binding energies of the protonated dimers BH{sup +}B. The bonding energies also vary with rotation about the hydrogen bond, and they decrease in rotamers where the internal dipoles of the components are aligned efficiently for inter-ring repulsion. For compounds substituted at the 3 or 4 (meta or para) positions, the lowest energy rotamers are T-shaped with the planes of the two rings rotated by 90° about the hydrogen bond, while the planar rotamers are weakened by repulsion between the ortho hydrogen atoms of the two rings. Conversely, in ortho-substituted (1,2-diazine){sub 2}H{sup +} and (2-F-pyridine){sub 2}H{sup +}, attractive interactions between the ortho (C–H) hydrogen atoms of one ring and the electronegative ortho atoms (N or F) of the other ring are stabilizing, and increase the protonated dimer binding energies by up to 4 kcal/mol. In all of the dimers, rotation about the hydrogen bond can involve a 2–4 kcal/mol barrier due to the relative energies of the rotamers.« less

Characterization of the proton binding sites of extracellular polymeric substances in an anaerobic membrane bioreactor.

PubMed

Liu, Yi; Chang, Sheng; Defersha, Fantahun M

2015-07-01

This paper focuses on the characterization of the chemical compositions and acidic constants of the extracellular polymeric substances (EPSs) in an anaerobic membrane bioreactor treating synthetic brewery wastewater by using chemical analysis, linear programming analysis (LPA) of titration data, and FT-IR analysis. The linear programming analysis of titration data revealed that the EPSs have proton binding sites with pKa values from pKa ≤ 6, between 6 and 7, and approximately 9.8. The strong acidic sites (pKa ≤ 6) and some weak acidic sites (7.5 < pKa < 9.0) were found to be readily removed by 0.45-μm membrane filtration. In addition, the FT-IR analysis confirmed the presence of proteins, carbohydrates, nucleic acids, and lipids in the EPS samples. Based on the FT-IR analysis and the main chemical functional groups at the bacterial cell surfaces, the identified proton binding sites were related to carboxyl, phosphate, and hydroxyl/amine groups with pKa values of 4.6 ± 0.7, 6.6 ± 0.01, and 9.7 ± 0.1, respectively, with the corresponding respective intensities of 0.31 ± 0.05, 0.96 ± 0.3, and 1.53 ± 0.3 mmole/g-EPS. The pKa values and intensities of the proton binding sites are the fundamental molecular properties of EPSs that affect the EPS charge, molecular interactions, and metal complexation characteristics. Determination of such properties can advance Derjaguin-Landau-Verwey-Overbeek (DLVO)-based concentration polarization modeling, facilitate the estimation of the osmotic pressure of the EPS concentration polarization layers, and lead to a deeper understanding of the role of metal complexation in membrane fouling. Copyright © 2015 Elsevier Ltd. All rights reserved.

Cyanide binding to human plasma heme-hemopexin: A comparative study

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

Ascenzi, Paolo, E-mail: ascenzi@uniroma3.it; Istituto Nazionale di Biostrutture e Biosistemi, Roma; Leboffe, Loris

Highlights: Black-Right-Pointing-Pointer Cyanide binding to ferric HHPX-heme-Fe. Black-Right-Pointing-Pointer Cyanide binding to ferrous HHPX-heme-Fe. Black-Right-Pointing-Pointer Dithionite-mediated reduction of ferric HHPX-heme-Fe-cyanide. Black-Right-Pointing-Pointer Cyanide binding to HHPX-heme-Fe is limited by ligand deprotonation. Black-Right-Pointing-Pointer Cyanide dissociation from HHPX-heme-Fe-cyanide is limited by ligand protonation. -- Abstract: Hemopexin (HPX) displays a pivotal role in heme scavenging and delivery to the liver. In turn, heme-Fe-hemopexin (HPX-heme-Fe) displays heme-based spectroscopic and reactivity properties. Here, kinetics and thermodynamics of cyanide binding to ferric and ferrous hexa-coordinate human plasma HPX-heme-Fe (HHPX-heme-Fe(III) and HHPX-heme-Fe(II), respectively), and for the dithionite-mediated reduction of the HHPX-heme-Fe(III)-cyanide complex, at pH 7.4 and 20.0 Degree-Sign C,more » are reported. Values of thermodynamic and kinetic parameters for cyanide binding to HHPX-heme-Fe(III) and HHPX-heme-Fe(II) are K = (4.1 {+-} 0.4) Multiplication-Sign 10{sup -6} M, k{sub on} = (6.9 {+-} 0.5) Multiplication-Sign 10{sup 1} M{sup -1} s{sup -1}, and k{sub off} = 2.8 Multiplication-Sign 10{sup -4} s{sup -1}; and H = (6 {+-} 1) Multiplication-Sign 10{sup -1} M, h{sub on} = 1.2 Multiplication-Sign 10{sup -1} M{sup -1} s{sup -1}, and h{sub off} = (7.1 {+-} 0.8) Multiplication-Sign 10{sup -2} s{sup -1}, respectively. The value of the rate constant for the dithionite-mediated reduction of the HHPX-heme-Fe(III)-cyanide complex is l = 8.9 {+-} 0.8 M{sup -1/2} s{sup -1}. HHPX-heme-Fe reactivity is modulated by proton acceptor/donor amino acid residue(s) (e.g., His236) assisting the deprotonation and protonation of the incoming and outgoing ligand, respectively.« less

Comparative studies on the human serum albumin binding of the clinically approved EGFR inhibitors gefitinib, erlotinib, afatinib, osimertinib and the investigational inhibitor KP2187.

PubMed

Dömötör, Orsolya; Pelivan, Karla; Borics, Attila; Keppler, Bernhard K; Kowol, Christian R; Enyedy, Éva A

2018-05-30

Binding interactions between human serum albumin (HSA) and four approved epidermal growth factor receptor (EGFR) inhibitors gefitinib (GEF), erlotinib (ERL), afatinib (AFA), osimertinib (OSI), as well as the experimental drug KP2187, were investigated by means of spectrofluorometric and molecular modelling methods. Steady-state and time resolved spectrofluorometric techniques were carried out, including direct quenching of protein fluorescence and site marker displacement measurements. Proton dissociation processes and solvent dependent fluorescence properties were investigated as well. The EGFR inhibitors were predominantly presented in their single protonated form (HL + ) at physiological pH except ERL, which is charge-neutral. Significant solvent dependent fluorescence properties were found for GEF, ERL and KP2187, namely their emission spectra show strong dependence on the polarity and the hydrogen bonding ability of the solvents. The inhibitors proved to be bound at site I of HSA (in subdomain IIA) in a weak-to-moderate fashion (logK' 3.9-4.9) using spectrofluorometry. OSI (logK' 4.3) and KP2187 can additionally bind in site II (in subdomain IIIA), while GEF, ERL and AFA clearly show no interaction here. Docking methods qualitatively confirmed binding site preferences of compounds GEF and KP2187, and indicated that they probably bind to HSA in their neutral forms. Binding constants calculated on the basis of the various experimental data indicate a weak-to-moderate binding on HSA, only OSI exhibits somewhat higher affinity towards this protein. However, model calculations performed at physiological blood concentrations of HSA resulted in high (ca. 90%) bound fractions for the inhibitors, highlighting the importance of plasma protein binding. Copyright © 2018 Elsevier B.V. All rights reserved.

Modeling Ionization Events iduced by Protein Protein Binding

NASA Astrophysics Data System (ADS)

Mitra, Rooplekha; Shyam, Radhey; Alexov, Emil

2009-11-01

The association of two or more biological macromolecules dramatically change the environment of the amino acids situated at binding interface and could change ionization states of titratable groups. The change of ionization due to the binding results in proton uptake/release and causes pH-dependence of the binding free energy. We apply computational method, as implemented in Multi Conformation Continuum Electrostatics (MCCE) algorithm, to study protonation evens on a large set of protein-protein complexes. Our results indicate that proton uptake/release is a common phenomena in protein binding since in vast majority of the cases (70%) the binding caused at least 0.5 units proton change. The proton uptake/release was further investigated with respect to interfacial area and charges of the monomers and it was found that macroscopic characteristics are not important determinants. Instead, charge complementarity across the interface and the number of unpaired ionizable groups at the interface are the primary source of proton uptake/release.

Stretched proton-neutron configurations in fp-shell nuclei (II). Systematics

NASA Astrophysics Data System (ADS)

von Neumann-Cosel, P.; Fister, U.; Jahn, R.; Schenk, P.; Trelle, T. K.; Wenzel, D.; Wienands, U.

1994-03-01

The systematics of the binding energies of stretched proton-neutron configurations ( f{7}/{2}, g{9}/{2}) 8 -, ( p{3}/{2}, g{9}/{2}) 6 -, ( g{9}/{2}, p{3}/{2}) 6- and ( g{9}/{2}) 29 + are studied over a wide range of f p-shell nuclei. The effective proton-neutron interaction energies deduced from the data are nearly constant for ( p{3}/{2}, g{9}/{2}) 6 -and ( g{9}/{2}) 29 + states while the ( f{7}/{2}, g{9}/{2}) 8 - configuration reveals an additional repulsive term proportional to the partial filling of the f{7}/{2} orbit in the target ground state. Two-body matrix elements are extracted. A crude shell model, which predicts that the excitation energy of a stretched state is equal to the sum of the single-particle energies, works well for the 6 - and 9 + states, but fails for the 8 - levels due to neglect of the additional interactions described above. The physics underlying the empirically introduced basic assumptions of the crude shell model is discussed. The binding energies are found to be linearly dependent on the mass number A and the isospin Tz component and are well described by the weak-coupling model of Bansal and French. The derived parameters agree with averaged values of a similar analysis for the single-particle states in the corresponding odd-even neighbours. The data indicate a significant change of the particle-hole energies with closure of the proton f{7}/{2} shell.

Mechanism for pH-dependent gene regulation by amino-terminus-mediated homooligomerization of Bacillus subtilis anti-trp RNA-binding attenuation protein

PubMed Central

Sachleben, Joseph R.; McElroy, Craig A.; Gollnick, Paul; Foster, Mark P.

2010-01-01

Anti-TRAP (AT) is a small zinc-binding protein that regulates tryptophan biosynthesis in Bacillus subtilis by binding to tryptophan-bound trp RNA-binding attenuation protein (TRAP), thereby preventing it from binding RNA, and allowing transcription and translation of the trpEDCFBA operon. Crystallographic and sedimentation studies have shown that AT can homooligomerize to form a dodecamer, AT12, composed of a tetramer of trimers, AT3. Structural and biochemical studies suggest that only trimeric AT is active for binding to TRAP. Our chromatographic and spectroscopic data revealed that a large fraction of recombinantly overexpressed AT retains the N-formyl group (fAT), presumably due to incomplete N-formyl-methionine processing by peptide deformylase. Hydrodynamic parameters from NMR relaxation and diffusion measurements showed that fAT is exclusively trimeric (AT3), while (deformylated) AT exhibits slow exchange between both trimeric and dodecameric forms. We examined this equilibrium using NMR spectroscopy and found that oligomerization of active AT3 to form inactive AT12 is linked to protonation of the amino terminus. Global analysis of the pH dependence of the trimer-dodecamer equilibrium revealed a near physiological pKa for the N-terminal amine of AT and yielded a pH-dependent oligomerization equilibrium constant. Estimates of excluded volume effects due to molecular crowding suggest the oligomerization equilibrium may be physiologically important. Because deprotonation favors “active” trimeric AT and protonation favors “inactive” dodecameric AT, our findings illuminate a possible mechanism for sensing and responding to changes in cellular pH. PMID:20713740

Kinetic and equilibrium studies of acrylonitrile binding to cytochrome c peroxidase and oxidation of acrylonitrile by cytochrome c peroxidase compound I.

PubMed

Chinchilla, Diana; Kilheeney, Heather; Vitello, Lidia B; Erman, James E

2014-01-03

Ferric heme proteins bind weakly basic ligands and the binding affinity is often pH dependent due to protonation of the ligand as well as the protein. In an effort to find a small, neutral ligand without significant acid/base properties to probe ligand binding reactions in ferric heme proteins we were led to consider the organonitriles. Although organonitriles are known to bind to transition metals, we have been unable to find any prior studies of nitrile binding to heme proteins. In this communication we report on the equilibrium and kinetic properties of acrylonitrile binding to cytochrome c peroxidase (CcP) as well as the oxidation of acrylonitrile by CcP compound I. Acrylonitrile binding to CcP is independent of pH between pH 4 and 8. The association and dissociation rate constants are 0.32±0.16 M(-1) s(-1) and 0.34±0.15 s(-1), respectively, and the independently measured equilibrium dissociation constant for the complex is 1.1±0.2 M. We have demonstrated for the first time that acrylonitrile can bind to a ferric heme protein. The binding mechanism appears to be a simple, one-step association of the ligand with the heme iron. We have also demonstrated that CcP can catalyze the oxidation of acrylonitrile, most likely to 2-cyanoethylene oxide in a "peroxygenase"-type reaction, with rates that are similar to rat liver microsomal cytochrome P450-catalyzed oxidation of acrylonitrile in the monooxygenase reaction. CcP compound I oxidizes acrylonitrile with a maximum turnover number of 0.61 min(-1) at pH 6.0. Copyright © 2013 Elsevier Inc. All rights reserved.

Kinetic and equilibrium studies of acrylonitrile binding to cytochrome c peroxidase and oxidation of acrylonitrile by cytochrome c peroxidase compound I

PubMed Central

Chinchilla, Diana; Kilheeney, Heather; Vitello, Lidia B.; Erman, James E.

2013-01-01

Ferric heme proteins bind weakly basic ligands and the binding affinity is often pH dependent due to protonation of the ligand as well as the protein. In an effort to find a small, neutral ligand without significant acid/base properties to probe ligand binding reactions in ferric heme proteins we were led to consider the organonitriles. Although organonitriles are known to bind to transition metals, we have been unable to find any prior studies of nitrile binding to heme proteins. In this communication we report on the equilibrium and kinetic properties of acrylonitrile binding to cytochrome c peroxidase (CcP) as well as the oxidation of acrylonitrile by CcP compound I. Acrylonitrile binding to CcP is independent of pH between pH 4 and 8. The association and dissociation rate constants are 0.32 ± 0.16 M−1s−1 and 0.34 ± 0.15 s−1, respectively, and the independently measured equilibrium dissociation constant for the complex is 1.1 ± 0.2 M. We have demonstrated for the first time that acrylonitrile can bind to a ferric heme protein. The binding mechanism appears to be a simple, one-step association of the ligand with the heme iron. We have also demonstrated that CcP can catalyze the oxidation of acrylonitrile, most likely to 2-cyanoethylene oxide in a “peroxygenase”-type reaction, with rates that are similar to rat liver microsomal cytochrome P450-catalyzed oxidation of acrylonitrile in the monooxygenase reaction. CcP compound I oxidizes acrylonitrile with a maximum turnover number of 0.61 min−1 at pH 6.0. PMID:24291498

Nonadiabatic rate constants for proton transfer and proton-coupled electron transfer reactions in solution: Effects of quadratic term in the vibronic coupling expansion

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

Soudackov, Alexander; Hammes-Schiffer, Sharon

2015-11-17

Rate constant expressions for vibronically nonadiabatic proton transfer and proton-coupled electron transfer reactions are presented and analyzed. The regimes covered include electronically adiabatic and nonadiabatic reactions, as well as high-frequency and low-frequency regimes for the proton donor-acceptor vibrational mode. These rate constants differ from previous rate constants derived with the cumulant expansion approach in that the logarithmic expansion of the vibronic coupling in terms of the proton donor-acceptor distance includes a quadratic as well as a linear term. The analysis illustrates that inclusion of this quadratic term does not significantly impact the rate constants derived using the cumulant expansion approachmore » in any of the regimes studied. The effects of the quadratic term may become significant when using the vibronic coupling expansion in conjunction with a thermal averaging procedure for calculating the rate constant, however, particularly at high temperatures and for proton transfer interfaces with extremely soft proton donor-acceptor modes that are associated with extraordinarily weak hydrogen bonds. Even with the thermal averaging procedure, the effects of the quadratic term for weak hydrogen-bonding systems are less significant for more physically realistic models that prevent the sampling of unphysical short proton donor-acceptor distances, and the expansion of the coupling can be avoided entirely by calculating the couplings explicitly for the range of proton donor-acceptor distances. This analysis identifies the regimes in which each rate constant expression is valid and thus will be important for future applications to proton transfer and proton-coupled electron transfer in chemical and biological processes. We are grateful for support from National Institutes of Health Grant GM056207 (applications to enzymes) and the Center for Molecular Electrocatalysis, an Energy Frontier Research Center funded by the U.S. Department of Energy, Office of Science, Basic Energy Sciences (applications to molecular electrocatalysts).« less

Temperature-Dependent Estimation of Gibbs Energies Using an Updated Group-Contribution Method.

PubMed

Du, Bin; Zhang, Zhen; Grubner, Sharon; Yurkovich, James T; Palsson, Bernhard O; Zielinski, Daniel C

2018-06-05

Reaction-equilibrium constants determine the metabolite concentrations necessary to drive flux through metabolic pathways. Group-contribution methods offer a way to estimate reaction-equilibrium constants at wide coverage across the metabolic network. Here, we present an updated group-contribution method with 1) additional curated thermodynamic data used in fitting and 2) capabilities to calculate equilibrium constants as a function of temperature. We first collected and curated aqueous thermodynamic data, including reaction-equilibrium constants, enthalpies of reaction, Gibbs free energies of formation, enthalpies of formation, entropy changes of formation of compounds, and proton- and metal-ion-binding constants. Next, we formulated the calculation of equilibrium constants as a function of temperature and calculated the standard entropy change of formation (Δ f S ∘ ) using a model based on molecular properties. The median absolute error in estimating Δ f S ∘ was 0.013 kJ/K/mol. We also estimated magnesium binding constants for 618 compounds using a linear regression model validated against measured data. We demonstrate the improved performance of the current method (8.17 kJ/mol in median absolute residual) over the current state-of-the-art method (11.47 kJ/mol) in estimating the 185 new reactions added in this work. The efforts here fill in gaps for thermodynamic calculations under various conditions, specifically different temperatures and metal-ion concentrations. These, to our knowledge, new capabilities empower the study of thermodynamic driving forces underlying the metabolic function of organisms living under diverse conditions. Copyright © 2018 Biophysical Society. Published by Elsevier Inc. All rights reserved.

The fast release of sticky protons: Kinetics of substrate binding and proton release in a multidrug transporter

PubMed Central

Adam, Yoav; Tayer, Naama; Rotem, Dvir; Schreiber, Gideon; Schuldiner, Shimon

2007-01-01

EmrE is an Escherichia coli H+-coupled multidrug transporter that provides a unique experimental paradigm because of its small size and stability, and because its activity can be studied in detergent solution. In this work, we report a study of the transient kinetics of substrate binding and substrate-induced proton release in EmrE. For this purpose, we measured transient changes in the tryptophan fluorescence upon substrate binding and the rates of substrate-induced proton release. The fluorescence of the essential and fully conserved Trp residue at position 63 is sensitive to the occupancy of the binding site with either protons or substrate. The maximal rate of binding to detergent-solubilized EmrE of TPP+, a high-affinity substrate, is 2 × 107 M−1·s−1, a rate typical of diffusion-limited reactions. Rate measurements with medium- and low-affinity substrates imply that the affinity is determined mainly by the koff of the substrate. The rates of substrate binding and substrate-induced release of protons are faster at basic pHs and slower at lower pHs. These findings imply that the substrate-binding rates are determined by the generation of the species capable of binding; this is controlled by the high affinity to protons of the glutamate at position 14, because an Asp replacement with a lower pK is faster at the same pHs. PMID:17984053

The binding of aluminum to mugineic acid and related compounds as studied by potentiometric titration.

PubMed

Yoshimura, Etsuro; Kohdr, Hicham; Mori, Satoshi; Hider, Robert C

2011-08-01

The phytosiderophores, mugineic acid (MA) and epi-hydroxymugineic acid (HMA), together with a related compound, nicotianamine (NA), were investigated for their ability to bind Al(III). Potentiometric titration analysis demonstrated that MA and HMA bind Al(III), in contrast to NA which does not under normal physiological conditions. With MA and HMA, in addition to the Al complex (AlL), the protonated (AlLH) and deprotonated (AlLH(-1)) complexes were identified from an analysis of titration curves, where L denotes the phytosiderophore form in which all the carboxylate functions are ionized. The equilibrium formation constants of the Al(III) phytosiderophore complexes are much smaller than those of the corresponding Fe(III) complexes. The higher selectivity of phytosiderophores for Fe(III) over Al(III) facilitates Fe(III) acquisition in alkaline conditions where free Al(III) levels are higher than free Fe(III) levels.

Δ isobars and nuclear saturation

NASA Astrophysics Data System (ADS)

Ekström, A.; Hagen, G.; Morris, T. D.; Papenbrock, T.; Schwartz, P. D.

2018-02-01

We construct a nuclear interaction in chiral effective field theory with explicit inclusion of the Δ -isobar Δ (1232 ) degree of freedom at all orders up to next-to-next-to-leading order (NNLO). We use pion-nucleon (π N ) low-energy constants (LECs) from a Roy-Steiner analysis of π N scattering data, optimize the LECs in the contact potentials up to NNLO to reproduce low-energy nucleon-nucleon scattering phase shifts, and constrain the three-nucleon interaction at NNLO to reproduce the binding energy and point-proton radius of 4He. For heavier nuclei we use the coupled-cluster method to compute binding energies, radii, and neutron skins. We find that radii and binding energies are much improved for interactions with explicit inclusion of Δ (1232 ) , while Δ -less interactions produce nuclei that are not bound with respect to breakup into α particles. The saturation of nuclear matter is significantly improved, and its symmetry energy is consistent with empirical estimates.

Application of a continuous distribution model for proton binding by humic acids extracted from acidic lake sediments

NASA Astrophysics Data System (ADS)

Rhea, James R.; Young, Thomas C.

1987-10-01

The proton binding characteristics of humic acids extracted from the sediments of Cranberry Pond, an acidic water body located in the Adirondack Mountain region of New York State, were explored by the application of a multiligand distribution model. The model characterizes a class of proton binding sites by mean log K values and the standard deviations of log K values about the mean. Mean log K values and their relative abundances were determined directly from experimental titration data. The model accurately predicts the binding of protons by the humic acids for pH values in the range 3.5 to 10.0.

Application of a continuous distribution model for proton binding by humic acids extracted from acidic lake sediments

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

Rhea, J.R.; Young, T.C.

1987-01-01

The proton binding characteristics of humic acids extracted from the sediments of Cranberry Pond, an acidic water body located in the Adirondack Mountain region of New York State, were explored by the application of a nultiligand distribution model. The model characterizes a class of proton binding sites by mean log K values and the standard deviations of log K values and the mean. Mean log K values and their relative abundances were determined directly from experimental titration data. The model accurately predicts the binding of protons by the humic acids for pH values in the range 3.5 to 10.0.

Molecular beads on a charged molecular string

NASA Astrophysics Data System (ADS)

Zhang, Haizhen; Ferrell, Tyler A.; Asplund, Matthew C.; Dearden, David V.

2007-09-01

Complexes of [alpha],[omega]-alkyldiammonium cations [H3N+(CH2)nNH3+, n = 2-10] with the cyclic, hollow ligand cucurbit[6]uril (CB6) were characterized in the gas phase using Fourier transform ion cyclotron resonance mass spectrometry with energy resolved sustained off-resonance irradiation (SORI) collision induced dissociation, in combination with HF/6-31G* and B3LYP/6-31G* computational methods. All the complexes have the diammonium cation threaded through the cavity of CB6. The modeled supramolecular geometries, the SORI energies required for dissociation of the complexes and for appearance of singly protonated diamine product ions, and the branching ratios for the various dissociation channels all suggest that the optimum [alpha],[omega]-alkyldiammonium chain length for binding CB6 in the gas phase occurs for n = 4. This contrasts with observed complex stability constants in aqueous formic acid, which are maximum for n = 6, reflecting solvent stabilization of the ammonium groups that is not possible in the gas phase. At the B3LYP/6-31G* level of theory, the binding energy for the n = 4 complex with respect to dissociation to singly protonated butanediamine and protonated CB6 is 204 kJ mol-1. The n = 6 complex exhibits especially low dissociation thresholds, perhaps reflecting compression of the diammonium cation upon complexation with CB6, forming a loaded "molecular spring."

Host-guest chemistry of dendrimer-drug complexes. 4. An in-depth look into the binding/encapsulation of guanosine monophosphate by dendrimers.

PubMed

Hu, Jingjing; Fang, Min; Cheng, Yiyun; Zhang, Jiahai; Wu, Qinglin; Xu, Tongwen

2010-06-03

In the present study, we investigated the host-guest chemistry of dendrimer/guanosine monophosphate (GMP) and present an in-depth look into the binding/encapsulation of GMP by dendrimers using NMR studies. (1)H NMR spectra showed a significant downfield shift of methylene protons in the outmost layer of the G5 dendrimer, indicating the formation of ion pairs between cationic amine groups of dendrimer and anionic phosphate groups of GMP. Chemical shift titration results showed that the binding constant between G5 dendrimer and GMP is 17,400 M(-1) and each G5 dendrimer has 107 binding sites. The binding of GMP to dendrimers prevents its aggregation in aqueous solutions and thereby enhances its stability. Nuclear Overhauser effect measurements indicated that a GMP binding and encapsulation balance occurs on the surface and in the interior of dendrimer. The binding/encapsulation transitions can be easily tailored by altering the surface and interior charge densities of the dendrimer. All these findings provide a new insight into the host-guest chemistry of dendrimer/guest complexes and may play important roles in the study of dendrimer/DNA aggregates by a "bottom-up" strategy.

Location and ion-binding of membrane-associated valinomycin, a proton nuclear magnetic resonance study.

PubMed

Meers, P; Feigenson, G W

1988-03-03

Valinomycin, incorporated in small unilamellar vesicles of perdeuterated dimyristoylphosphatidylcholine, reveals several well-resolved 1H-NMR resonances. These resonances were used to examine the location, orientation and ion-binding of membrane-bound valinomycin. The order of affinity of membrane-bound valinomycin for cations is Rb+ greater than K+ greater than Cs+ greater than Ba2+, and binding is sensitive to surface change. The exchange between bound and free forms is fast on the NMR time scale. The intrinsic binding constants, extrapolated to zero anion concentration, are similar to those determined in aqueous solution. Rb+ and K+ show 1:1 binding to valinomycin, whereas the stoichiometry of Cs+ and Ba2+ is not certain. Paramagnetic chemical shift reagents and nitroxide spin label relaxation probes were used to study the location and orientation of valinomycin in the membrane. Despite relatively fast exchange of bound cations, the time average location of the cation-free form of valinomycin is deep within the bilayer under the conditions of these experiments. Upon complexation to K+, valinomycin moves closer to the interfacial region.

Protein Structure Validation and Refinement Using Amide Proton Chemical Shifts Derived from Quantum Mechanics

PubMed Central

Christensen, Anders S.; Linnet, Troels E.; Borg, Mikael; Boomsma, Wouter; Lindorff-Larsen, Kresten; Hamelryck, Thomas; Jensen, Jan H.

2013-01-01

We present the ProCS method for the rapid and accurate prediction of protein backbone amide proton chemical shifts - sensitive probes of the geometry of key hydrogen bonds that determine protein structure. ProCS is parameterized against quantum mechanical (QM) calculations and reproduces high level QM results obtained for a small protein with an RMSD of 0.25 ppm (r = 0.94). ProCS is interfaced with the PHAISTOS protein simulation program and is used to infer statistical protein ensembles that reflect experimentally measured amide proton chemical shift values. Such chemical shift-based structural refinements, starting from high-resolution X-ray structures of Protein G, ubiquitin, and SMN Tudor Domain, result in average chemical shifts, hydrogen bond geometries, and trans-hydrogen bond (h3 JNC') spin-spin coupling constants that are in excellent agreement with experiment. We show that the structural sensitivity of the QM-based amide proton chemical shift predictions is needed to obtain this agreement. The ProCS method thus offers a powerful new tool for refining the structures of hydrogen bonding networks to high accuracy with many potential applications such as protein flexibility in ligand binding. PMID:24391900

Charge-Neutral Constant pH Molecular Dynamics Simulations Using a Parsimonious Proton Buffer.

PubMed

Donnini, Serena; Ullmann, R Thomas; Groenhof, Gerrit; Grubmüller, Helmut

2016-03-08

In constant pH molecular dynamics simulations, the protonation states of titratable sites can respond to changes of the pH and of their electrostatic environment. Consequently, the number of protons bound to the biomolecule, and therefore the overall charge of the system, fluctuates during the simulation. To avoid artifacts associated with a non-neutral simulation system, we introduce an approach to maintain neutrality of the simulation box in constant pH molecular dynamics simulations, while maintaining an accurate description of all protonation fluctuations. Specifically, we introduce a proton buffer that, like a buffer in experiment, can exchange protons with the biomolecule enabling its charge to fluctuate. To keep the total charge of the system constant, the uptake and release of protons by the buffer are coupled to the titration of the biomolecule with a constraint. We find that, because the fluctuation of the total charge (number of protons) of a typical biomolecule is much smaller than the number of titratable sites of the biomolecule, the number of buffer sites required to maintain overall charge neutrality without compromising the charge fluctuations of the biomolecule, is typically much smaller than the number of titratable sites, implying markedly enhanced simulation and sampling efficiency.

Solvent Dependence of Double Proton Transfer in the Formic Acid-Formamidine Complex: Path Integral Molecular Dynamics Investigation.

PubMed

Kungwan, Nawee; Ngaojampa, Chanisorn; Ogata, Yudai; Kawatsu, Tsutomu; Oba, Yuki; Kawashima, Yukio; Tachikawa, Masanori

2017-10-05

Solvent dependence of double proton transfer in the formic acid-formamidine (FA-FN) complex at room temperature was investigated by means of ab initio path integral molecular dynamics (AIPIMD) simulation with taking nuclear quantum and thermal effects into account. The conductor-like screening model (COSMO) was applied for solvent effect. In comparison with gas phase, double proton delocalization between two heavy atoms (O and N) in FA-FN were observed with reduced proton transfer barrier height in low dielectric constant medium (<4.8). For dielectric constant medium at 4.8, the chance of finding these two protons are more pronounced due to the solvent effect which completely washes out the proton transfer barrier. In the case of higher dielectric constant medium (>4.8), the ionic species becomes more stable than the neutral ones and the formate anion and formamidium cation are thermodynamically stable. For ab initio molecular dynamics simulation, in low dielectric constant medium (<4.8) a reduction of proton transfer barrier with solvent effect is found to be less pronounced than the AIPIMD due to the absence of nuclear quantum effect. Moreover, the motions of FA-FN complex are significantly different with increasing dielectric constant medium. Such a difference is revealed in detail by the principal component analysis.

Acid-base and copper-binding properties of three organic matter fractions isolated from a forest floor soil solution

NASA Astrophysics Data System (ADS)

van Schaik, Joris W. J.; Kleja, Dan B.; Gustafsson, Jon Petter

2010-02-01

Vast amounts of knowledge about the proton- and metal-binding properties of dissolved organic matter (DOM) in natural waters have been obtained in studies on isolated humic and fulvic (hydrophobic) acids. Although macromolecular hydrophilic acids normally make up about one-third of DOM, their proton- and metal-binding properties are poorly known. Here, we investigated the acid-base and Cu-binding properties of the hydrophobic (fulvic) acid fraction and two hydrophilic fractions isolated from a soil solution. Proton titrations revealed a higher total charge for the hydrophilic acid fractions than for the hydrophobic acid fraction. The most hydrophilic fraction appeared to be dominated by weak acid sites, as evidenced by increased slope of the curve of surface charge versus pH at pH values above 6. The titration curves were poorly predicted by both Stockholm Humic Model (SHM) and NICA-Donnan model calculations using generic parameter values, but could be modelled accurately after optimisation of the proton-binding parameters (pH ⩽ 9). Cu-binding isotherms for the three fractions were determined at pH values of 4, 6 and 9. With the optimised proton-binding parameters, the SHM model predictions for Cu binding improved, whereas the NICA-Donnan predictions deteriorated. After optimisation of Cu-binding parameters, both models described the experimental data satisfactorily. Iron(III) and aluminium competed strongly with Cu for binding sites at both pH 4 and pH 6. The SHM model predicted this competition reasonably well, but the NICA-Donnan model underestimated the effects significantly at pH 6. Overall, the Cu-binding behaviour of the two hydrophilic acid fractions was very similar to that of the hydrophobic acid fraction, despite the differences observed in proton-binding characteristics. These results show that for modelling purposes, it is essential to include the hydrophilic acid fraction in the pool of 'active' humic substances.

Insights into the functional role of protonation states in the HIV-1 protease-BEA369 complex: molecular dynamics simulations and free energy calculations.

PubMed

Chen, Jianzhong; Yang, Maoyou; Hu, Guodong; Shi, Shuhua; Yi, Changhong; Zhang, Qinggang

2009-10-01

The molecular mechanics Poisson-Boltzmann surface area (MM-PBSA) method combined with molecular dynamics (MD) simulations were used to investigate the functional role of protonation in human immunodeficiency virus type 1 (HIV-1) protease complexed with the inhibitor BEA369. Our results demonstrate that protonation of two aspartic acids (Asp25/Asp25') has a strong influence on the dynamics behavior of the complex, the binding free energy of BEA369, and inhibitor-residue interactions. Relative binding free energies calculated using the MM-PBSA method show that protonation of Asp25 results in the strongest binding of BEA369 to HIV-1 protease. Inhibitor-residue interactions computed by the theory of free energy decomposition also indicate that protonation of Asp25 has the most favorable effect on binding of BEA369. In addition, hydrogen-bond analysis based on the trajectories of the MD simulations shows that protonation of Asp25 strongly influences the water-mediated link of a conserved water molecule, Wat301. We expect that the results of this study will contribute significantly to binding calculations for BEA369, and to the design of high affinity inhibitors.

Effect of Temperature on the Protonation of the TALSPEAK Ligands: Lactic and Diethylenetrinitropentaacetic Acids

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

Tian, Guoxin; Rao, Linfeng

2009-10-20

The protonation reactions of two ligands that play important roles in the TALSPEAK process for the separation of trivalent actinides from lanthanides, lactic acid and diethylenetrinitropentaacetic acid (DTPA), have been studied at variable temperatures. The protonation constants at 10-70 C were determined by titration potentiometry and the protonation enthalpies were determined at 25 C by titration microcalorimetry. The protonation constants remain essentially unchanged (25-70 C) within the experimental uncertainties, indicating that the effect of temperature on the protonation of lactate is insignificant. In contrast, the protonation constants of DTPA (log {beta}H's) generally decrease as the temperature is increased. Results frommore » this study indicate that the effect of temperature on the protonation of DTPA could alter the speciation of metal ions (actinides and lanthanides) in the TALSPEAK system, since lower values of log{beta}H at higher temperatures suggest that the hydrogen ions would compete less strongly with the metal ions for the complexation of DTPA at higher temperatures.« less

Poly(amidoamine) dendrimers on lipid bilayers II: Effects of bilayer phase and dendrimer termination.

PubMed

Kelly, Christopher V; Leroueil, Pascale R; Orr, Bradford G; Banaszak Holl, Mark M; Andricioaei, Ioan

2008-08-07

The molecular structures and enthalpy release of poly(amidoamine) (PAMAM) dendrimers binding to 1,2-dimyristoyl- sn-glycero-3-phosphocholine (DMPC) bilayers were explored through atomistic molecular dynamics. Three PAMAM dendrimer terminations were examined: protonated primary amine, neutral acetamide, and deprotonated carboxylic acid. Fluid and gel lipid phases were examined to extract the effects of lipid tail mobility on the binding of generation-3 dendrimers, which are directly relevant to the nanoparticle interactions involving lipid rafts, endocytosis, lipid removal, and/or membrane pores. Upon binding to gel phase lipids, dendrimers remained spherical, had a constant radius of gyration, and approximately one-quarter of the terminal groups were in close proximity to the lipids. In contrast, upon binding to fluid phase bilayers, dendrimers flattened out with a large increase in their asphericity and radii of gyration. Although over twice as many dendrimer-lipid contacts were formed on fluid versus gel phase lipids, the dendrimer-lipid interaction energy was only 20% stronger. The greatest enthalpy release upon binding was between the charged dendrimers and the lipid bilayer. However, the stronger binding to fluid versus gel phase lipids was driven by the hydrophobic interactions between the inner dendrimer and lipid tails.

Study of the hydrolysis and ionization constants of Schiff base from pyridoxal 5'-phosphate and n-hexylamine in partially aqueous solvents. An application to phosphorylase b.

PubMed Central

Donoso, J; Muñoz, F; García Del Vado, A; Echevarría, G; García Blanco, F

1986-01-01

Formation and hydrolysis rate constants as well as equilibrium constants of the Schiff base derived from pyridoxal 5'-phosphate and n-hexylamine were determined between pH 3.5 and 7.5 in ethanol/water mixtures (3:17, v/v, and 49:1, v/v). The results indicate that solvent polarity scarcely alters the values of these constants but that they are dependent on the pH. Spectrophotometric titration of this Schiff base was also carried out. We found that a pKa value of 6.1, attributed in high-polarity media to protonation of the pyridine nitrogen atom, is independent of solvent polarity, whereas the pKa of the monoprotonated form of the imine falls from 12.5 in ethanol/water (3:17) to 11.3 in ethanol/water (49:1). Fitting of the experimental results for the hydrolysis to a theoretical model indicates the existence of a group with a pKa value of 6.1 that is crucial in the variation of kinetic constant of hydrolysis with pH. Studies of the reactivity of the coenzyme (pyridoxal 5'-phosphate) of glycogen phosphorylase b with hydroxylamine show that this reaction only occurs when the pH value of solution is below 6.5 and the hydrolysis of imine bond has started. We propose that the decrease in activity of phosphorylase b when the pH value is less than 6.2 must be caused by the cleavage of enzyme-coenzyme binding and that this may be related with protonation of the pyridine nitrogen atom of pyridoxal 5'-phosphate. PMID:3099764

Protonated Nitrous Oxide, NNOH(+): Fundamental Vibrational Frequencies and Spectroscopic Constants from Quartic Force Fields

NASA Technical Reports Server (NTRS)

Huang, Xinchuan; Fortenberry, Ryan C.; Lee, Timothy J.

2013-01-01

The interstellar presence of protonated nitrous oxide has been suspected for some time. Using established high-accuracy quantum chemical techniques, spectroscopic constants and fundamental vibrational frequencies are provided for the lower energy O-protonated isomer of this cation and its deuterated isotopologue. The vibrationally-averaged B0 and C0 rotational constants are within 6 MHz of their experimental values and the D(subJ) quartic distortion constants agree with experiment to within 3%. The known gas phase O-H stretch of NNOH(+) is 3330.91 cm(exp-1), and the vibrational configuration interaction computed result is 3330.9 cm(exp-1). Other spectroscopic constants are also provided, as are the rest of the fundamental vibrational frequencies for NNOH(+) and its deuterated isotopologue. This high-accuracy data should serve to better inform future observational or experimental studies of the rovibrational bands of protonated nitrous oxide in the ISM and the laboratory.

Solution and fluorescence properties of symmetric dipicolylamine-containing dichlorofluorescein-based Zn2+ sensors.

PubMed

Wong, Brian A; Friedle, Simone; Lippard, Stephen J

2009-05-27

The mechanism by which dipicolylamine (DPA) chelate-appended fluorophores respond to zinc was investigated by the synthesis and study of five new analogues of the 2',7'-dichlorofluorescein-based Zn(2+) sensor Zinpyr-1 (ZP1). With the use of absorption and emission spectroscopy in combination with potentiometric titrations, a detailed molecular picture has emerged of the Zn(2+) and H(+) binding properties of the ZP1 family of sensors. The two separate N(3)O donor atom sets on ZP1 converge to form binding pockets in which all four heteroatoms participate in coordination to either Zn(2+) or protons. The position of the pyridyl group nitrogen atom, 2-pyridyl or 4-pyridyl, has a large impact on the fluorescence response of the dyes to protons despite relatively small changes in pK(a) values. The fluorescence quenching effects of such multifunctional electron-donating units are often taken as a whole. Despite the structural complexity of ZP1, however, we provide evidence that the pyridyl arms of the DPA appendages participate in the quenching process, in addition to the contribution from the tertiary nitrogen amine atom. Potentiometric titrations reveal ZP1 dissociation constants (K(d)) for Zn(2+) of 0.04 pM and 1.2 nM for binding to the first and second binding pockets of the ligand, respectively, the second of which correlates with the value observed by fluorescence titration. This result demonstrates that both binding pockets of this symmetric, ditopic sensor need to be occupied in order for full fluorescence turn-on to be achieved. These results have significant implications for the design and implementation of fluorescent sensors for studies of mobile zinc ions in biology.

A fluorescent combinatorial logic gate with Na+, H+-enabled OR and H+-driven low-medium-high ternary logic functions.

PubMed

Spiteri, Jasmine M A; Mallia, Carl J; Scerri, Glenn J; Magri, David C

2017-12-06

A novel fluorescent molecular logic gate with a 'fluorophore-spacer 1 -receptor 1 -spacer 2 -receptor 2 ' format is demonstrated in 1 : 1 (v/v) methanol/water. The molecule consists of an anthracene fluorophore, and tertiary alkyl amine and N-(2-methoxyphenyl)aza-15-crown-5 ether receptors. In the presence of threshold concentrations of H + and Na + , the molecule switches 'on' as an AND logic gate with a fluorescence quantum yield of 0.21 with proton and sodium binding constants of log β H+ = 9.0 and log β Na+ = 3.2, respectively. At higher proton levels, protonation also occurs at the anilinic nitrogen atom ether with a log β H+ = 4.2, which allows for Na + , H + -enabled OR (OR + AND circuit) and H + -driven ternary logic functions. The reported molecule is compared and contrasted to classic anthracene-based Na + and H + logic gates. We propose that such logic-based molecules could be useful tools for probing the vicinity of Na + , H + antiporters in biological systems.

Reduction and Reoxidation of Humic Acid: Influence on Spectroscopic Properties and Proton Binding

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

Maurer, F.; Christl, I; Kretzschmar, R

2010-01-01

Previous studies on proton and metal binding to humic substances have not considered a potential influence of reduction and oxidation of functional groups. Therefore, we investigated how proton binding of a purified soil humic acid was affected by reduction. Reduction of the humic acid was carried out using an electrochemical cell that allowed us to measure the amounts of electrons and protons involved in reduction reactions. We further applied spectroscopic methods (UV-vis, fluorescence, FT-IR, C-1s NEXAFS) to detect possible chemical changes in the humic acid induced by reduction and reoxidation. The effect of reduction on proton binding was determined withmore » acid-base titrations in the pH range 4-10 under controlled redox conditions. During reduction, 0.54 mol kg{sup -1} protons and 0.55 mol kg{sup -1} electrons were transferred to humic acid. NICA-Donnan modeling revealed an equivalent increase in proton-reactive sites (0.52 mol kg{sup -1}) in the alkaline pH-range. Our results indicate that reduction of humic acid increased the amount of proton-reactive sites by 15% compared to the untreated state. Spectroscopic differences between the untreated and reduced humic acid were minor, apart from a lower UV-vis absorption of the reduced humic acid between 400 and 700 nm.« less

Molecular modeling and simulation studies of recombinant laccase from Yersinia enterocolitica suggests significant role in the biotransformation of non-steroidal anti-inflammatory drugs

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

Singh, Deepti; Rawat, Surender; Waseem, Mohd

The YacK gene from Yersinia enterocolitica strain 7, cloned in pET28a vector and expressed in Escherichia coli BL21 (DE3), showed laccase activity when oxidized with 2,2′-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS) and guaiacol. The recombinant laccase protein was purified and characterized biochemically with a molecular mass of ≈58 KDa on SDS-PAGE and showed positive zymogram with ABTS. The protein was highly robust with optimum pH 9.0 and stable at 70 °C upto 12 h with residual activity of 70%. Kinetic constants, K{sub m} values, for ABTS and guaiacol were 675 μM and 2070 μM, respectively, with corresponding Vmax values of 0.125 μmol/ml/min and 6500 μmol/ml/min. It also possess antioxidative propertymore » against BSA and Cu{sup 2+}/H{sub 2}O{sub 2} model system. Constant pH MD simulation studies at different protonation states of the system showed ABTS to be most stable at acidic pH, whereas, diclofenac at neutral pH. Interestingly, aspirin drifted out of the binding pocket at acidic and neutral pH, but showed stable binding at alkaline pH. The biotransformation of diclofenac and aspirin by laccase also corroborated the in silico results. This is the first report on biotransformation of non-steroidal anti-inflammatory drugs (NSAIDs) using recombinant laccase from gut bacteria, supported by in silico simulation studies. - Highlights: • Laccase from Yersinia enterocolitica strain 7 was expressed in Escherichia coli BL21 (DE3). • Recombinant laccase was found to be thermostable and alkali tolerant. • The in silico and experimental studied proves the biotransformation of NSAIDs. • Laccase binds to ligands differentially under different protonation state. • Laccase also possesses free radical scavenging property.« less

Spectroscopic and TDDFT investigation on highly selective fluorogenic chemosensor and construction of molecular logic gates.

PubMed

Basheer, Sabeel M; Kumar, Saravana Loganathan Ashok; Kumar, Moorthy Saravana; Sreekanth, Anandaram

2017-03-01

1,5-Bis(2-fluorene)thiocarbohydrazone (FBTC) was designed and synthesized for selective sensing of fluoride and copper ions. The binding constants of FBTC towards fluoride and copper ions have been calculated using the Benesi-Hildebrand equation, and FBTC has more binding affinity towards copper ion than fluoride ion. The 1 H NMR and 13 C NMR titration studies strongly support the deprotonation was taken from the N-H protons followed by the formation of hydrogen bond via N-H … F. To understand the fluoride ion sensing mechanism, theoretical investigation had been carried out using the density functional theory and time-dependent density functional theory. The theoretical data well reproduced the experimental results. The deprotonation process has a moderate transition barrier (481.55kcal/mol). The calculated ΔE and ΔG values (-253.92 and -192.41kcal/mol respectively) suggest the feasibility of sensing process. The potential energy curves give the optimized structures of FBTC-F complex in the ground state and excited state, which states the proton transition occurs at the excited state. The excited state proton transition mechanism was further confirmed with natural bond orbital analysis. The reversibility of the sensor was monitored by the alternate addition of F - and Cu 2+ ions, which was explained with "Read-Erase-Write-Read" behaviour. The multi-ion detection of sensor used to construct the molecular logic gate, such as AND, OR, NOR and INHIBITION logic gates. Copyright © 2016 Elsevier B.V. All rights reserved.

Magnetic properties and core electron binding energies of liquid water

NASA Astrophysics Data System (ADS)

Galamba, N.; Cabral, Benedito J. C.

2018-01-01

The magnetic properties and the core and inner valence electron binding energies of liquid water are investigated. The adopted methodology relies on the combination of molecular dynamics and electronic structure calculations. Born-Oppenheimer molecular dynamics with the Becke and Lee-Yang-Parr functionals for exchange and correlation, respectively, and includes an empirical correction (BLYP-D3) functional and classical molecular dynamics with the TIP4P/2005-F model were carried out. The Keal-Tozer functional was applied for predicting magnetic shielding and spin-spin coupling constants. Core and inner valence electron binding energies in liquid water were calculated with symmetry adapted cluster-configuration interaction. The relationship between the magnetic shielding constant σ(17O), the role played by the oxygen atom as a proton acceptor and donor, and the tetrahedral organisation of liquid water are investigated. The results indicate that the deshielding of the oxygen atom in water is very dependent on the order parameter (q) describing the tetrahedral organisation of the hydrogen bond network. The strong sensitivity of magnetic properties on changes of the electronic density in the nuclei environment is illustrated by a correlation between σ(17O) and the energy gap between the 1a1[O1s] (core) and the 2a1 (inner valence) orbitals of water. Although several studies discussed the eventual connection between magnetic properties and core electron binding energies, such a correlation could not be clearly established. Here, we demonstrate that for liquid water this correlation exists although involving the gap between electron binding energies of core and inner valence orbitals.

/sup 2/H NMR demonstration of amino acid - nucleotide interactions

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

Khaled, M.A.; Watkins, C.L.; Lacey, J.C. Jr.

Deuterium (/sup 2/H) NMR was used to investigate the interaction of L-Phenylalanine (with aromatic protons replaced by deuterons (Phe-D/sub 5/)), with 5'-AMP and polyadenylic acid (poly A). A considerable change in line width of the aromatic deuteron signals of Phe was observed. These data were plotted using a Scatchard-type equation, and yielded apparent binding constants for L-Phe to 5'-AMP and poly A of 7 and 11 M/sup -1/, respectively. Future applications of /sup 2/H-NMR in the study of nucleic acid-protein interactions are discussed.

Ion adsorption at the rutile-water interface: linking molecular and macroscopic properties.

PubMed

Zhang, Z; Fenter, P; Cheng, L; Sturchio, N C; Bedzyk, M J; Predota, M; Bandura, A; Kubicki, J D; Lvov, S N; Cummings, P T; Chialvo, A A; Ridley, M K; Bénézeth, P; Anovitz, L; Palmer, D A; Machesky, M L; Wesolowski, D J

2004-06-08

A comprehensive picture of the interface between aqueous solutions and the (110) surface of rutile (alpha-TiO2) is being developed by combining molecular-scale and macroscopic approaches, including experimental measurements, quantum calculations, molecular simulations, and Gouy-Chapman-Stern models. In situ X-ray reflectivity and X-ray standing-wave measurements are used to define the atomic arrangement of adsorbed ions, the coordination of interfacial water molecules, and substrate surface termination and structure. Ab initio calculations and molecular dynamics simulations, validated through direct comparison with the X-ray results, are used to predict ion distributions not measured experimentally. Potentiometric titration and ion adsorption results for rutile powders having predominant (110) surface expression provide macroscopic constraints of electrical double layer (EDL) properties (e.g., proton release) which are evaluated by comparison with a three-layer EDL model including surface oxygen proton affinities calculated using ab initio bond lengths and partial charges. These results allow a direct correlation of the three-dimensional, crystallographically controlled arrangements of various species (H2O, Na+, Rb+, Ca2+, Sr2+, Zn2+, Y3+, Nd3+) with macroscopic observables (H+ release, metal uptake, zeta potential) and thermodynamic/electrostatic constraints. All cations are found to be adsorbed as "inner sphere" species bonded directly to surface oxygen atoms, while the specific binding geometries and reaction stoichiometries are dependent on ionic radius. Ternary surface complexes of sorbed cations with electrolyte anions are not observed. Finally, surface oxygen proton affinities computed using the MUSIC model are improved by incorporation of ab initio bond lengths and hydrogen bonding information derived from MD simulations. This multitechnique and multiscale approach demonstrates the compatibility of bond-valence models of surface oxygen proton affinities and Stern-based models of the EDL structure, with the actual molecular interfacial distributions observed experimentally, revealing new insight into EDL properties including specific binding sites and hydration states of sorbed ions, interfacial solvent properties (structure, diffusivity, dielectric constant), surface protonation and hydrolysis, and the effect of solution ionic strength.

Note: Proton microbeam formation with continuously variable kinetic energy using a compact system for three-dimensional proton beam writing

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

Ohkubo, T., E-mail: ohkubo.takeru@jaea.go.jp; Ishii, Y.

A compact focused gaseous ion beam system has been developed to form proton microbeams of a few hundreds of keV with a penetration depth of micrometer range in 3-dimensional proton beam writing. Proton microbeams with kinetic energies of 100-140 keV were experimentally formed on the same point at a constant ratio of the kinetic energy of the object side to that of the image side. The experimental results indicate that the beam diameters were measured to be almost constant at approximately 6 μm at the same point with the kinetic energy range. These characteristics of the system were experimentally andmore » numerically demonstrated to be maintained as long as the ratio was constant.« less

Complexity in Acid–Base Titrations: Multimer Formation Between Phosphoric Acids and Imines

PubMed Central

Malm, Christian; Kim, Heejae; Wagner, Manfred

2017-01-01

Abstract Solutions of Brønsted acids with bases in aprotic solvents are not only common model systems to study the fundamentals of proton transfer pathways but are also highly relevant to Brønsted acid catalysis. Despite their importance the light nature of the proton makes characterization of acid–base aggregates challenging. Here, we track such acid–base interactions over a broad range of relative compositions between diphenyl phosphoric acid and the base quinaldine in dichloromethane, by using a combination of dielectric relaxation and NMR spectroscopy. In contrast to what one would expect for an acid–base titration, we find strong deviations from quantitative proton transfer from the acid to the base. Even for an excess of the base, multimers consisting of one base and at least two acid molecules are formed, in addition to the occurrence of proton transfer from the acid to the base and simultaneous formation of ion pairs. For equimolar mixtures such multimers constitute about one third of all intermolecular aggregates. Quantitative analysis of our results shows that the acid‐base association constant is only around six times larger than that for the acid binding to an acid‐base dimer, that is, to an already protonated base. Our findings have implications for the interpretation of previous studies of reactive intermediates in organocatalysis and provide a rationale for previously observed nonlinear effects in phosphoric acid catalysis. PMID:28597513

NMR Chemical Exchange as a Probe for Ligand-Binding Kinetics in a Theophylline-Binding RNA Aptamer

PubMed Central

Latham, Michael P.; Zimmermann, Grant R.; Pardi, Arthur

2009-01-01

The apparent on- and off-rate constants for theophylline binding to its RNA aptamer in the absence of Mg2+ were determined here by 2D 1H-1H NMR ZZ-exchange spectroscopy. Analysis of the build-up rate of the exchange cross peaks for several base-paired imino protons in the RNA yielded an apparent kon of 600 M-1 s-1. This small apparent kon results from the free RNA existing as a dynamic equilibrium of inactive states rapidly interconverting with a low population of active species. The data here indicate that the RNA aptamer employs a conformational selection mechanism for binding theophylline in the absence of Mg2+. The kinetic data here also explain a very unusual property of this RNA-theophylline system, slow exchange on the NMR chemical shift timescale for a weak-binding complex. To our knowledge, it is unprecedented to have such a weak binding complex (Kd ≈ 3.0 mM at 15 °C) show slow exchange on the NMR chemical shift timescale, but the results clearly demonstrate that slow exchange and weak binding are readily rationalized by a small kon. Comparisons with other ligand-receptor interactions are presented. PMID:19317486

Rabeprazole: the role of proton pump inhibitors in Helicobacter pylori eradication.

PubMed

Sharara, Ala I

2005-12-01

Proton pump inhibitors have become one of the cornerstones in the treatment of Helicobacter pylori infection. Rabeprazole (Pariet) is a substituted benzimidazole proton pump inhibitor with potent gastric acid suppression properties. Its high acid-base dissociation constant allows activation over a broader pH range, resulting in quick, irreversible binding to the H+/K+-ATPase pump, and a more rapid onset of action compared with omeprazole, lansoprazole and pantoprazole. Unlike other proton pump inhibitors, the metabolism of rabeprazole is primarily via a nonenzymatic reduction to the thioether derivative, and the cytochrome P450 isoenzyme 2C19 is only partly involved in its metabolism. The effect of genetic polymorphism in cytochrome P450 isoenzyme 2C19 on the pharmacokinetics and pharmacodynamics of rabeprazole is therefore limited. In humans, once-daily dosing of 5-40 mg of rabeprazole inhibits gastric acid secretion in a dose-dependent manner. In vitro studies have shown that rabeprazole possesses more potent antibacterial properties against the growth of H. pylori than other proton pump inhibitors. Furthermore, its thioether derivative has more potent inhibitory in vitro activity against the growth and motility of clarithromycin-resistant H. pylori than other proton pump inhibitors or commonly used antimicrobials. Despite these inherent favorable characteristics of rabeprazole, randomized controlled trials have largely shown equivalence amongst proton pump inhibitors when used with two antibiotics in the eradication of H. pylori, with cure rates of 75-89% on an intent-to-treat basis. However, rabeprazole appears to consistently achieve such comparable eradication rates even when used at reduced doses (10 mg twice daily) as part of clarithromycin-based triple therapy.

Modeling solubility and acid-base properties of some amino acids in aqueous NaCl and (CH3)4NCl aqueous solutions at different ionic strengths and temperatures.

PubMed

Bretti, Clemente; Giuffrè, Ottavia; Lando, Gabriele; Sammartano, Silvio

2016-01-01

New potentiometric experiments have been performed in NaCl and in (CH3)4NCl media, to determine the protonation constants, the protonation enthalpy changes and the solubility of six natural α-amino acids, namely Glycine (Gly), Alanine (Ala), Valine (Val), Leucine (Leu), Serine (Ser) and Phenylalanine (Phe). The aim of the work is the rationalization of the protonation thermodynamics (log [Formula: see text], solubility and [Formula: see text]) in NaCl, determining recommended, tentative or provisional values in selected experimental conditions and to report, for the first time, data in a weak interacting medium, as (CH3)4NCl. Literature data analysis was performed selecting the most reliable values, analyzed together with new data here reported. Significant trends and similarities were observed in the behavior of the six amino acids, and in some cases it was possible to determine common parameters for the ionic strength and temperature dependence. In general, the first protonation step, relative to the amino group, is significantly exothermic (average value is [Formula: see text] = -44.5 ± 0.4 kJ mol(-1) at infinite dilution and T = 298.15 K), and the second, relative to the carboxylate group, is fairly close to zero ([Formula: see text] = -2.5 ± 1.6, same conditions). In both cases, the main contribution to the proton binding reaction is mainly entropic in nature. For phenylalanine and leucine, solubility measurements at different concentrations of supporting electrolyte allowed to determine total and specific solubility values, then used to obtain the Setschenow and the activity coefficients of all the species involved in the protonation equilibria. The values of the first protonation constant in (CH3)4NCl are lower than the corresponding values in NaCl, due to the weak interaction between the deprotonated amino group and (CH3)4N(+). In this light, differences between the protonation functions in NaCl and (CH3)4NCl were used for the quantification of the stability of the weak [(CH3)4N(+)-L(-)] complexes that resulted log K = -0.38 ± 0.07 as an average value for the six amino acids.

Proton movement and coupling in the POT family of peptide transporters

PubMed Central

Parker, Joanne L.; Li, Chenghan; Brinth, Allete; Wang, Zhi; Vogeley, Lutz; Solcan, Nicolae; Ledderboge-Vucinic, Gregory; Swanson, Jessica M. J.; Caffrey, Martin; Voth, Gregory A.

2017-01-01

POT transporters represent an evolutionarily well-conserved family of proton-coupled transport systems in biology. An unusual feature of the family is their ability to couple the transport of chemically diverse ligands to an inwardly directed proton electrochemical gradient. For example, in mammals, fungi, and bacteria they are predominantly peptide transporters, whereas in plants the family has diverged to recognize nitrate, plant defense compounds, and hormones. Although recent structural and biochemical studies have identified conserved sites of proton binding, the mechanism through which transport is coupled to proton movement remains enigmatic. Here we show that different POT transporters operate through distinct proton-coupled mechanisms through changes in the extracellular gate. A high-resolution crystal structure reveals the presence of ordered water molecules within the peptide binding site. Multiscale molecular dynamics simulations confirm proton transport occurs through these waters via Grotthuss shuttling and reveal that proton binding to the extracellular side of the transporter facilitates a reorientation from an inward- to outward-facing state. Together these results demonstrate that within the POT family multiple mechanisms of proton coupling have likely evolved in conjunction with variation of the extracellular gate. PMID:29180426

Proton movement and coupling in the POT family of peptide transporters.

PubMed

Parker, Joanne L; Li, Chenghan; Brinth, Allete; Wang, Zhi; Vogeley, Lutz; Solcan, Nicolae; Ledderboge-Vucinic, Gregory; Swanson, Jessica M J; Caffrey, Martin; Voth, Gregory A; Newstead, Simon

2017-12-12

POT transporters represent an evolutionarily well-conserved family of proton-coupled transport systems in biology. An unusual feature of the family is their ability to couple the transport of chemically diverse ligands to an inwardly directed proton electrochemical gradient. For example, in mammals, fungi, and bacteria they are predominantly peptide transporters, whereas in plants the family has diverged to recognize nitrate, plant defense compounds, and hormones. Although recent structural and biochemical studies have identified conserved sites of proton binding, the mechanism through which transport is coupled to proton movement remains enigmatic. Here we show that different POT transporters operate through distinct proton-coupled mechanisms through changes in the extracellular gate. A high-resolution crystal structure reveals the presence of ordered water molecules within the peptide binding site. Multiscale molecular dynamics simulations confirm proton transport occurs through these waters via Grotthuss shuttling and reveal that proton binding to the extracellular side of the transporter facilitates a reorientation from an inward- to outward-facing state. Together these results demonstrate that within the POT family multiple mechanisms of proton coupling have likely evolved in conjunction with variation of the extracellular gate. Copyright © 2017 the Author(s). Published by PNAS.

Squeezing at Entrance of Proton Transport Pathway in Proton-translocating Pyrophosphatase upon Substrate Binding*

PubMed Central

Huang, Yun-Tzu; Liu, Tseng-Huang; Lin, Shih-Ming; Chen, Yen-Wei; Pan, Yih-Jiuan; Lee, Ching-Hung; Sun, Yuh-Ju; Tseng, Fan-Gang; Pan, Rong-Long

2013-01-01

Homodimeric proton-translocating pyrophosphatase (H+-PPase; EC 3.6.1.1) is indispensable for many organisms in maintaining organellar pH homeostasis. This unique proton pump couples the hydrolysis of PPi to proton translocation across the membrane. H+-PPase consists of 14–16 relatively hydrophobic transmembrane domains presumably for proton translocation and hydrophilic loops primarily embedding a catalytic site. Several highly conserved polar residues located at or near the entrance of the transport pathway in H+-PPase are essential for proton pumping activity. In this investigation single molecule FRET was employed to dissect the action at the pathway entrance in homodimeric Clostridium tetani H+-PPase upon ligand binding. The presence of the substrate analog, imidodiphosphate mediated two sites at the pathway entrance moving toward each other. Moreover, single molecule FRET analyses after the mutation at the first proton-carrying residue (Arg-169) demonstrated that conformational changes at the entrance are conceivably essential for the initial step of H+-PPase proton translocation. A working model is accordingly proposed to illustrate the squeeze at the entrance of the transport pathway in H+-PPase upon substrate binding. PMID:23720778

Calculation of site affinity constants and cooperativity coefficients for binding of ligands and/or protons to macromolecules. II. Relationships between chemical model and partition function algorithm.

PubMed

Fisicaro, E; Braibanti, A; Lamb, J D; Oscarson, J L

1990-05-01

The relationships between the chemical properties of a system and the partition function algorithm as applied to the description of multiple equilibria in solution are explained. The partition functions ZM, ZA, and ZH are obtained from powers of the binary generating functions Jj = (1 + kappa j gamma j,i[Y])i tau j, where i tau j = p tau j, q tau j, or r tau j represent the maximum number of sites in sites in class j, for Y = M, A, or H, respectively. Each term of the generating function can be considered an element (ij) of a vector Jj and each power of the cooperativity factor gamma ij,i can be considered an element of a diagonal cooperativity matrix gamma j. The vectors Jj are combined in tensor product matrices L tau = (J1) [J2]...[Jj]..., thus representing different receptor-ligand combinations. The partition functions are obtained by summing elements of the tensor matrices. The relationship of the partition functions with the total chemical amounts TM, TA, and TH has been found. The aim is to describe the total chemical amounts TM, TA, and TH as functions of the site affinity constants kappa j and cooperativity coefficients bj. The total amounts are calculated from the sum of elements of tensor matrices Ll. Each set of indices (pj..., qj..., rj...) represents one element of a tensor matrix L tau and defines each term of the summation. Each term corresponds to the concentration of a chemical microspecies. The distinction between microspecies MpjAqjHrj with ligands bound on specific sites and macrospecies MpAqHR corresponding to a chemical stoichiometric composition is shown. The translation of the properties of chemical model schemes into the algorithms for the generation of partition functions is illustrated with reference to a series of examples of gradually increasing complexity. The equilibria examined concern: (1) a unique class of sites; (2) the protonation of a base with two classes of sites; (3) the simultaneous binding of ligand A and proton H to a macromolecule or receptor M with four classes of sites; and (4) the binding to a macromolecule M of ligand A which is in turn a receptor for proton H. With reference to a specific example, it is shown how a computer program for least-squares refinement of variables kappa j and bj can be organized. The chemical model from the free components M, A, and H to the saturated macrospecies MpAQHR, with possible complex macrospecies MpAq and AHR, is defined first.(ABSTRACT TRUNCATED AT 250 WORDS)

Temperature dependence of (+)-catechin pyran ring proton coupling constants as measured by NMR and modeled using GMMX search methodology

Treesearch

Fred L. Tobiason; Stephen S. Kelley; M. Mark Midland; Richard W. Hemingway

1997-01-01

The pyran ring proton coupling constants for (+)-catechin have been experimentally determined in deuterated methanol over a temperature range of 213 K to 313 K. The experimental coupling constants were simulated to 0.04 Hz on the average at a 90 percent confidence limit using a LAOCOON method. The temperature dependence of the coupling constants was reproduced from the...

Binding of oxytocin and 8-arginine-vasopressin to neurophysin studied by /sup 15/N NMR using magnetization transfer and indirect detection via protons

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

Live, D.H.; Cowburn, D.

1987-10-06

NMR was used to monitor the binding to neurophysin of oxytocin and 8-arginine-vasopressin, /sup 15/N labeling being used to identify specific backbone /sup 15/N and /sup 1/H signals. The most significant effects of binding were large downfield shifts in the amino nitrogen resonance of Phe-3 of vasopressin and in its associated proton, providing evidence that the peptide bond between residues 2 and 3 of the hormones is hydrogen-bonded to the protein within hormone-neurophysin complexes. Suggestive evidence for hydrogen bonding of the amino nitrogen of Tyr-2 was also obtained in the form of decreased proton exchange rates on binding; however, themore » chemical shift changes of this nitrogen and its associated proton indicated that such hydrogen bonding, if present, is probably weak. Shifts in the amino nitrogen of Asn-5 and in the -NH protons of both Asn-5 and Cys-6 demonstrated that these residues are significantly perturbed by binding, suggesting conformational changes of the ring on binding and/or the presence of binding sites on the hormone outside the 1-3 region. No support was obtained for the thesis that there is a significant second binding site for vasopressin on each neutrophysin chain. The behavior of both oxytocin and vasopressin on binding was consistent with formation of 1:1 complexes in slow exchange with the free state under most pH conditions. At low pH there was evidence of an increased exchange rate. Additionally, broadening of /sup 15/N resonances in the bound state at low pH occurred without a corresponding change in the resonances of equilibrating free hormone. The results suggest significant conformational alteration in neurophysin-hormone complexes at low pH possibly associated with protonation of the carboxyl group of the hormone-protein salt bridge.« less

Positive and negative ion mode ESI-MS and MS/MS for studying drug-DNA complexes

NASA Astrophysics Data System (ADS)

Rosu, Frédéric; Pirotte, Sophie; Pauw, Edwin De; Gabelica, Valérie

2006-07-01

We report systematic investigation of duplex DNA complexes with minor groove binders (Hoechsts 33258 and 33342, netropsin and DAPI) and intercalators (daunomycin, doxorubicin, actinomycin D, ethidium, cryptolepine, neocryptolepine, m-Amsacrine, proflavine, ellipticine and mitoxantrone) by ESI-MS and ESI-MS/MS in the negative ion mode and in the positive ion mode. The apparent solution phase equilibrium binding constants can be determined by measuring relative intensities in the ESI-MS spectrum. While negative ion mode gives reliable results, positive ion mode gives a systematic underestimation of the binding constants and even a complete suppression of the complexes for intercalators lacking functional groups capable of interacting in the grooves. In the second part of the paper we systematically compare MS/MS fragmentation channels and breakdown curves in the positive and the negative modes, and discuss the possible uses and caveats of MS/MS in drug-DNA complexes. In the negative mode, the drugs can be separated in three groups: (1) those that leave the complex with no net charge; (2) those that leave the complex with a negative charge; and (3) those that remain attached on the strands upon dissociation of the duplex due to their positive charge. In the positive ion mode, all complexes fragment via the loss of protonated drug. Information on the stabilization of the complex by drug-DNA noncovalent interactions can be obtained straightforwardly only in the case of neutral drug loss. In all other cases, proton affinity (in the positive ion mode), gas-phase basicity (in the negative ion mode) and coulombic repulsion are the major factors influencing the fragmentation channel and the dissociation kinetics.

An enzyme kinetics study of the pH dependence of chloride activation of oxygen evolution in photosystem II.

PubMed

Baranov, Sergei; Haddy, Alice

2017-03-01

Oxygen evolution by photosystem II (PSII) involves activation by Cl - ion, which is regulated by extrinsic subunits PsbQ and PsbP. In this study, the kinetics of chloride activation of oxygen evolution was studied in preparations of PSII depleted of the PsbQ and PsbP subunits (NaCl-washed and Na 2 SO 4 /pH 7.5-treated) over a pH range from 5.3 to 8.0. At low pH, activation by chloride was followed by inhibition at chloride concentrations >100 mM, whereas at high pH activation continued as the chloride concentration increased above 100 mM. Both activation and inhibition were more pronounced at lower pH, indicating that Cl - binding depended on protonation events in each case. The simplest kinetic model that could account for the complete data set included binding of Cl - at two sites, one for activation and one for inhibition, and four protonation steps. The intrinsic (pH-independent) dissociation constant for Cl - activation, K S , was found to be 0.9 ± 0.2 mM for both preparations, and three of the four pK a s were determined, with the fourth falling below the pH range studied. The intrinsic inhibition constant, K I , was found to be 64 ± 2 and 103 ± 7 mM for the NaCl-washed and Na 2 SO 4 /pH7.5-treated preparations, respectively, and is considered in terms of the conditions likely to be present in the thylakoid lumen. This enzyme kinetics analysis provides a more complete characterization of chloride and pH dependence of O 2 evolution activity than has been previously presented.

Isothermal lipid phase transitions.

PubMed

Cevc, G

1991-03-01

In liotropic lipid systems phase transitions can be induced isothermally by changing the solvent concentration or composition; alternatively, lipid composition can be modified by (bio)chemical means. The probability for isothermal phase transitions increases with the decreasing transition entropy; it is proportional to the magnitude of the transition temperature shift caused by transformation-inducing system variation. Manipulations causing large thermodynamic effects, such as lipid (de)hydration, binding of protons or divalent ions and macromolecular adsorption, but also close bilayer approach are, therefore, likely to cause structural lipid change(s) at a constant temperature. Net lipid charges enhance the membrane susceptibility to salt-induced isothermal phase transitions; a large proportion of this effect is due to the bilayer dehydration, however, rather than being a consequence of the decreased Coulombic electrostatic interactions. Membrane propensity for isothermal phase transitions, consequently, always increases with the hydrophilicity of the lipid heads, as well as with the desaturation and shortening of the lipid chains. Upon a phase change at a constant temperature, some of the interfacially bound solutes (e.g. protons or calcium) are released in the solution. Membrane permeability and fusogenicity simultaneously increase. In mixed systems, isothermal phase transitions, moreover, may result in lateral phase separation. All this opens up ways for the involvement of isothermal phase transitions in the regulation of biological processes.

Kinetics of binding of chicken cystatin to papain.

PubMed

Björk, I; Alriksson, E; Ylinenjärvi, K

1989-02-21

The kinetics of binding of chicken cystatin to papain were studied by stopped-flow fluorometry under pseudo-first-order conditions, i.e., with an excess of inhibitor. All reactions showed first-order behavior, and the observed pseudo-first-order rate constant increased linearly with the cystatin concentration up to the highest concentration that could be studied, 35 microM. The analyses thus provided no evidence for a limiting rate resulting from a conformational change stabilizing an initial encounter complex, in contrast with previous studies of reactions between serine proteinases and their protein inhibitors. The second-order association rate constant for complex formation was 9.9 X 10(6) M-1 s-1 at 25 degrees C, pH 7.4, I = 0.15, for both forms of cystatin, 1 and 2. This value approaches that expected for a diffusion-controlled rate. The temperature dependence of the association rate constant gave an enthalpy of activation at 25 degrees C of 31.5 kJ mol-1 and an entropy of activation at 25 degrees C of -7 J K-1 mol-1, compatible with no appreciable conformational change during the reaction. The association rate constant was independent of pH between pH 6 and 8 but decreased at lower and higher pH in a manner consistent with involvement of an unprotonated acid group with a pKa of 4-4.5 and a protonated basic group with a pKa of 9-9.5 in the interaction. The association rate constant was unaffected by ionic strengths between 0.15 and 1.0 but decreased somewhat at lower ionic strengths. Incubation of the complex between cystatin 2 and papain with an excess of cystatin 1 resulted in slow displacement of cystatin 2 from the complex.(ABSTRACT TRUNCATED AT 250 WORDS)

Route, mechanism, and implications of proton import during Na+/K+ exchange by native Na+/K+-ATPase pumps

PubMed Central

Vedovato, Natascia

2014-01-01

A single Na+/K+-ATPase pumps three Na+ outwards and two K+ inwards by alternately exposing ion-binding sites to opposite sides of the membrane in a conformational sequence coupled to pump autophosphorylation from ATP and auto-dephosphorylation. The larger flow of Na+ than K+ generates outward current across the cell membrane. Less well understood is the ability of Na+/K+ pumps to generate an inward current of protons. Originally noted in pumps deprived of external K+ and Na+ ions, as inward current at negative membrane potentials that becomes amplified when external pH is lowered, this proton current is generally viewed as an artifact of those unnatural conditions. We demonstrate here that this inward current also flows at physiological K+ and Na+ concentrations. We show that protons exploit ready reversibility of conformational changes associated with extracellular Na+ release from phosphorylated Na+/K+ pumps. Reversal of a subset of these transitions allows an extracellular proton to bind an acidic side chain and to be subsequently released to the cytoplasm. This back-step of phosphorylated Na+/K+ pumps that enables proton import is not required for completion of the 3 Na+/2 K+ transport cycle. However, the back-step occurs readily during Na+/K+ transport when external K+ ion binding and occlusion are delayed, and it occurs more frequently when lowered extracellular pH raises the probability of protonation of the externally accessible carboxylate side chain. The proton route passes through the Na+-selective binding site III and is distinct from the principal pathway traversed by the majority of transported Na+ and K+ ions that passes through binding site II. The inferred occurrence of Na+/K+ exchange and H+ import during the same conformational cycle of a single molecule identifies the Na+/K+ pump as a hybrid transporter. Whether Na+/K+ pump–mediated proton inflow may have any physiological or pathophysiological significance remains to be clarified. PMID:24688018

Producing the deuteron in stars: anthropic limits on fundamental constants

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

Barnes, Luke A.; Lewis, Geraint F., E-mail: luke.barnes@sydney.edu.au, E-mail: gfl@physics.usyd.edu.au

2017-07-01

Stellar nucleosynthesis proceeds via the deuteron (D), but only a small change in the fundamental constants of nature is required to unbind it. Here, we investigate the effect of altering the binding energy of the deuteron on proton burning in stars. We find that the most definitive boundary in parameter space that divides probably life-permitting universes from probably life-prohibiting ones is between a bound and unbound deuteron. Due to neutrino losses, a ball of gas will undergo rapid cooling or stabilization by electron degeneracy pressure before it can form a stable, nuclear reaction-sustaining star. We also consider a less-bound deuteron,more » which changes the energetics of the pp and pep reactions. The transition to endothermic pp and pep reactions, and the resulting beta-decay instability of the deuteron, do not seem to present catastrophic problems for life.« less

Application of physical adsorption thermodynamics to heterogeneous chemistry on polar stratospheric clouds

NASA Technical Reports Server (NTRS)

Elliott, Scott; Turco, Richard P.; Toon, Owen B.; Hamill, Patrick

1991-01-01

Laboratory isotherms for the binding of several nonheterogeneously active atmospheric gases and for HCl to water ice are translated into adsorptive equilibrium constants and surface enthalpies. Extrapolation to polar conditions through the Clausius Clapeyron relation yields coverage estimates below the percent level for N2, Ar, CO2, and CO, suggesting that the crystal faces of type II stratospheric cloud particles may be regarded as clean with respect to these species. For HCl, and perhaps HF and HNO3, estimates rise to several percent, and the adsorbed layer may offer acid or proton sources alternate to the bulk solid for heterogeneous reactions with stratospheric nitrates. Measurements are lacking for many key atmospheric molecules on water ice, and almost entirely for nitric acid trihydrate as substrate. Adsorptive equilibria enter into gas to particle mass flux descriptions, and the binding energy determines rates for desorption of, and encounter between, potential surface reactants.

Complexity in Acid-Base Titrations: Multimer Formation Between Phosphoric Acids and Imines.

PubMed

Malm, Christian; Kim, Heejae; Wagner, Manfred; Hunger, Johannes

2017-08-10

Solutions of Brønsted acids with bases in aprotic solvents are not only common model systems to study the fundamentals of proton transfer pathways but are also highly relevant to Brønsted acid catalysis. Despite their importance the light nature of the proton makes characterization of acid-base aggregates challenging. Here, we track such acid-base interactions over a broad range of relative compositions between diphenyl phosphoric acid and the base quinaldine in dichloromethane, by using a combination of dielectric relaxation and NMR spectroscopy. In contrast to what one would expect for an acid-base titration, we find strong deviations from quantitative proton transfer from the acid to the base. Even for an excess of the base, multimers consisting of one base and at least two acid molecules are formed, in addition to the occurrence of proton transfer from the acid to the base and simultaneous formation of ion pairs. For equimolar mixtures such multimers constitute about one third of all intermolecular aggregates. Quantitative analysis of our results shows that the acid-base association constant is only around six times larger than that for the acid binding to an acid-base dimer, that is, to an already protonated base. Our findings have implications for the interpretation of previous studies of reactive intermediates in organocatalysis and provide a rationale for previously observed nonlinear effects in phosphoric acid catalysis. © 2017 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA.

Steric Effects on the Binding of Phosphate and Polyphosphate Anions by Zinc(II) and Copper(II) Dinuclear Complexes of m-Xylyl-bis-cyclen.

PubMed

Esteves, Catarina V; Esteban-Gómez, David; Platas-Iglesias, Carlos; Tripier, Raphaël; Delgado, Rita

2018-05-11

The triethylbenzene-bis-cyclen (cyclen = 1,4,7,10-tetraazacyclododecane) compound (tbmce) was designed with an imposed structural rigidity at the m-xylyl spacer to be compared to a less restrained and known parent compound (bmce). The framework of both compounds differs only in the substituents of the m-xylyl spacer. The study was centered in the differences observed in the acid-base reactions of both compounds, their copper(II) and zinc(II) complexation behaviors, as well as in the uptake of phosphate and polyphosphate anions (HPPi 3- , ATP 4- , ADP 3- , AMP 2- , PhPO 4 2- , and HPO 4 2- ). On the one hand, the acid-base reactions showed lower values for the third and fourth protonation constants of tbmce than for bmce, suggesting that the ethyl groups of the spacer in tbmce force the two cyclen units to more conformational restricted positions. On the other hand, the stability constant values for copper(II) and zinc(II) complexes revealed that bmce is a better chelator than tbmce pointing out to additional conformational restraints imposed by the triethylbenzene spacer. The binding studies of phosphates by the dinuclear copper(II) and zinc(II) complexes showed much smaller effective association constants for the dicopper complexes. Single-crystal X-ray and computational (density functional theory) studies suggest that anion binding promotes the formation of tetranuclear entities in which anions are bridging the metal centers. Our studies also revealed the dinuclear zinc(II) complex of bmce as a promising receptor for phosphate anions, with the largest effective association constant of 5.94 log units being observed for the formation of [Zn 2 bmce(HPPi)] + . Accordingly, a colorimetric study via an indicator displacement assay to detect phosphates in aqueous solution found that the [Zn 2 bmce] 4+ complex acts as the best receptor for pyrophosphate displaying a detection limit of 2.5 nM by changes visible to naked eye.

Rational design of a conformation-switchable Ca2+- and Tb3+-binding protein without the use of multiple coupled metal-binding sites.

PubMed

Li, Shunyi; Yang, Wei; Maniccia, Anna W; Barrow, Doyle; Tjong, Harianto; Zhou, Huan-Xiang; Yang, Jenny J

2008-10-01

Ca2+, as a messenger of signal transduction, regulates numerous target molecules via Ca2+-induced conformational changes. Investigation into the determinants for Ca2+-induced conformational change is often impeded by cooperativity between multiple metal-binding sites or protein oligomerization in naturally occurring proteins. To dissect the relative contributions of key determinants for Ca2+-dependent conformational changes, we report the design of a single-site Ca2+-binding protein (CD2.trigger) created by altering charged residues at an electrostatically sensitive location on the surface of the host protein rat Cluster of Differentiation 2 (CD2).CD2.trigger binds to Tb3+ and Ca2+ with dissociation constants of 0.3 +/- 0.1 and 90 +/- 25 microM, respectively. This protein is largely unfolded in the absence of metal ions at physiological pH, but Tb3+ or Ca2+ binding results in folding of the native-like conformation. Neutralization of the charged coordination residues, either by mutation or protonation, similarly induces folding of the protein. The control of a major conformational change by a single Ca2+ ion, achieved on a protein designed without reliance on sequence similarity to known Ca2+-dependent proteins and coupled metal-binding sites, represents an important step in the design of trigger proteins.

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

PubMed

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

2013-09-17

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

Multiscale Simulations Reveal Key Aspects of the Proton Transport Mechanism in the ClC-ec1 Antiporter

PubMed Central

Lee, Sangyun; Swanson, Jessica M.J.; Voth, Gregory A.

2016-01-01

Multiscale reactive molecular dynamics simulations are used to study proton transport through the central region of ClC-ec1, a widely studied ClC transporter that enables the stoichiometric exchange of 2 Cl– ions for 1 proton (H+). It has long been known that both Cl– and proton transport occur through partially congruent pathways, and that their exchange is strictly coupled. However, the nature of this coupling and the mechanism of antiporting remain topics of debate. Here multiscale simulations have been used to characterize proton transport between E203 (Gluin) and E148 (Gluex), the internal and external intermediate proton binding sites, respectively. Free energy profiles are presented, explicitly accounting for the binding of Cl– along the central pathway, the dynamically coupled hydration changes of the central region, and conformational changes of Gluin and Gluex. We find that proton transport between Gluin and Gluex is possible in both the presence and absence of Cl– in the central binding site, although it is facilitated by the anion presence. These results support the notion that the requisite coupling between Cl– and proton transport occurs elsewhere (e.g., during proton uptake or release). In addition, proton transport is explored in the E203K mutant, which maintains proton permeation despite the substitution of a basic residue for Gluin. This collection of calculations provides for the first time, to our knowledge, a detailed picture of the proton transport mechanism in the central region of ClC-ec1 at a molecular level. PMID:27028643

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

Modification by protons of frog skeletal muscle KATP channels: effects on ion conduction and nucleotide inhibition.

PubMed Central

Vivaudou, M; Forestier, C

1995-01-01

1. The molecular mechanisms underlying pH regulation of skeletal muscle ATP-sensitive K+ (KATP) channels were studied using the patch clamp technique in the inside-out configuration. Two effects of intracellular protons were studied in detail: the decrease in magnitude of single-channel currents and the increase in open probability (Po) of nucleotide-inhibited channels. 2. The pH dependence of inward unit currents under different ionic conditions was in poor agreement with either a direct block of the pore by protons or an indirect proton-induced conformational change, but was compatible with the protonation of surface charges located near the cytoplasmic entrance of the pore. This latter electrostatic mechanism was modelled using Gouy-Chapman-Stern theory, which predicted the data accurately with a surface charge density of about 0.1 negative elementary charges per square nanometre and a pK (pH value for 50% effect) value for protonation of these charges of 6.25. The same mechanism, i.e. neutralization of negative surface charges by cation binding, could also account for the previously reported reduction of inward unit currents by Mg2+. 3. Intracellular alkalization did not affect Po of the KATP channels. Acidification increased Po. In the presence of 0.1 mM ATP (no Mg2+), the channel activation vs. pH relationship could be fitted with a sigmoid curve with a Hill coefficient slightly above 2 and a pK value of 6. This latter value was dependent on the ATP concentration, decreasing from 6.3 in 30 microM ATP to 5.3 in 1 microM ATP. 4. Conversely, the channel inhibition vs. ATP concentration curve was shifted to the right when the pH was lowered. At pH 7.1, the ATP concentration causing half-maximal inhibition was about 10 microM. At pH 5.4, it was about 400 microM. The Hill coefficient values remained slightly below 2. Similar effects were observed when ADP was used as the inhibitory nucleotide. 5. These results confirm that a reciprocal competitive link exists between proton and nucleotide binding sites. Quantitatively, they are in full agreement with a steady-state model of a KATP channel possessing four identical protonation sites (microscopic pK, 6) allosterically connected to the channel open state and two identical nucleotide sites (microscopic ATP dissociation constant, approximately 30 microM) connected to the closed state. Images Figure 13 PMID:7473225

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

Constant pH Molecular Dynamics of Proteins in Explicit Solvent with Proton Tautomerism

PubMed Central

Goh, Garrett B.; Hulbert, Benjamin S.; Zhou, Huiqing; Brooks, Charles L.

2015-01-01

pH is a ubiquitous regulator of biological activity, including protein-folding, protein-protein interactions and enzymatic activity. Existing constant pH molecular dynamics (CPHMD) models that were developed to address questions related to the pH-dependent properties of proteins are largely based on implicit solvent models. However, implicit solvent models are known to underestimate the desolvation energy of buried charged residues, increasing the error associated with predictions that involve internal ionizable residue that are important in processes like hydrogen transport and electron transfer. Furthermore, discrete water and ions cannot be modeled in implicit solvent, which are important in systems like membrane proteins and ion channels. We report on an explicit solvent constant pH molecular dynamics framework based on multi-site λ-dynamics (CPHMDMSλD). In the CPHMDMSλD framework, we performed seamless alchemical transitions between protonation and tautomeric states using multi-site λ-dynamics, and designed novel biasing potentials to ensure that the physical end-states are predominantly sampled. We show that explicit solvent CPHMDMSλD simulations model realistic pH-dependent properties of proteins such as the Hen-Egg White Lysozyme (HEWL), binding domain of 2-oxoglutarate dehydrogenase (BBL) and N-terminal domain of ribosomal L9 (NTL9), and the pKa predictions are in excellent agreement with experimental values, with a RMSE ranging from 0.72 to 0.84 pKa units. With the recent development of the explicit solvent CPHMDMSλD framework for nucleic acids, accurate modeling of pH-dependent properties of both major class of biomolecules – proteins and nucleic acids is now possible. PMID:24375620

Potentiometric and NMR complexation studies of phenylboronic acid PBA and its aminophosphonate analog with selected catecholamines

NASA Astrophysics Data System (ADS)

Ptak, Tomasz; Młynarz, Piotr; Dobosz, Agnieszka; Rydzewska, Agata; Prokopowicz, Monika

2013-05-01

Boronic acids are a class of intensively explored compounds, which according to their specific properties have been intensively explored in last decades. Among them phenylboronic acids and their derivatives are most frequently examined as receptors for diverse carbohydrates. In turn, there is a large gap in basic research concerning complexation of catecholamines by these compounds. Therefore, we decided to undertake studies on interaction of chosen catecholamines, namely: noradrenaline (norephinephrine), dopamine, L-DOPA, DOPA-P (phosphonic analog of L-DOPA) and catechol, with simple phenyl boronic acid PBA by means of potentiometry and NMR spectroscopy. For comparison, the binding properties of recently synthesized phenylboronic receptor 1 bearing aminophosphonate function in meta-position were investigated and showed promising ability to bind catecholamines. The protonation and stability constants of PBA and receptor 1 complexes were examined by potentiometry. The obtained results demonstrated that PBA binds the catecholamines with the following affinity order: noradrenaline ⩾ dopamine ≈ L-DOPA > catechol > DOPA-P, while its modified analog 1 reveals slightly different preferences: dopamine > noradrenaline > catechol > L-DOPA > DOPA-P.

Terminal base pairs of oligodeoxynucleotides: imino proton exchange and fraying.

PubMed

Nonin, S; Leroy, J L; Guéron, M

1995-08-22

We have estimated the dissociation constant of the terminal base pairs of the B-DNA duplexes formed by 5'-d(CGCGATCGCG) and 5'-d(TAGCGCTA) by two methods, one based on the change in imino proton chemical shift with temperature and the other on the apparent pK shift of the imino proton, as monitored by the change in chemical shift of aromatic protons. These methods do not rely on imino proton exchange, whose rate was also measured. (1) The effect of ammonia on the imino proton exchange rate of the terminal pair of the 5'-d(CGCGATCGCG) duplex is 67 times less than on the isolated nucleoside. This provides an upper limit on the exchange rate from the closed pair. In fact, the effect is just as predicted from the dissociation constant, assuming that there is no exchange at all from the closed pair and that, as has been argued previously, external catalysts act on the open state as they do on the isolated nucleoside. The inhibition of catalyzed proton exchange in the closed pair, despite exposure of one face of the pair to solvent, is a new feature of the exchange process. It will allow determination of the dissociation constant of terminal pairs from the exchange rate. (2) Intrinsic catalysis of proton exchange is less efficient for the terminal pair than for an internal one. A possible explanation is that proton transfer across the water bridge responsible for intrinsic catalysis is slower, as expected if the open-state separation of the bases is larger in a terminal pair. This observation may lead to a direct method for the study of fraying. (3) At 0 degrees C, the dissociation constant of the second pair of the 5'-d(CGCGATCGCG) duplex is close to the square of the constant for the terminal pair, as predicted from a simple model of fraying. The enthalpy and entropy of opening of the terminal pairs may be compared with those of nearest neighbor interactions derived from calorimetry [Breslauer, K. J., et al. (1986) Proc. Natl. Acad. Sci. U.S.A. 83, 3746-3750].

Substitution and protonation effects on spin-spin coupling constants in prototypical aromatic rings: C6H6, C5H5N and C5H5P.

PubMed

Del Bene, Janet E; Elguero, José

2006-08-01

Ab initio equation-of-motion coupled cluster calculations have been carried out to evaluate one-, two-, and three-bond 13C-13C, 15N-13C, 31P-13C coupling constants in benzene, pyridine, pyridinium, phosphinine, and phosphininium. The introduction of N or P heteroatoms into the aromatic ring not only changes the magnitudes of the corresponding X-C coupling constants (J, for X = C, N, or P) but also the signs and magnitudes of corresponding reduced coupling constants (K). Protonation of the heteroatoms also produces dramatic changes in coupling constants and, by removing the lone pair of electrons from the sigma-electron framework, leads to the same signs for corresponding reduced coupling constants for benzene, pyridinium, and phosphininium. C-C coupling constants are rather insensitive to the presence of the heteroatoms and protonation. All terms that contribute to the total coupling constant (except for the diamagnetic spin-orbit (DSO) term) must be computed if good agreement with experimental data is to be obtained. Copyright 2006 John Wiley & Sons, Ltd.

Binding of pixantrone to DNA at CpA dinucleotide sequences and bulge structures.

PubMed

Konda, Shyam K; Wang, Haiqiang; Cutts, Suzanne M; Phillips, Don R; Collins, J Grant

2015-06-07

The binding of the anti-cancer drug pixantrone to three oligonucleotide sequences, d(TCATATGA)2, d(CCGAGAATTCCGG)2 {double bulge = DB} and the non-self complementary d(TACGATGAGTA) : d(TACCATCGTA) {single bulge = SB}, has been studied by NMR spectroscopy and molecular modelling. The upfield shifts observed for the aromatic resonances of pixantrone upon addition of the drug to each oligonucleotide confirmed the drug bound by intercalation. For the duplex sequence d(TCATATGA)2, NOEs were observed from the pixantrone aromatic H7/8 and aliphatic Ha/Hb protons to the H6/H8 and H1' protons of the C2, A3, T6 and G7 nucleotides, demonstrating that pixantrone preferentially binds at the symmetric CpA sites. However, weaker NOEs observed to various protons from the T4 and A5 residues indicated alternative minor binding sites. NOEs from the H7/H8 and Ha/Hb protons to both major (H6/H8) and minor groove (H1') protons indicated approximately equal proportions of intercalation was from the major and minor groove at the CpA sites. Intermolecular NOEs were observed between the H7/H8 and H4 protons of pixantrone and the A4H1' and G3H1' protons of the oligonucleotide that contains two symmetrically related bulge sites (DB), indicative of binding at the adenine bulge sites. For the oligonucleotide that only contains a single bulge site (SB), NOEs were observed from pixantrone protons to the SB G7H1', A8H1' and G9H1' protons, confirming that the drug bound selectively at the adenine bulge site. A molecular model of pixantrone-bound SB could be constructed with the drug bound from the minor groove at the A8pG9 site that was consistent with the observed NMR data. The results demonstrate that pixantrone preferentially intercalates at adenine bulge sites, compared to duplex DNA, and predominantly from the minor groove.

An NMR strategy to detect conformational differences in a protein complexed with highly analogous inhibitors in solution.

PubMed

Persons, John D; Khan, Shahid N; Ishima, Rieko

2018-04-12

This manuscript presents an NMR strategy to investigate conformational differences in protein-inhibitor complexes, when the inhibitors tightly bind to a protein at sub-nanomolar dissociation constants and are highly analogous to each other. Using HIV-1 protease (PR), we previously evaluated amide chemical shift differences, ΔCSPs, of PR bound to darunavir (DRV) compared to PR bound to several DRV analogue inhibitors, to investigate subtle but significant long-distance conformation changes caused by the inhibitor's chemical moiety variation [Khan, S. N., Persons, J. D. Paulsen, J. L., Guerrero, M., Schiffer, C. A., Kurt-Yilmaz, N., and Ishima, R., Biochemistry, (2018), 57, 1652-1662]. However, ΔCSPs are not ideal for investigating subtle PR-inhibitor interface differences because intrinsic differences in the electron shielding of the inhibitors affect protein ΔCSPs. NMR relaxation is also not suitable as it is not sensitive enough to detect small conformational differences in rigid regions among similar PR-inhibitor complexes. Thus, to gain insight into conformational differences at the inhibitor-protein interface, we recorded 15 N-half filtered NOESY spectra of PR bound to two highly analogous inhibitors and assessed NOEs between PR amide protons and inhibitor protons, between PR amide protons and hydroxyl side chains, and between PR amide protons and water protons. We also verified the PR amide-water NOEs using 2D water-NOE/ROE experiments. Differences in water-amide proton NOE peaks, possibly due to amide-protein hydrogen bonds, were observed between subunit A and subunit B, and between the DRV-bound form and an analogous inhibitor-bound form, which may contribute to remote conformational changes. Copyright © 2018 Elsevier Inc. All rights reserved.

The Hydrogenase Activity of the Molybdenum/Copper-containing Carbon Monoxide Dehydrogenase of Oligotropha carboxidovorans*

PubMed Central

Wilcoxen, Jarett; Hille, Russ

2013-01-01

The reaction of the air-tolerant CO dehydrogenase from Oligotropha carboxidovorans with H2 has been examined. Like the Ni-Fe CO dehydrogenase, the enzyme can be reduced by H2 with a limiting rate constant of 5.3 s−1 and a dissociation constant Kd of 525 μm; both kred and kred/Kd, reflecting the breakdown of the Michaelis complex and the reaction of free enzyme with free substrate in the low [S] regime, respectively, are largely pH-independent. During the reaction with H2, a new EPR signal arising from the Mo/Cu-containing active site of the enzyme is observed which is distinct from the signal seen when the enzyme is reduced by CO, with greater g anisotropy and larger hyperfine coupling to the active site 63,65Cu. The signal also exhibits hyperfine coupling to at least two solvent-exchangeable protons of bound substrate that are rapidly exchanged with solvent. Proton coupling is also evident in the EPR signal seen with the dithionite-reduced native enzyme, and this coupling is lost in the presence of bicarbonate. We attribute the coupled protons in the dithionite-reduced enzyme to coordinated water at the copper site in the native enzyme and conclude that bicarbonate is able to displace this water from the copper coordination sphere. On the basis of our results, a mechanism for H2 oxidation is proposed which involves initial binding of H2 to the copper of the binuclear center, displacing the bound water, followed by sequential deprotonation through a copper-hydride intermediate to reduce the binuclear center. PMID:24165123

Effects of organic ligands on fractionation of rare earth elements (REEs) in hydroponic plants: an application to the determination of binding capacities by humic acid for modeling.

PubMed

Ding, ShiMing; Liang, Tao; Zhang, ChaoSheng; Yan, JunCai; Zhang, ZiLi

2006-12-01

Previous studies have revealed the fractionation processes of rare earth elements (REEs) in hydroponic plants, with a heavy REE (HREE, the elements from Gd to Lu) enrichment in leaves. In this study, effects on the HREE enrichment in soybean leaves with additions of carboxylic acids (acetate, malate, citrate, NTA, EDTA and DTPA) and two soil humic acids (HAs) were investigated. REE speciation in carboxylic acid and HA solutions was simulated using Visual MINTEQ and Model V, respectively. The results showed that the effects caused by carboxylic acids were strongly dependent on the differences between their binding strengths for light REEs (LREEs, the elements from La to Eu) and those for HREEs. A good correlation existed between these effects and the changes of free REE ions in solutions. This relationship was also observed for the HA treatments, provided that the intrinsic equilibrium constants of REEs for cation-proton exchange with HA (i.e., pK(MHA)) in Model V were estimated using a free-energy relationship with the stability constants for REE complexation with lactic acid. It is suggested that this set of pK(MHA) values is more suitable for use in Model V for the simulation of REE complexation with HA.

Visualizing the kinetic power stroke that drives proton-coupled Zn(II) transport

PubMed Central

Gupta, Sayan; Chai, Jin; Cheng, Jie; D'Mello, Rhijuta; Chance, Mark R.; Fu, Dax

2014-01-01

The proton gradient is a principal energy source for respiration-dependent active transport, but the structural mechanisms of proton-coupled transport processes are poorly understood. YiiP is a proton-coupled zinc transporter found in the cytoplasmic membrane of E. coli, and the transport-site of YiiP receives protons from water molecules that gain access to its hydrophobic environment and transduces the energy of an inward proton gradient to drive Zn(II) efflux1,2. This membrane protein is a well characterized member3-7 of the protein family of cation diffusion facilitators (CDFs) that occurs at all phylogenetic levels8-10. X-ray mediated hydroxyl radical labeling of YiiP and mass spectrometric analysis showed that Zn(II) binding triggered a highly localized, all-or-none change of water accessibility to the transport-site and an adjacent hydrophobic gate. Millisecond time-resolved dynamics revealed a concerted and reciprocal pattern of accessibility changes along a transmembrane helix, suggesting a rigid-body helical reorientation linked to Zn(II) binding that triggers the closing of the hydrophobic gate. The gated water access to the transport-site enables a stationary proton gradient to facilitate the conversion of zinc binding energy to the kinetic power stroke of a vectorial zinc transport. The kinetic details provide energetic insights into a proton-coupled active transport reaction. PMID:25043033

Tautomeric transformation of temozolomide, their proton affinities and chemical reactivities: A theoretical approach.

PubMed

Sang-Aroon, Wichien; Ruangpornvisuti, Vithaya; Amornkitbamrung, Vittaya

2016-05-01

The gas-phase geometry optimizations of bare, mono- and dihydrated complexes of temozolomide isomers were carried out using density functional calculation at the M06-2X/6-31+G(d,p) level of the theory. The structures and protonation energies of protonated species of temozolomide are reported. Chemical indices of all isomers and protonated species are also reported. Energies, thermodynamic quantities, rate constants and equilibrium constants of tautomeric and rotameric transformations of all isomers I1↔TZM↔HIa↔HIb↔I2↔I3 in bare and hydrated systems were obtained. Copyright © 2016 Elsevier Inc. All rights reserved.

Species-dependent chelation of (241)Am by DTPA Di-ethyl ester.

PubMed

Huckle, James E; Sadgrove, Matthew P; Mumper, Russell J; Jay, Michael

2015-04-01

Diethylenetriaminepentaacetic acid (DTPA) is an FDA-approved chelating agent for enhancing the elimination of transuranic elements such as americium from the body. Early access to therapy minimizes deposition of these radionuclides in tissues such as the bone. Due to its poor oral bioavailability, DTPA is administered as an IV injection, delaying access. Therefore, a diethyl-ester analog of DTPA, named C2E2, was synthesized as a means to increase oral absorption. As a hexadentate ligand, it was hypothesized that C2E2 was capable of binding americium directly. Therefore, the protonation constants and americium stability constant for C2E2 were determined by potentiometric titration and a solvent extraction method, respectively. C2E2 was shown to bind americium with a log K of 19.6. The concentrations of C2E2, its metabolite C2E1, and DTPA required to achieve effective binding in rat, beagle, and human plasma were studied in vitro. Dose response curves for each ligand were established, and the 50% maximal effective concentrations were determined for each species. As expected, higher concentrations of C2E2 were required to achieve the same degree of binding as DTPA. The results indicated that chelation in beagle plasma is more representative of the human response than rats. Finally, the pharmacokinetics of C2E2 were investigated in beagles, and the data was fit to a two-compartment model with elimination from the central compartment, along with first-order absorption. Based on the in vitro data, a 100 mg kg dose of C2E2 can be expected to have an effective duration of action of 3.8 h in beagles.

Species-Dependent Chelation of 241Am by DTPA Di-ethyl Ester

PubMed Central

Huckle, James E.; Sadgrove, Matthew P.; Mumper, Russell J.; Jay, Michael

2014-01-01

Diethylenetriaminepentaacetic acid (DTPA) is an FDA approved chelating agent for enhancing the elimination of transuranic elements such as americium from the body. Early access to therapy minimizes deposition of these radionuclides in tissues such as the bone. Due to its poor oral bioavailability, DTPA is administered as an IV injection, delaying access. Therefore a diethyl-ester analog of DTPA, named C2E2, was synthesized as a means to increase oral absorption. As a hexadentate ligand, it was hypothesized that C2E2 was capable of binding americium directly. Therefore, the protonation constants and americium stability constant for C2E2 were determined by potentiometric titration and a solvent extraction method, respectively. C2E2 was shown to bind americium with a log K of 19.6. The concentrations of C2E2, its metabolite C2E1, and DTPA required to achieve effective binding in rat, beagle, and human plasma were studied in vitro. Dose response curves for each ligand were established and the 50% maximal effective concentrations were determined for each species. As expected, higher concentrations of C2E2 were required to achieve the same degree of binding as DTPA. The results indicated that chelation in beagle plasma is more representative of the human response than rats. Finally, the pharmacokinetics of C2E2 were investigated in beagles and the data was fit to a two-compartment model with elimination from the central compartment, along with first-order absorption. Based on the in vitro data, a 100 mg kg−1 dose of C2E2 can be expected to have an effective duration of action of 3.8 hours in beagles. PMID:25706138

External protons destabilize the activated voltage sensor in hERG channels.

PubMed

Shi, Yu Patrick; Cheng, Yen May; Van Slyke, Aaron C; Claydon, Tom W

2014-03-01

Extracellular acidosis shifts hERG channel activation to more depolarized potentials and accelerates channel deactivation; however, the mechanisms underlying these effects are unclear. External divalent cations, e.g., Ca(2+) and Cd(2+), mimic these effects and coordinate within a metal ion binding pocket composed of three acidic residues in hERG: D456 and D460 in S2 and D509 in S3. A common mechanism may underlie divalent cation and proton effects on hERG gating. Using two-electrode voltage clamp, we show proton sensitivity of hERG channel activation (pKa = 5.6), but not deactivation, was greatly reduced in the presence of Cd(2+) (0.1 mM), suggesting a common binding site for the Cd(2+) and proton effect on activation and separable effects of protons on activation and deactivation. Mutational analysis confirmed that D509 plays a critical role in the pH dependence of activation, as shown previously, and that cooperative actions involving D456 and D460 are also required. Importantly, neutralization of all three acidic residues abolished the proton-induced shift of activation, suggesting that the metal ion binding pocket alone accounts for the effects of protons on hERG channel activation. Voltage-clamp fluorimetry measurements demonstrated that protons shifted the voltage dependence of S4 movement to more depolarized potentials. The data indicate a site and mechanism of action for protons on hERG activation gating; protonation of D456, D460 and D509 disrupts interactions between these residues and S4 gating charges to destabilize the activated configuration of S4.

Impact of mutation on proton transfer reactions in ketosteroid isomerase: insights from molecular dynamics simulations.

PubMed

Chakravorty, Dhruva K; Hammes-Schiffer, Sharon

2010-06-02

The two proton transfer reactions catalyzed by ketosteroid isomerase (KSI) involve a dienolate intermediate stabilized by hydrogen bonds with Tyr14 and Asp99. Molecular dynamics simulations based on an empirical valence bond model are used to examine the impact of mutating these residues on the hydrogen-bonding patterns, conformational changes, and van der Waals and electrostatic interactions during the proton transfer reactions. While the rate constants for the two proton transfer steps are similar for wild-type (WT) KSI, the simulations suggest that the rate constant for the first proton transfer step is smaller in the mutants due to the significantly higher free energy of the dienolate intermediate relative to the reactant. The calculated rate constants for the mutants D99L, Y14F, and Y14F/D99L relative to WT KSI are qualitatively consistent with the kinetic experiments indicating a significant reduction in the catalytic rates along the series of mutants. In the simulations, WT KSI retained two hydrogen-bonding interactions between the substrate and the active site, while the mutants typically retained only one hydrogen-bonding interaction. A new hydrogen-bonding interaction between the substrate and Tyr55 was observed in the double mutant, leading to the prediction that mutation of Tyr55 will have a greater impact on the proton transfer rate constants for the double mutant than for WT KSI. The electrostatic stabilization of the dienolate intermediate relative to the reactant was greater for WT KSI than for the mutants, providing a qualitative explanation for the significantly reduced rates of the mutants. The active site exhibited restricted motion during the proton transfer reactions, but small conformational changes occurred to facilitate the proton transfer reactions by strengthening the hydrogen-bonding interactions and by bringing the proton donor and acceptor closer to each other with the proper orientation for proton transfer. Thus, these calculations suggest that KSI forms a preorganized active site but that the structure of this preorganized active site is altered upon mutation. Moreover, small conformational changes due to stochastic thermal motions are required within this preorganized active site to facilitate the proton transfer reactions.

Imino proton exchange rates imply an induced-fit binding mechanism for the VEGF165-targeting aptamer, Macugen

PubMed Central

Lee, Joon-Hwa; Jucker, Fiona; Pardi, Arthur

2008-01-01

The 2′-fluoro/2′-O-methyl modified RNA aptamer Macugen is a potent inhibitor of the angiogenic regulatory protein, VEGF165. Macugen binds with high affinity to the heparin-binding domain (HBD) of VEGF165. Hydrogen exchange rates of the imino protons were measured for free Macugen and Macugen bound to the HBD or full-length VEGF to better understand the mechanism for high affinity binding. The results here show that the internal loop and hairpin loop of Macugen are highly dynamic in the free state and are greatly stabilized and/or protected from solvent upon protein binding. PMID:18485899

Conformational Changes Represent the Rate-Limiting Step in the Transport Cycle of Maize SUCROSE TRANSPORTER1[C][W

PubMed Central

Derrer, Carmen; Wittek, Anke; Bamberg, Ernst; Carpaneto, Armando; Dreyer, Ingo; Geiger, Dietmar

2013-01-01

Proton-driven Suc transporters allow phloem cells of higher plants to accumulate Suc to more than 1 M, which is up to ∼1000-fold higher than in the surrounding extracellular space. The carrier protein can accomplish this task only because proton and Suc transport are tightly coupled. This study provides insights into this coupling by resolving the first step in the transport cycle of the Suc transporter SUT1 from maize (Zea mays). Voltage clamp fluorometry measurements combining electrophysiological techniques with fluorescence-based methods enable the visualization of conformational changes of SUT1 expressed in Xenopus laevis oocytes. Using the Suc derivate sucralose, binding of which hinders conformational changes of SUT1, the association of protons to the carrier could be dissected from transport-associated movements of the protein. These combined approaches enabled us to resolve the binding of protons to the carrier and its interrelationship with the alternating movement of the protein. The data indicate that the rate-limiting step of the reaction cycle is determined by the accessibility of the proton binding site. This, in turn, is determined by the conformational change of the SUT1 protein, alternately exposing the binding pockets to the inward and to the outward face of the membrane. PMID:23964025

Mechanistic characterization of the HDV genomic ribozyme: a mutant of the C41 motif provides insight into the positioning and thermodynamic linkage of metal ions and protons.

PubMed

Nakano, Shu-ichi; Bevilacqua, Philip C

2007-03-20

Binding of two Mg2+ and two H+ ions influences the self-cleavage activity of the genomic HDV ribozyme. The positioning of these four ligands and their thermodynamic linkage are not fully resolved. Protonated C41 engages in a base triple, whereas protonated C75 has been implicated as an acid-base catalyst in bond cleavage. Prior studies led to the identification of one structural inner-sphere ion and one catalytic outer-sphere ion. In the present study, the contributions of the C41 base triple to the metal ion- and pH-dependence of the reaction are examined. Experiments were conducted on a CG to UA double mutant (DM), which changes the base triple to one involving an unprotonated C41. Below pH 6, the DM has a steeper dependence on pH than the wild-type (WT), consistent with a single protonation misfolding the core; this conclusion is also supported by thermal denaturation studies. Between pH 6 and 8, the WT and DM display nearly identical catalytic metal ion and H+ binding profiles. In contrast, over the same pH range, the WT and DM have distinct structural ion binding profiles; for the WT, binding is favored at lower pH, whereas the DM shows no pH dependence. These data localize the structural ion to the vicinity of the C41 motif. An overall model is presented that accommodates binding affinity, coupling, and positioning of the two metal ions and the two protons within the ribozyme. The data suggest that a protonated base triple allows the WT ribozyme to maintain appreciable activity at acidic pH, which could play an important role in the life cycle of the virus.

Characterization of lithium coordination sites with magic-angle spinning NMR

NASA Astrophysics Data System (ADS)

Haimovich, A.; Goldbourt, A.

2015-05-01

Lithium, in the form of lithium carbonate, is one of the most common drugs for bipolar disorder. Lithium is also considered to have an effect on many other cellular processes hence it possesses additional therapeutic as well as side effects. In order to quantitatively characterize the binding mode of lithium, it is required to identify the interacting species and measure their distances from the metal center. Here we use magic-angle spinning (MAS) solid-state NMR to study the binding site of lithium in complex with glycine and water (LiGlyW). Such a compound is a good enzyme mimetic since lithium is four-coordinated to one water molecule and three carboxylic groups. Distance measurements to carbons are performed using a 2D transferred echo double resonance (TEDOR) MAS solid-state NMR experiment, and water binding is probed by heteronuclear high-resolution proton-lithium and proton-carbon correlation (wPMLG-HETCOR) experiments. Both HETCOR experiments separate the main complex from impurities and non-specifically bound lithium species, demonstrating the sensitivity of the method to probe the species in the binding site. Optimizations of the TEDOR pulse scheme in the case of a quadrupolar nucleus with a small quadrupole coupling constant show that it is most efficient when pulses are positioned on the spin-1/2 (carbon-13) nucleus. Since the intensity of the TEDOR signal is not normalized, careful data analysis that considers both intensity and dipolar oscillations has to be performed. Nevertheless we show that accurate distances can be extracted for both carbons of the bound glycine and that these distances are consistent with the X-ray data and with lithium in a tetrahedral environment. The lithium environment in the complex is very similar to the binding site in inositol monophosphatase, an enzyme associated with bipolar disorder and the putative target for lithium therapy. A 2D TEDOR experiment applied to the bacterial SuhB gene product of this enzyme was designed to probe direct correlations between lithium, the enzyme inhibitor, and the closest carboxyl carbons of the binding site. At this point, the chemical shift of the bound carboxyl groups in this 29 kDa enzyme could be determined.

The transition in hemoglobin proton-binding characteristics within the basal actinopterygian fishes.

PubMed

Regan, Matthew Daniel; Brauner, Colin J

2010-04-01

Carbon dioxide (CO(2)) transport in the blood of fishes is aided by the proton-binding properties of hemoglobin (Hb) through either a high-intrinsic buffer value and small oxylabile proton binding (Haldane effect), or a low buffer value and large Haldane effect. Primitive species, such as elasmobranchs and sarcopterygians have been shown to rely on the former, while derived species, such as teleosts rely on the latter. Both strategies are effective in the transport of CO(2) in the blood. However, there is a paucity of information on the nature of the transition between these two strategies that appears to occur within the intermediate group of fishes, the basal actinopterygians. The objective of the present study was to simultaneously assess the intrinsic Hb buffer values and Haldane effects of species within the basal actinopterygian lineage to characterize the transition in Hb-proton-binding strategy seen among the fishes. Expressed in order of most basal to most derived, the species investigated included American paddlefish (Polyodon spathula), white sturgeon (Acipenser transmontanus), spotted gar (Lepisosteus oculatus), alligator gar (Atractosteus spatula), bowfin (Amia calva), and mooneye (Hiodon tergisus). Hemolysates from these species were prepared and Hb titrations (under oxygenated and deoxygenated conditions) were performed in both the presence and absence of saturating levels of organic phosphates (GTP). The findings suggest that the nature of the Hb-proton-binding transition may have been punctuated rather than gradual, with the Hb buffer value decreasing and the Haldane effect increasing significantly in bowfin from fairly steady ancestral levels in the four more basal species. This change is coupled with the initial appearance of the choroid rete, as well as an increase in the magnitude and onset pH of the Root effect in bowfin, suggesting that the change in Hb-proton-binding strategy may be associated with the evolution of enhanced O(2) delivery to the eye and an in vivo operational Root effect.

2JHH-resolved HSQC: Exclusive determination of geminal proton-proton coupling constants

NASA Astrophysics Data System (ADS)

Marcó, Núria; Nolis, Pau; Gil, Roberto R.; Parella, Teodor

2017-09-01

The measurement of two-bond proton-proton coupling constants (2JHH) in prochiral CH2 groups from the F2 dimension of 2D spectra is not easy due to the usual presence of complex multiplet J patterns, line broadening effects and strong coupling artifacts. These drawbacks are particularly pronounced and frequent in AB spin systems, as those normally exhibited by the pair of diastereotopic CH2 protons. Here, a novel 2JHH-resolved HSQC experiment for the exclusive and accurate determination of the magnitude of 2JHH from the doublet displayed along the highly-resolved indirect F1 dimension is described. A pragmatic 2JHH NMR profile affords a fast overview of the full range of existing 2JHH values. In addition, a 2JHH/δ(13C)-scaled version proves to be an efficient solution when severe signal overlapping complicate a rigorous analysis. The performance of the method is compared with other current techniques and illustrated by the determination of challenging residual dipolar 2DHH coupling constants of small molecules dissolved in weakly orienting media.

A novel salt of antidiabetic drug metformin resulting from a proton transfer reaction: Synthesis, characterization, crystal structure and solution studies

NASA Astrophysics Data System (ADS)

Ghasemi, Fatemeh; Ghasemi, Khaled; Rezvani, Ali Reza; Shokrollahi, Ardeshir; Refahi, Masoud; García-Granda, Santiago; Mendoza-Meroño, Rafael

2017-03-01

Reaction between N,N-dimethylebiguanidine, Met = Metformin, and 4-hydroxy-2,6-pyridinedicarboxylic acid, HO-dipicH2, results in the formation of a novel proton transfer compound, [MetH2][HO-dipicH]2·H2O, 1. The characterization was performed using FTIR, UV-Vis, 1H and 13C NMR spectroscopy and X-ray crystallography. The crystal system is triclinic with space group P 1 bar and two molecules per unit cell. The protonation constants of O-dipic and Met, in all of probability protonated forms, and the equilibrium constants for the O-dipic-Met proton transfer system were investigated by the potentiometric pH titration method using the Hyperquad2008 program. The stoichiometry of the proton transfer species in solution were in agreement with the solid state result.

Improved version of a binding energy formula for heavy and superheavy nuclei with Z{>=}90 and N{>=}140

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

Dong Tiekuang; Joint Center for Particle, Nuclear Physics and Cosmology, Nanjing 210008; Ren Zhongzhou

2008-06-15

A local formula of binding energy for heavy and superheavy nuclei has very recently been proposed [Dong and Ren, Phys. Rev. C 72, 064331 (2005)]. In this paper, the limit of the predictive ability of this local formula is investigated. It is found that the neutron-proton correlations should be considered when higher precision is required. On the one hand, we introduce a new term |N-Z-50|/A, and on the other hand we consider the different strengths of proton-proton, neutron-neutron, and neutron-proton pairing correlations. For the first time, the standard deviation {radical}({sigma}{sup 2}) of the binding energies for 117 nuclei with Z{>=}90more » and N{>=}140 is reduced to 0.105 MeV. The {alpha} decay energies Q{sub {alpha}} and half-lives T{sub {alpha}} of nuclei with Z=102-118 are reproduced quite well. The proton drip line of superheavy elements from Md (Z=101) to Ds (Z=110) are predicted.« less

Design of Broad-Spectrum Inhibitors of Influenza A Virus M2 Proton Channels: A Molecular Modeling Approach.

PubMed

Klimochkin, Yuri N; Shiryaev, Vadim A; Petrov, Pavel V; Radchenko, Eugene V; Palyulin, Vladimir A; Zefirov, Nikolay S

2016-01-01

The influenza A virus M2 proton channel plays a critical role in its life cycle. However, known M2 inhibitors have lost their clinical efficacy due to the spread of resistant mutant channels. Thus, the search for broad-spectrum M2 channel inhibitors is of great importance. The goal of the present work was to develop a general approach supporting the design of ligands interacting with multiple labile targets and to propose on its basis the potential broad-spectrum inhibitors of the M2 proton channel. The dynamic dimer-of-dimers structures of the three primary M2 target variants, wild-type, S31N and V27A, were modeled by molecular dynamics and thoroughly analyzed in order to define the inhibitor binding sites. The potential inhibitor structures were identified by molecular docking and their binding was verified by molecular dynamics simulation. The binding sites of the M2 proton channel inhibitors were analyzed, a number of potential broad-spectrum inhibitors were identified and the binding modes and probable mechanisms of action of one promising compound were clarified. Using the molecular dynamics and molecular docking techniques, we have refined the dynamic dimer-ofdimers structures of the WT, S31N and V27A variants of the M2 proton channel of the influenza A virus, analyzed the inhibitor binding sites, identified a number of potential broad-spectrum inhibitor structures targeting them, and clarified the binding modes and probable mechanisms of action of one promising compound. The proposed approach is also suitable for the design of ligands interacting with other multiple labile targets.

Crystallographic Location and Mutational Analysis of Zn and Cd Inhibitory Sites and Role of Lipidic Carboxylates in Rescuing Proton Path Mutants in Cytochrome c Oxidase†

PubMed Central

Qin, Ling; Mills, Denise A.; Hiser, Carrie; Murphree, Anna; Garavito, R. Michael; Ferguson-Miller, Shelagh; Hosler, Jonathan

2008-01-01

Cytochrome c oxidase (CcO) transfers protons from the inner surface of the enzyme to the buried O2 reduction site through two different pathways, termed K and D, and from the outer surface via an undefined route. These proton paths can be inhibited by metals such as zinc or cadmium, but the sites of inhibition have not been established. Anomalous difference Fourier analyses of Rhodobacter sphaeroides CcO crystals, with cadmium added, reveal metal binding sites that include the proposed initial proton donor/acceptor of the K pathway, Glu-101 of subunit II. Mutant forms of CcO that lack Glu-101II (E101A and E101A/H96A) exhibit low activity and eliminate metal binding at this site. Significant activity is restored to E101A and E101A/H96A by adding the lipophilic carboxylic compounds, arachidonic acid and cholic acid, but not by their non-carboxylic analogues. These amphipathic acids likely provide their carboxylic groups as substitute proton donors/acceptors in the absence of Glu-101II, as previously observed for arachidonic acid in mutants that alter Asp-132I of the D pathway. The activity of E101A/H96A is still inhibited by zinc, but this remaining inhibition is nearly eliminated by removal of subunit III, which is known to alter the D pathway. The results identify the Glu-101/His-96 site of subunit II as the site of metal binding that inhibits the uptake of protons into the K pathway and indicate that subunit III contributes to zinc binding and/or inhibition of the D pathway. By removing subunit III from E101A/H96A, thereby eliminating zinc inhibition of the uptake of protons from the inner surface of CcO, we confirm that an external zinc binding site is involved in inhibiting the backflow of protons to the active site. PMID:17477548

Photochemical reaction of 2-(3-benzoylphenyl)propionic acid (ketoprofen) with basic amino acids and dipeptides.

PubMed

Suzuki, Tadashi; Shinoda, Mio; Osanai, Yohei; Isozaki, Tasuku

2013-08-22

Photoreaction of 2-(3-benzoylphenyl)propionic acid (ketoprofen, KP) with basic amino acids (histidine, lysine, and arginine) and dipeptides (carnosine and anserine) including a histidine moiety in phosphate buffer solution (pH 7.4) has been investigated with transient absorption spectroscopy. With UV irradiation KP(-) gave rise to a carbanion through a decarboxylation reaction, and the carbanion easily abstracted a proton from the surrounding molecule to yield a 3-ethylbenzophenone ketyl biradical (EBPH). The dipeptides as well as the basic amino acids were found to accelerate the proton transfer reaction whereas alanine and glycine had no effect on the reaction, revealing that these amino acids having a protonated side chain act as a proton donor. The formation quantum yield of EBPH was estimated to be fairly large by means of an actinometrical method with benzophenone, and the bimolecular reaction rate constant for the proton transfer between the carbanion and the protonated basic amino acids or the protonated dipeptides was successfully determined. It has become apparent that the bimolecular reaction rate constant for the proton transfer depended on the acid dissociation constant for the side chain of the amino acids for the first time. This reaction mechanism was interpreted by difference of the heat of reaction for each basic amino acid based on the thermodynamical consideration. These results strongly suggest that the side chain of the basic amino acid residue in protein should play an important role for photochemistry of KP in vivo.

Influence of the protonation state on the binding mode of methyl orange with cucurbiturils

NASA Astrophysics Data System (ADS)

He, Suhang; Sun, Xuzhuo; Zhang, Haibo

2016-03-01

Binding modes of methyl orange (MO) with cucurbiturils (CBs) have been investigated by Single Crystal X-ray Diffraction and NMR Spectroscopy. Detailed study of intermolecular interactions was supported by the Hirshfeld surface analysis. Protonation state of the anionic part of methyl orange has greatly influenced the binding mode of the complex. Stabilized by hydrogen bonding at the portal, hydrophobic and dispersion interactions in the cavity, the protonated methyl orange was deeply inserted into the cavity. On the contrary, the anionic methyl orange has been pushed towards the outside of the cavity by the electrostatic repulsion between the azo group and the portal oxygen. A ;water bridge; was found in MO@CB8 linking both host and guest via hydrogen bonds.

Investigation of phosphoserine and cytidine 5'-phosphate by heteronuclear two-dimensional spectroscopy: samples with strong proton coupling

NASA Astrophysics Data System (ADS)

Bolton, Philip H.

Heteronuclear two-dimensional magnetic resonance is a novel method for investigating the conformations of cellular phosphates. The two-dimensional proton spectra are detected indirectly via the phosphorus-31 nucleus and thus allow determination of proton chemical shifts and coupling constants in situations in which the normal proton spectrum is obscured. Previous investigations of cellular phosphates with relatively simple spin systems have shown that the two-dimensional proton spectrum can be readily related to the normal proton spectrum by subspectral analysis. The normal proton spectrum can be decomposed into two subspectra, one for each polarization of the phosphorus-31 nucleus. The two-dimensional spectrum arises from the difference between the subspectra, and the normal proton spectrum is the sum. This allows simulation of the two-dimensional spectra and hence determination of the proton chemical shifts and coupling constants. Many cellular phosphates of interest, such as 5'-nucleotides and phosphoserine, contain three protons coupled to the phosphorus which are strongly coupled to one another. These samples are amenable to the two-dimensional method and the straightforward subspectral analysis is preserved when a 90° pulse is applied to the protons in the magnetization transfer step. The two-dimensional proton spectra of the samples investigated here have higher resolution than the normal proton spectra, revealing spectral features not readily apparent in the normal proton spectra.

Constant pH molecular dynamics of proteins in explicit solvent with proton tautomerism.

PubMed

Goh, Garrett B; Hulbert, Benjamin S; Zhou, Huiqing; Brooks, Charles L

2014-07-01

pH is a ubiquitous regulator of biological activity, including protein-folding, protein-protein interactions, and enzymatic activity. Existing constant pH molecular dynamics (CPHMD) models that were developed to address questions related to the pH-dependent properties of proteins are largely based on implicit solvent models. However, implicit solvent models are known to underestimate the desolvation energy of buried charged residues, increasing the error associated with predictions that involve internal ionizable residue that are important in processes like hydrogen transport and electron transfer. Furthermore, discrete water and ions cannot be modeled in implicit solvent, which are important in systems like membrane proteins and ion channels. We report on an explicit solvent constant pH molecular dynamics framework based on multi-site λ-dynamics (CPHMD(MSλD)). In the CPHMD(MSλD) framework, we performed seamless alchemical transitions between protonation and tautomeric states using multi-site λ-dynamics, and designed novel biasing potentials to ensure that the physical end-states are predominantly sampled. We show that explicit solvent CPHMD(MSλD) simulations model realistic pH-dependent properties of proteins such as the Hen-Egg White Lysozyme (HEWL), binding domain of 2-oxoglutarate dehydrogenase (BBL) and N-terminal domain of ribosomal protein L9 (NTL9), and the pKa predictions are in excellent agreement with experimental values, with a RMSE ranging from 0.72 to 0.84 pKa units. With the recent development of the explicit solvent CPHMD(MSλD) framework for nucleic acids, accurate modeling of pH-dependent properties of both major class of biomolecules-proteins and nucleic acids is now possible. © 2013 Wiley Periodicals, Inc.

Biotic ligand modeling approach: Synthesis of the effect of major cations on the toxicity of metals to soil and aquatic organisms.

PubMed

Ardestani, Masoud M; van Straalen, Nico M; van Gestel, Cornelis A M

2015-10-01

The biotic ligand model (BLM) approach is used to assess metal toxicity, taking into account the competition of other cations with the free metal ions for binding to the biotic ligand sites of aquatic and soil organisms. The bioavailable fraction of metals, represented by the free metal ion, is a better measure than the total concentration for assessing their potential risk to the environment. Because BLMs are relating toxicity to the fraction of biotic ligands occupied by the metal, they can be useful for investigating factors affecting metal bioaccumulation and toxicity. In the present review, the effects of major cations on the toxicity of metals to soil and aquatic organisms were comprehensively studied by performing a meta-analysis of BLM literature data. Interactions at the binding sites were shown to be species- and metal-specific. The main factors affecting the relationships between toxicity and conditional binding constants for metal binding at the biotic ligand appeared to be Ca(2+) , Mg(2+) , and protons. Other important characteristics of the exposure medium, such as levels of dissolved organic carbon and concentrations of other cations, should also be considered to obtain a proper assessment of metal toxicity to soil and aquatic organisms. © 2015 SETAC.

Synthetic water soluble di-/tritopic molecular receptors exhibiting Ca2+/Mg2+ exchange.

PubMed

Lavie-Cambot, Aurélie; Tron, Arnaud; Ducrot, Aurélien; Castet, Frédéric; Kauffmann, Brice; Beauté, Louis; Allouchi, Hassan; Pozzo, Jean-Luc; Bonnet, Célia S; McClenaghan, Nathan D

2017-05-23

Structural integration of two synthetic water soluble receptors for Ca 2+ and Mg 2+ , namely 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA) and o-aminophenol-N,N,O-triacetic acid (APTRA), respectively, gave novel di- and tritopic ionophores (1 and 2). As Mg 2+ and Ca 2+ cannot be simultaneously complexed by the receptors, allosteric control of complexation results. Potentiometric measurements established stepwise protonation constants and showed high affinity for Ca 2+ (log K = 6.08 and 8.70 for 1 and 2, respectively) and an excellent selectivity over Mg 2+ (log K = 3.70 and 5.60 for 1 and 2, respectively), which is compatible with magnesium-calcium ion exchange. While ion-exchange of a single Mg 2+ for a single Ca 2+ is possible in both 1 and 2, the simultaneous binding of two Mg 2+ by 2 appears prohibitive for replacement of these two ions by a single Ca 2+ . Ion-binding and exchange was further rationalized by DFT calculations.

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

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.

Determination of the side-reaction coefficient of the trihydroxamate siderophore desferrioxamine B in metal-free seawater

NASA Astrophysics Data System (ADS)

Schijf, J.; Burns, S. M.

2016-02-01

Desferrioxamines are a class of trihydroxamate siderophores, members of which occur in surface seawater at low-picomolar concentrations. The total synthesis of desferrioxamine B (DFOB), achieved in the late 1980s and prompted by its use in the treatment of human iron-overload disorders, has ensured a steady commercial supply enabling extensive laboratory studies of its properties. While highly specific for Fe3+, DFOB binds many di-, tri-, and tetravalent metals with substantial affinity and has consequently been employed as a model for strong organic ligands that ostensibly dominate the speciation of several bio-essential metals in the ocean, yet remain largely unidentified. Such comparisons are only meaningful if we know the side-reaction coefficient of DFOB in seawater, which accounts for its binding with the divalent cations Mg2+ and Ca2+. Although quite weak, this has a potentially important effect on the availability of the free ligand, due to the great abundance of these sea salt constituents. We have performed potentiometric titrations to measure the sequential binding of Mg and Ca to the three hydroxamate groups of DFOB, quantified by stability constants β1, β2, and β3. Values of β1 are reported for the first time, however no evidence was found for binding with the terminal amine of DFOB and the corresponding stability constant β4 was thus omitted from the regression model constructed to fit the titration curves. We also examined Mg and Ca binding to methanesulfonate (MSA), a common DFOB counter-ion, by measuring the stability of their complexes with acetohydroxamate in the presence and absence of MSA. Whereas stabilities of metal-MSA complexes have not been published, their similarity to sulfate complexes suggests that MSA may compete with DFOB for Mg and Ca in the titrations. Our calculated side-reaction coefficient is consistent with a previous estimate, but should properly be expressed in terms of protonated forms of DFOB, resulting in a much lower value.

Intracellular diffusion in the presence of mobile buffers. Application to proton movement in muscle.

PubMed

Irving, M; Maylie, J; Sizto, N L; Chandler, W K

1990-04-01

Junge and McLaughlin (1987) derived an expression for the apparent diffusion constant of protons in the presence of both mobile and immobile buffers. Their derivation applies only to cases in which the values of pH are considerably greater than the largest pK of the individual buffers, a condition that is not expected to hold in skeletal muscle or many other cell types. Here we show that, if the pH gradients are small, the same expression for the apparent diffusion constant of protons can be derived without such constraints on the values of the pK's. The derivation is general and can be used to estimate the apparent diffusion constant of any substance that diffuses in the presence of both mobile and immobile buffers. The apparent diffusion constant of protons is estimated to be 1-2 x 10(-6) cm2/s at 18 degrees C inside intact frog twitch muscle fibers. It may be smaller inside cut fibers, owing to a reduction in the concentration of mobile myoplasmic buffers, so that in this preparation a pH gradient, if established within a sarcomere following action potential stimulation, could last 10 ms or longer after stimulation ceased.

Purification of the lactose:H+ carrier of Escherichia coli and characterization of galactoside binding and transport.

PubMed

Wright, J K; Overath, P

1984-02-01

The lactose carrier, a galactoside:H+ symporter in Escherichia coli, has been purified from cytoplasmic membranes by pre-extraction of the membranes with 5-sulfosalicylate, solubilization in dodecyl-O-beta-D-maltoside, Ecteola-column chromatography, and removal of residual impurities by anti-impurity antibodies. Subsequently, the purified carrier was reincorporated into E. coli phospholipid vesicles. Purification was monitored by tracer N-[3H]ethylmaleimide-labeled carrier and by binding of the substrate p-nitrophenyl-alpha-D-galactopyranoside. All purified carrier molecules were active in substrate binding and the purified protein was at least 95% pure by several criteria. Substrate binding to the purified carrier in detergent micelles and in reconstituted proteoliposomes yielded a stoichiometry close to one molecule substrate bound per polypeptide chain. Large unilamellar proteoliposomes (1-5-micron diameter) were prepared from initially small reconstituted vesicles by freeze-thaw cycles and low-speed centrifugation. These proteoliposomes catalyzed facilitated diffusion and active transport in response to artificially imposed electrochemical proton gradients (delta mu H+) or one of its components (delta psi or delta pH). Comparison of the steady-state level of galactoside accumulation and the nominal value of the driving gradients yielded cotransport stoichiometries up to 0.7 proton/galactoside, suggesting that the carrier protein is the only component required for active galactoside transport. The half-saturation constants for active uptake of lactose (KT = 200 microM) or beta-D-galactosyl-1-thio-beta-D-galactoside (KT = 50-80 microM) by the purified carrier were found to be similar to be similar to those measured in cells or cytoplasmic membrane vesicles. The maximum rate for active transport expressed as a turnover number was similar in proteoliposomes and cytoplasmic membrane vesicles (kcat = 3-4 s-1 for lactose) but considerably smaller than in cells (kcat = 40-60 s-1). Possible reasons for this discrepancy are discussed.

Enzyme-substrate and enzyme-inhibitor complexes of triose phosphate isomerase studied by 31P nuclear magnetic resonance.

PubMed Central

Campbell, I D; Jones, R B; Kiener, P A; Waley, S G

1979-01-01

The complex formed between the enzyme triose phosphate isomerase (EC 5.3.1.1.), from rabbit and chicken muscle, and its substrate dihydroxyacetone phosphate was studied by 31P n.m.r. Two other enzyme-ligant complexes examined were those formed by glycerol 3-phosphate (a substrate analogue) and by 2-phosphoglycollate (potential transition-state analogue). Separate resonances were observed in the 31P n.m.r. spectrum for free and bound 2-phosphoglycollate, and this sets an upper limit to the rate constant for dissociation of the enzyme-inhibitor complex; the linewidth of the resonance assigned to the bound inhibitor provided further kinetic information. The position of this resonance did not vary with pH but remained close to that of the fully ionized form of the free 2-phosphoglycollate. It is the fully ionized form of this ligand that binds to the enzyme. The proton uptake that accompanies binding shows protonation of a group on the enzyme. On the basis of chemical and crystallographic information [Hartman (1971) Biochemistry 10, 146--154; Miller & Waley (1971) Biochem. J. 123, 163--170; De la Mare, Coulson, Knowles, Priddle & Offord )1972) Biochem. J. 129, 321--331; Phillips, Rivers, Sternberg, Thornton & Wilson (1977) Biochem. Soc. Trans. 5, 642--647] this group is believed to be glutamate-165. On the other hand, the position of the resonance of D-glycerol 3 phosphate (sn-glycerol 1-phosphate) in the enzyme-ligand complex changes with pH, and both monoanion and dianon of the ligand bind, although dianion binds better. The substrate, dihydroxyacetone phosphate, behaves essentially like glycerol 3-phosphate. The experiments with dihydroxy-acetone phosphate and triose phosphate isomerase have to be carried out at 1 degree C because at 37 degrees C there is conversion into methyl glyoxal and orthophosphate. The mechanismof the enzymic reaction and the reasons for rate-enhancement are considered, and aspects of the pH-dependence are discussed in an Appendix. PMID:38777

The binding of carbon dioxide by horse haemoglobin

PubMed Central

Kilmartin, J. V.; Rossi-Bernardi, L.

1971-01-01

1. Three modified horse haemoglobins have been prepared: (i) αc2βc2, in which both the α-amino groups of the α- and β-chains have reacted with cyanate, (ii) αc2β2, in which the α-amino groups of the α-chains have reacted with cyanate, and (iii) α2βc2, in which the two α-amino groups of the β-chain have reacted with cyanate. 2. The values of n (the Hill constant) for αc2βc2, α2βc2 and αc2β2 were (respectively) 2.5, 2.0 and 2.6, indicating the presence of co-operative interactions between the haem groups for all derivatives. 3. In the alkaline pH range (about pH8.0) all the derivatives show the same charge as normal haemoglobin whereas in the acid pH range (about pH6.0) αc2βc2 differs by four protonic charges and αc2β2, α2βc2 by two protonic charges from normal haemoglobin, indicating that the expected number of ionizing groups have been removed. 4. αc2β2 and αc2βc2 show a 25% decrease in the alkaline Bohr effect, in contrast with α2βc2, which has the same Bohr effect as normal haemoglobin. 5. The deoxy form of αc2βc2 does not bind more CO2 than the oxy form of αc2βc2, whereas αc2β2 and α2βc2 show intermediate binding. 6. The results reported confirm the hypothesis that, under physiological conditions, haemoglobin binds CO2 through the four terminal α-amino groups and that the two terminal α-amino groups of α-chains are involved in the Bohr effect. ImagesPLATE 1 PMID:5166592

Characterization and copper binding of humic and nonhumic organic matter isolated from the South Platte River: Evidence for the presence of nitrogenous binding site

USGS Publications Warehouse

Croue, J.-P.; Benedetti, M.F.; Violleau, D.; Leenheer, J.A.

2003-01-01

Humic substances typically constitute 40-60% of the dissolved organic matter (DOM) in surface waters. However, little information is available regarding the metal binding properties of the nonhumic hydrophilic portion of the DOM. In this study, humic and nonhumic DOM samples were isolated from the South Platte River (Colorado, DOC = 2.6 mg??L-1, SUVA254 = 2.4 L/mg??m) using a two-column array of XAD-8 and XAD-4 resins. The three major isolated fractions of DOM, which accounted for 57% of the bulk DOM, were characterized using a variety of analytical tools. Proton and copper binding properties were studied for each fraction. The main objective of this work was to compare the structural and chemical characteristics of the isolated fractions and test models describing DOM reactivity toward metal ions. The characterization work showed significant structural differences between the three isolated fractions of DOM. The hydrophobic acid fraction (i.e., humic substances isolated from the XAD-8 resin) gave the largest C/H, C/O, and C/N ratios and aromatic carbon content among the three isolated fractions. The transphilic acid (TPHA) fraction ("transphilic" meaning fraction of intermediate polarity isolated from the XAD-4 resin) was found to incorporate the highest proportion of polysaccharides, whereas the transphilic neutral (TPHN) fraction was almost entirely proteinaceous. The gradual increase of the charge with pH for the three DOM fractions is most likely caused by a large distribution of proton affinity constants for the carboxylic groups, as well as a second type of group more generally considered to be phenolic. In the case of the DOM fraction enriched in proteinaceous material (i.e., TPHN fraction), the results showed that the amino groups are reponsible for the charge reversal. For low copper concentrations, nitrogen-containing functional groups similar to those of amino acids are likely to be involved in complexation, in agreement with previously published data.

Guanidinium Group Remains Protonated in a Strongly Basic Arginine Solution

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

Xu, Bo; Jacobs, Michael I.; Kostko, Oleg

Knowledge of the acid dissociation constant of an amino acid has very important ramifications in the biochemistry of proteins and lipid bilayers in aqueous environments because charge and proton transfer depend on its value. The acid dissociation constant for the guanidinium group in arginine has historically been posited as 12.5, but there is substantial variation in published values over the years. Recent experiments suggest that the dissociation constant for arginine is much higher than 12.5, which explains why the arginine guanidinium group retains its positive charge under all physiological conditions. Here, we use X-ray photoelectron spectroscopy to study unsupported, aqueousmore » arginine nanoparticles. By varying the pH of the constituent solution, we provide evidence that the guanidinium group is protonated even in a very basic solution. By analyzing the energy shifts in the C and N X-ray photoelectron spectra, we establish a molecular level picture of how charge and proton transport in aqueous solutions of arginine occur.« less

Guanidinium Group Remains Protonated in a Strongly Basic Arginine Solution

DOE PAGES

Xu, Bo; Jacobs, Michael I.; Kostko, Oleg; ...

2017-05-16

Knowledge of the acid dissociation constant of an amino acid has very important ramifications in the biochemistry of proteins and lipid bilayers in aqueous environments because charge and proton transfer depend on its value. The acid dissociation constant for the guanidinium group in arginine has historically been posited as 12.5, but there is substantial variation in published values over the years. Recent experiments suggest that the dissociation constant for arginine is much higher than 12.5, which explains why the arginine guanidinium group retains its positive charge under all physiological conditions. Here, we use X-ray photoelectron spectroscopy to study unsupported, aqueousmore » arginine nanoparticles. By varying the pH of the constituent solution, we provide evidence that the guanidinium group is protonated even in a very basic solution. By analyzing the energy shifts in the C and N X-ray photoelectron spectra, we establish a molecular level picture of how charge and proton transport in aqueous solutions of arginine occur.« less

Double binding energy differences: Mean-field or pairing effect?

NASA Astrophysics Data System (ADS)

Qi, Chong

2012-10-01

In this Letter we present a systematic analysis on the average interaction between the last protons and neutrons in atomic nuclei, which can be extracted from the double differences of nuclear binding energies. The empirical average proton-neutron interaction Vpn thus derived from experimental data can be described in a very simple form as the interplay of the nuclear mean field and the pairing interaction. It is found that the smooth behavior as well as the local fluctuations of the Vpn in even-even nuclei with N ≠ Z are dominated by the contribution from the proton-neutron monopole interactions. A strong additional contribution from the isoscalar monopole interaction and isovector proton-neutron pairing interaction is seen in the Vpn for even-even N = Z nuclei and for the adjacent odd-A nuclei with one neutron or proton being subtracted.

pH-selective mutagenesis of protein-protein interfaces: in silico design of therapeutic antibodies with prolonged half-life.

PubMed

Spassov, Velin Z; Yan, Lisa

2013-04-01

Understanding the effects of mutation on pH-dependent protein binding affinity is important in protein design, especially in the area of protein therapeutics. We propose a novel method for fast in silico mutagenesis of protein-protein complexes to calculate the effect of mutation as a function of pH. The free energy differences between the wild type and mutants are evaluated from a molecular mechanics model, combined with calculations of the equilibria of proton binding. The predicted pH-dependent energy profiles demonstrate excellent agreement with experimentally measured pH-dependency of the effect of mutations on the dissociation constants for the complex of turkey ovomucoid third domain (OMTKY3) and proteinase B. The virtual scanning mutagenesis identifies all hotspots responsible for pH-dependent binding of immunoglobulin G (IgG) to neonatal Fc receptor (FcRn) and the results support the current understanding of the salvage mechanism of the antibody by FcRn based on pH-selective binding. The method can be used to select mutations that change the pH-dependent binding profiles of proteins and guide the time consuming and expensive protein engineering experiments. As an application of this method, we propose a computational strategy to search for mutations that can alter the pH-dependent binding behavior of IgG to FcRn with the aim of improving the half-life of therapeutic antibodies in the target organism. Copyright © 2013 Wiley Periodicals, Inc.

Modulation of dimerization by residues distant from the interface in bovine neurophysin-II.

PubMed

Zheng, C; Peyton, D; Breslow, E

1997-09-01

The crystal structure of bovine neurophysin-II in its liganded state (Chen et al. [1991] Proc. Natl. Acad. Sci. USA 88, 4240-4244) indicates that the 1-6 sequence has a disordered conformation, lacks noncovalent contacts to other regions of the protein and is distant from the monomer-monomer interface. Cleavage of the 1-6 sequence by Staphylococcus protease V8 yielded a protein that, for the first time, crystallized in both liganded and unliganded states. Insights into the role of the 1-6 sequence in the unliganded state were obtained by NMR and related biophysical comparisons of the native and des-1-6 proteins. NMR spectra demonstrated that the environment and/or conformation of residues in the 1-6 sequence differed in liganded and unliganded states. Additionally, the unliganded des-1-6 protein exhibited a dimerization constant four to five times that of the native protein, potentially accounting for the observation that its peptide affinity was also increased. NMR studies further indicated that the increased dimerization constant of the des-1-6 protein correlated with the presence in the native protein of two isoenergetic forms of the monomer, in contrast to only a single form in the des-1-6 protein, as evidenced by signals from an internal dimerization-sensitive alpha-proton. Thus, the 1-6 sequence reduces the dimerization constant by stabilization of an alternative monomer conformation. A second product of Staphylococcus protease V8 digestion of the native protein was identified as the des-1-6 protein with an internal clip after binding site residue Glu-47, the clip presumably breaking the short 3,10 helix that most directly connects the interface to the interface to the binding site. This product, although unable to bind peptide, retained the dimerization constant of the des-1-6 protein, suggesting a lack of importance of the helix in dimerization and contrasting with the effects of the 1-6 sequence. A model is proposed in which the 1-6 sequence stabilizes the second conformation of the unliganded monomer via interactions affecting the loop region that separates the two neurophysin domains and which has been shown to influence neurophysin self-association.

p-Dimethylaminobenzamide as an ICT dual fluorescent neutral receptor for anions under proton coupled electron transfer sensing mechanism

NASA Astrophysics Data System (ADS)

Wu, Fang-Ying; Jiang, Yun-Bao

2002-04-01

The intramolecular charge transfer (ICT) dual fluorescence of p-dimethylaminobenzamide (DMABA) in acetonitrile was found to show highly sensitive response to HSO 4- over several other anions such as H 2PO 4-,AcO - and ClO 4-. In the presence of bisulfate anion the dual fluorescence intensity ratio and the total intensity of DMABA decreased while the dual emission band positions remained unchanged. Absorption titration indicated that a 1:1 hydrogen bonding complex was formed between bisulfate anion and DMABA, which gave a binding constant of 2.02×10 4 mol-1 l that is two orders of magnitude higher than those for other anions. The obvious isotopic effect observed in the fluorescence quenching [ K SV( HSO4-)/K SV( DSO4-)=1.63 ] suggests that the hydrogen atom moving is an important reaction coordinate. It was assumed that the dual fluorescence response was due to proton coupled electron transfer mediated by hydrogen bonds within the 1:1 HSO 4--DMABA hydrogen-bonding complex.

[15]aneN4S: synthesis, thermodynamic studies and potential applications in chelation therapy.

PubMed

Torres, Nuno; Gonçalves, Sandrina; Fernandes, Ana S; Machado, J Franco; de Brito, Maria J Villa; Oliveira, Nuno G; Castro, Matilde; Costa, Judite; Cabral, Maria F

2014-01-03

The purpose of this work was to synthesize and characterize the thiatetraaza macrocycle 1-thia-4,7,10,13-tetraazacyclopentadecane ([15]aneN4S). Its acid-base behaviour was studied by potentiometry at 25 °C and ionic strength 0.10 M in KNO3. The protonation sequence of this ligand was investigated by 1H-NMR titration that also allowed the determination of protonation constants in D2O. Binding studies of [15]aneN4S with Mn2+, Fe2+, Co2+, Ni2+, Cu2+, Zn2+, Cd2+, Hg2+ and Pb2+ metal ions were further performed under the same experimental conditions. The results demonstrated that this compound has a higher selectivity and thermodynamic stability for Hg2+ and Cu2+, followed by Ni2+. The UV-visible-near IR spectroscopies and magnetic moment data for the Co(II) and Ni(II) complexes indicated a tetragonal distorted coordination geometry for both metal centres. The value of magnetic moment and the X-band EPR spectra of the Cu(II) complex are consistent with a distorted square pyramidal geometry.

Regular square planer bis-(4,4,4-trifluoro-1-(thiophen-2-yl)butane-1,3-dione)/copper(II) complex: Trans/cis-DFT isomerization, crystal structure, thermal, solvatochromism, hirshfeld surface and DNA-binding analysis

NASA Astrophysics Data System (ADS)

Hema, M. K.; Karthik, C. S.; Warad, Ismail; Lokanath, N. K.; Zarrouk, Abdelkader; Kumara, Karthik; Pampa, K. J.; Mallu, P.

2018-04-01

Trans-[Cu(O∩O)2] complex, O∩O = 4,4,4-trifluoro-1-(thiophen-2-yl)butane-1,3-dione was reported with high potential toward CT-DNA binder. The solved XRD-structure of complex indicated a perfect regular square-planer geometry around the Cu(II) center. The trans/cis-DFT-isomerization calculation supported the XRD seen in reflecting the trans-isomer as the kinetic-favor isomer. The desired complex structure was also characterized by conductivity measurement, CHN-elemental analyses, MS, EDX, SEM, UV-Vis., FT-IR, HAS and TG/DTG. The Solvatochromism behavior of the complex was evaluated using four different polar solvents. MPE and Hirshfeld surface analysis (HSA) come to an agreement that fluoride and thiophene protons atoms are with suitable electro-potential environment to form non-classical H-bonds of type CThsbnd H⋯F. The DNA-binding properties were investigated by viscosity tests and spectrometric titrations, the results revealed the complex as strong calf-thymus DNA binder. High intrinsic-binding constants value ∼1.8 × 105 was collected.

Binding Thermodynamics of Ferredoxin:NADP+ Reductase: Two Different Protein Substrates and One Energetics

PubMed Central

Martínez-Júlvez, Marta; Medina, Milagros; Velázquez-Campoy, Adrián

2009-01-01

Abstract The thermodynamics of the formation of binary and ternary complexes between Anabaena PCC 7119 FNR and its substrates, NADP+ and Fd, or Fld, has been studied by ITC. Despite structural dissimilarities, the main difference between Fd and Fld binding to FNR relates to hydrophobicity, reflected in different binding heat capacity and number of water molecules released from the interface. At pH 8, the formation of the binary complexes is both enthalpically and entropically driven, accompanied by the protonation of at least one ionizable group. His299 FNR has been identified as the main responsible for the proton exchange observed. However, at pH 10, where no protonation occurs and intrinsic binding parameters can be obtained, the formation of the binary complexes is entropically driven, with negligible enthalpic contribution. Absence of the FMN cofactor in Fld does not alter significantly the strength of the interaction, but considerably modifies the enthalpic and entropic contributions, suggesting a different binding mode. Ternary complexes show negative cooperativity (6-fold and 11-fold reduction in binding affinity, respectively), and an increase in the enthalpic contribution (more favorable) and a decrease in the entropic contribution (less favorable), with regard to the binary complexes energetics. PMID:19527656

Fluorescence studies on binding of pyrene and its derivatives to humic acid

NASA Astrophysics Data System (ADS)

Nakashima, K.; Maki, M.; Ishikawa, F.; Yoshikawa, T.; Gong, Y.-K.; Miyajima, T.

2007-07-01

Binding of pyrene (PyH) and its derivatives to humic acid (HA) has been studied by fluorescence spectroscopy. The nature of the interaction between HA and pyrene derivatives are extensively investigated by employing three derivatives ranging from anionic to cationic compounds: 1-pyrenebutylic acid (PyA), 1-pyrenemethanol (PyM), and 1-pyrenebutyltrimethylammonium bromide (PyB). Binding constants between HA and PyX (X = H, A, M, B) are obtained by steady-state fluorescence quenching techniques, and it is found that PyB has a markedly large binding constant among the pyrene family. This is attributed to a strong electrostatic interaction between cationic PyB and anionic HA. The result suggests that an electrostatic interaction plays a dominant role in binding of pyrenes to humic acid. The importance of electrostatic interaction was also confirmed by a salt effect on the binding constant. Influence of collisional quenching on the binding constant, which causes overestimation of the binding constant, was examined by time-resolved fluorescence spectroscopy as well as temperature effect in steady-state fluorescence measurements. It is elucidated that collisional quenching does not much bring overestimation into the binding constants.

Cavitation Damage Experiments for Mercury Spallation Targets At the LANSCE WNR in 2008

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

Riemer, Bernie; Wendel, Mark W; Felde, David K

2010-01-01

Proton beam experiments investigating cavitation damage in short pulse mercury spallation targets were performed at LANSCE WNR in July of 2008. They included two main areas for investigation: damage dependence on mercury velocity using geometry more prototypic to the SNS target than previously employed and damage dependence on incident proton beam flux intensity. The flow dependence experiment employed six test targets with mercury velocity in the channel ranging from 0 to more than 4 m/s. Each was hit with 100 WNR beam pulses with peak proton flux equivalent to that of SNS operating at 2.7 MW. Damage dependence on incidentmore » proton beam flux intensity was also investigated with three intensity levels used on simple rectangular shaped targets without mercury flow. Intensity variation was imposed by focusing the beam differently while maintaining protons per pulse. This kept total energy deposited in each target constant. A fourth test target was hit with various beams: constant protons and varied spot size; constant spot size and varied protons. No damage will be assessed in this case. Instead, acoustic emissions associated with cavitation collapse were measured by laser Doppler vibrometer (LDV) from readings of exterior vessel motions as well as by mercury wetted acoustic transducers. This paper will provide a description of the experiment and present available results. Damage assessment will require several months before surface analysis can be completed and was not available in time for IWSMT-9.« less

Synthesis, characterization, crystal structure and solution studies of a novel proton transfer (charge transfer) complex of 2,2‧-dipyridylamine with 2,6-pyridine dicarboxylic acid

NASA Astrophysics Data System (ADS)

Ghasemi, Khaled; Rezvani, Ali Reza; Shokrollahi, Ardeshir; Zarghampour, Fereshteh; Moghimi, Abolghasem; García-Granda, Santiago; Mendoza-Meroño, Rafael

2015-06-01

Reaction between 2,2‧-dipyridylamine (DPA) and 2,6-pyridine dicarboxylic acid (dipicolinic acid, dipicH2), in water results in the formation of a proton transfer or charge transfer (CT) complex, (DPAH)+(dipicH)-·H2O, 1. The characterization was performed using 1H NMR and FTIR spectroscopy, elemental analysis and X-ray crystallography. The crystal system is triclinic with space group P1. The structural investigations exhibit that the hydrogen bonds and π-π stacking interactions stabilize the crystal structure of proton transfer complex. The protonation constants of 2,6-pyridine dicarboxylic acid, 2,2‧-dipyridylamine and the equilibrium constants for dipic-DPA (1:1) proton transfer system were calculated by potentiometric pH titration method using Hyperquad2008 program. The stoichiometries of the proton transfer species in solution was in agreement with the solid state result.

Manipulation and measurement of pH sensitive metal-ligand binding using electrochemical proton generation and metal detection.

PubMed

Read, Tania L; Joseph, Maxim B; Macpherson, Julie V

2016-01-31

Generator-detector electrodes can be used to both perturb and monitor pH dependant metal-ligand binding equilibria, in situ. In particular, protons generated at the generator locally influence the speciation of metal (Cu(2+)) in the presence of ligand (triethylenetetraamine), with the detector employed to monitor, in real time, free metal (Cu(2+)) concentrations.

Application of differential scanning calorimetry to measure the differential binding of ions, water and protons in the unfolding of DNA molecules.

PubMed

Olsen, Chris M; Shikiya, Ronald; Ganugula, Rajkumar; Reiling-Steffensmeier, Calliste; Khutsishvili, Irine; Johnson, Sarah E; Marky, Luis A

2016-05-01

The overall stability of DNA molecules globally depends on base-pair stacking, base-pairing, polyelectrolyte effect and hydration contributions. In order to understand how they carry out their biological roles, it is essential to have a complete physical description of how the folding of nucleic acids takes place, including their ion and water binding. To investigate the role of ions, water and protons in the stability and melting behavior of DNA structures, we report here an experimental approach i.e., mainly differential scanning calorimetry (DSC), to determine linking numbers: the differential binding of ions (Δnion), water (ΔnW) and protons (ΔnH(+)) in the helix-coil transition of DNA molecules. We use DSC and temperature-dependent UV spectroscopic techniques to measure the differential binding of ions, water, and protons for the unfolding of a variety of DNA molecules: salmon testes DNA (ST-DNA), one dodecamer, one undecamer and one decamer duplexes, nine hairpin loops, and two triplexes. These methods can be applied to any conformational transition of a biomolecule. We determined complete thermodynamic profiles, including all three linking numbers, for the unfolding of each molecule. The favorable folding of a DNA helix results from a favorable enthalpy-unfavorable entropy compensation. DSC thermograms and UV melts as a function of salt, osmolyte and proton concentrations yielded releases of ions and water. Therefore, the favorable folding of each DNA molecule results from the formation of base-pair stacks and uptake of both counterions and water molecules. In addition, the triplex with C(+)GC base triplets yielded an uptake of protons. Furthermore, the folding of a DNA duplex is accompanied by a lower uptake of ions and a similar uptake of four water molecules as the DNA helix gets shorter. In addition, the oligomer duplexes and hairpin thermodynamic data suggest ion and water binding depends on the DNA sequence rather than DNA composition. Copyright © 2015. Published by Elsevier B.V.

DFT Study on Nitrite Reduction Mechanism in Copper-Containing Nitrite Reductase.

PubMed

Lintuluoto, Masami; Lintuluoto, Juha M

2016-01-12

Dissimilatory reduction of nitrite by copper-containing nitrite reductase (CuNiR) is an important step in the geobiochemical nitrogen cycle. The proposed mechanisms for the reduction of nitrite by CuNiRs include intramolecular electron and proton transfers, and these two events are understood to couple. Proton-coupled electron transfer is one of the key processes in enzyme reactions. We investigated the geometric structure of bound nitrite and the mechanism of nitrite reduction on CuNiR using density functional theory calculations. Also, the proton transfer pathway, the key residues, and their roles in the reaction mechanism were clarified in this study. In our results, the reduction of T2 Cu site promotes the proton transfer, and the hydrogen bond network around the binding site has an important role not only to stabilize the nitrite binding but also to promote the proton transfer to nitrite.

A comparative study of nano-SiO2 and nano-TiO2 fillers on proton conductivity and dielectric response of a silicotungstic acid-H3PO4-poly(vinyl alcohol) polymer electrolyte.

PubMed

Gao, Han; Lian, Keryn

2014-01-08

The effects of nano-SiO2 and nano-TiO2 fillers on a thin film silicotungstic acid (SiWA)-H3PO4-poly(vinyl alcohol) (PVA) proton conducting polymer electrolyte were studied and compared with respect to their proton conductivity, environmental stability, and dielectric properties, across a temperature range from 243 to 323 K. Three major effects of these fillers have been identified: (a) barrier effect; (b) intrinsic dielectric constant effect; and (c) water retention effect. Dielectric analyses were used to differentiate these effects on polymer electrolyte-enabled capacitors. Capacitor performance was correlated to electrolyte properties through dielectric constant and dielectric loss spectra. Using a single-ion approach, proton density and proton mobility of each polymer electrolyte were derived as a function of temperature. The results allow us to deconvolute the different contributions to proton conductivity in SiWA-H3PO4-PVA-based electrolytes, especially in terms of the effects of fillers on the dynamic equilibrium of free protons and protonated water in the electrolytes.

Intrinsic Thermodynamics and Structure Correlation of Benzenesulfonamides with a Pyrimidine Moiety Binding to Carbonic Anhydrases I, II, VII, XII, and XIII

PubMed Central

Kišonaitė, Miglė; Zubrienė, Asta; Čapkauskaitė, Edita; Smirnov, Alexey; Smirnovienė, Joana; Kairys, Visvaldas; Michailovienė, Vilma; Manakova, Elena; Gražulis, Saulius; Matulis, Daumantas

2014-01-01

The early stage of drug discovery is often based on selecting the highest affinity lead compound. To this end the structural and energetic characterization of the binding reaction is important. The binding energetics can be resolved into enthalpic and entropic contributions to the binding Gibbs free energy. Most compound binding reactions are coupled to the absorption or release of protons by the protein or the compound. A distinction between the observed and intrinsic parameters of the binding energetics requires the dissection of the protonation/deprotonation processes. Since only the intrinsic parameters can be correlated with molecular structural perturbations associated with complex formation, it is these parameters that are required for rational drug design. Carbonic anhydrase (CA) isoforms are important therapeutic targets to treat a range of disorders including glaucoma, obesity, epilepsy, and cancer. For effective treatment isoform-specific inhibitors are needed. In this work we investigated the binding and protonation energetics of sixteen [(2-pyrimidinylthio)acetyl]benzenesulfonamide CA inhibitors using isothermal titration calorimetry and fluorescent thermal shift assay. The compounds were built by combining four sulfonamide headgroups with four tailgroups yielding 16 compounds. Their intrinsic binding thermodynamics showed the limitations of the functional group energetic additivity approach used in fragment-based drug design, especially at the level of enthalpies and entropies of binding. Combined with high resolution crystal structural data correlations were drawn between the chemical functional groups on selected inhibitors and intrinsic thermodynamic parameters of CA-inhibitor complex formation. PMID:25493428

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.

Emulating proton-induced conformational changes in the vesicular monoamine transporter VMAT2 by mutagenesis.

PubMed

Yaffe, Dana; Vergara-Jaque, Ariela; Forrest, Lucy R; Schuldiner, Shimon

2016-11-22

Neurotransporters located in synaptic vesicles are essential for communication between nerve cells in a process mediated by neurotransmitters. Vesicular monoamine transporter (VMAT), a member of the largest superfamily of transporters, mediates transport of monoamines to synaptic vesicles and storage organelles in a process that involves exchange of two H + per substrate. VMAT transport is inhibited by the competitive inhibitor reserpine, a second-line agent to treat hypertension, and by the noncompetitive inhibitor tetrabenazine, presently in use for symptomatic treatment of hyperkinetic disorders. During the transport cycle, VMAT is expected to occupy at least three different conformations: cytoplasm-facing, occluded, and lumen-facing. The lumen- to cytoplasm-facing transition, facilitated by protonation of at least one of the essential membrane-embedded carboxyls, generates a binding site for reserpine. Here we have identified residues in the cytoplasmic gate and show that mutations that disrupt the interactions in this gate also shift the equilibrium toward the cytoplasm-facing conformation, emulating the effect of protonation. These experiments provide significant insight into the role of proton translocation in the conformational dynamics of a mammalian H + -coupled antiporter, and also identify key aspects of the mode of action and binding of two potent inhibitors of VMAT2: reserpine binds the cytoplasm-facing conformation, and tetrabenazine binds the lumen-facing conformation.

Proton and gallium(III) binding properties of a biologically active salicylidene acylhydrazide.

PubMed

Hakobyan, Shoghik; Boily, Jean-François; Ramstedt, Madeleine

2014-09-01

Bacterial biofilm formation causes a range of problems in our society, especially in health care. Salicylidene acylhydrazides (hydrazones) are promising antivirulence drugs targeting secretion systems used during bacterial infection of host cells. When mixed with the gallium ion they become especially potent as bacterial and biofilm growth-suppressing agents, although the mechanisms through which this occurs are not fully understood. At the base of this uncertainty lies the nature of hydrazone-metal interactions. This study addresses this issue by resolving the equilibrium speciation of hydrazone-gallium aqueous solutions. The protonation constants of the target 2-oxo-2-[N-(2,4,6-trihydroxy-benzylidene)-hydrazino]-acetamide (ME0163) hydrazone species and of its 2,4,6-trihydroxybenzaldehyde and oxamic acid hydrazide building blocks were determined by UV-visible spectrophotometry to achieve this goal. These studies show that the hydrazone is an excessively strong complexing agent for gallium and that its antivirulence properties are predominantly ascribed to monomeric 1:1Ga-ME0163 complexes of various Ga hydrolysis and ME0163 protonation states. The chelation of Ga(III) to the hydrazone also increased the stability of the compounds against acid-induced hydrolysis, making this group of compounds very interesting for biological applications where the Fe-antagonist action of both Ga(III) and the hydrazone can be combined for enhanced biological effect. Copyright © 2014. Published by Elsevier Inc.

Measurement of the inclusive 3-jet production differential cross section in proton-proton collisions at 7 TeV and determination of the strong coupling constant in the TeV range.

PubMed

Khachatryan, V; Sirunyan, A M; Tumasyan, A; Adam, W; Bergauer, T; Dragicevic, M; Erö, J; Fabjan, C; Friedl, M; Frühwirth, R; Ghete, V M; Hartl, C; Hörmann, N; Hrubec, J; Jeitler, M; Kiesenhofer, W; Knünz, V; Krammer, M; Krätschmer, I; Liko, D; Mikulec, I; Rabady, D; Rahbaran, B; Rohringer, H; Schöfbeck, R; Strauss, J; Taurok, A; Treberer-Treberspurg, W; Waltenberger, W; Wulz, C-E; Mossolov, V; Shumeiko, N; Suarez Gonzalez, J; Alderweireldt, S; Bansal, M; Bansal, S; Cornelis, T; De Wolf, E A; Janssen, X; Knutsson, A; Luyckx, S; Ochesanu, S; Rougny, R; Van De Klundert, M; Van Haevermaet, H; Van Mechelen, P; Van Remortel, N; Van Spilbeeck, A; Blekman, F; Blyweert, S; D'Hondt, J; Daci, N; Heracleous, N; Keaveney, J; Lowette, S; Maes, M; Olbrechts, A; Python, Q; Strom, D; Tavernier, S; Van Doninck, W; Van Mulders, P; Van Onsem, G P; Villella, I; Caillol, C; Clerbaux, B; De Lentdecker, G; Dobur, D; Favart, L; Gay, A P R; Grebenyuk, A; Léonard, A; Mohammadi, A; Perniè, L; Reis, T; Seva, T; Thomas, L; 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This paper presents a measurement of the inclusive 3-jet production differential cross section at a proton-proton centre-of-mass energy of 7 TeV using data corresponding to an integrated luminosity of 5[Formula: see text]collected with the CMS detector. The analysis is based on the three jets with the highest transverse momenta. The cross section is measured as a function of the invariant mass of the three jets in a range of 445-3270 GeV and in two bins of the maximum rapidity of the jets up to a value of 2. A comparison between the measurement and the prediction from perturbative QCD at next-to-leading order is performed. Within uncertainties, data and theory are in agreement. The sensitivity of the observable to the strong coupling constant [Formula: see text] is studied. A fit to all data points with 3-jet masses larger than 664 GeV gives a value of the strong coupling constant of [Formula: see text].

Constraints on parton distribution functions and extraction of the strong coupling constant from the inclusive jet cross section in pp collisions at [Formula: see text][Formula: see text].

PubMed

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Russ, J; Vogel, H; Vorobiev, I; Cumalat, J P; Ford, W T; Gaz, A; Krohn, M; Luiggi Lopez, E; Nauenberg, U; Smith, J G; Stenson, K; Ulmer, K A; Wagner, S R; Alexander, J; Chatterjee, A; Chaves, J; Chu, J; Dittmer, S; Eggert, N; Mirman, N; Nicolas Kaufman, G; Patterson, J R; Ryd, A; Salvati, E; Skinnari, L; Sun, W; Teo, W D; Thom, J; Thompson, J; Tucker, J; Weng, Y; Winstrom, L; Wittich, P; Winn, D; Abdullin, S; Albrow, M; Anderson, J; Apollinari, G; Bauerdick, L A T; Beretvas, A; Berryhill, J; Bhat, P C; Bolla, G; Burkett, K; Butler, J N; Cheung, H W K; Chlebana, F; Cihangir, S; Elvira, V D; Fisk, I; Freeman, J; Gao, Y; Gottschalk, E; Gray, L; Green, D; Grünendahl, S; Gutsche, O; Hanlon, J; Hare, D; Harris, R M; Hirschauer, J; Hooberman, B; Jindariani, S; Johnson, M; Joshi, U; Kaadze, K; Klima, B; Kreis, B; Kwan, S; Linacre, J; Lincoln, D; Lipton, R; Liu, T; Lykken, J; Maeshima, K; Marraffino, J M; Martinez Outschoorn, V I; Maruyama, S; Mason, D; McBride, P; Merkel, P; Mishra, K; Mrenna, S; Musienko, Y; Nahn, S; Newman-Holmes, C; O'Dell, V; Prokofyev, O; Sexton-Kennedy, E; Sharma, S; Soha, A; Spalding, W J; Spiegel, L; Taylor, L; Tkaczyk, S; Tran, N V; Uplegger, L; Vaandering, E W; Vidal, R; Whitbeck, A; Whitmore, J; Yang, F; Acosta, D; Avery, P; Bortignon, P; Bourilkov, D; Carver, M; Cheng, T; Curry, D; Das, S; De Gruttola, M; Di Giovanni, G P; Field, R D; Fisher, M; Furic, I K; Hugon, J; Konigsberg, J; Korytov, A; Kypreos, T; Low, J F; Matchev, K; Milenovic, P; Mitselmakher, G; Muniz, L; Rinkevicius, A; Shchutska, L; Snowball, M; Sperka, D; Yelton, J; Zakaria, M; Hewamanage, S; Linn, S; Markowitz, P; Martinez, G; Rodriguez, J L; Adams, T; Askew, A; Bochenek, J; Diamond, B; Haas, J; Hagopian, S; Hagopian, V; Johnson, K F; Prosper, H; Veeraraghavan, V; Weinberg, M; Baarmand, M M; Hohlmann, M; Kalakhety, H; Yumiceva, F; Adams, M R; Apanasevich, L; Bazterra, V E; Berry, D; Betts, R R; Bucinskaite, I; Cavanaugh, R; Evdokimov, O; Gauthier, L; Gerber, C E; Hofman, D J; Khalatyan, S; Kurt, P; Moon, D H; O'Brien, C; Silkworth, C; Turner, P; Varelas, N; Bilki, B; Clarida, W; Dilsiz, K; Duru, F; Haytmyradov, M; Merlo, J-P; Mermerkaya, H; Mestvirishvili, A; Moeller, A; Nachtman, J; Ogul, H; Onel, Y; Ozok, F; Penzo, A; Rahmat, R; Sen, S; Tan, P; Tiras, E; Wetzel, J; Yi, K; Barnett, B A; Blumenfeld, B; Bolognesi, S; Fehling, D; Gritsan, A V; Maksimovic, P; Martin, C; Swartz, M; Baringer, P; Bean, A; Benelli, G; Bruner, C; Kenny, R P; Malek, M; Murray, M; Noonan, D; Sanders, S; Sekaric, J; Stringer, R; Wang, Q; Wood, J S; Chakaberia, I; Ivanov, A; Khalil, S; Makouski, M; Maravin, Y; Saini, L K; Shrestha, S; Skhirtladze, N; Svintradze, I; Gronberg, J; Lange, D; Rebassoo, F; Wright, D; Baden, A; Belloni, A; Calvert, B; Eno, S C; Gomez, J A; Hadley, N J; Kellogg, R G; Kolberg, T; Lu, Y; Marionneau, M; Mignerey, A C; Pedro, K; Skuja, A; Tonjes, M B; Tonwar, S C; Apyan, A; Barbieri, R; Bauer, G; Busza, W; Cali, I A; Chan, M; Di Matteo, L; Gomez Ceballos, G; Goncharov, M; Gulhan, D; Klute, M; Lai, Y S; Lee, Y-J; Levin, A; Luckey, P D; Ma, T; Paus, C; Ralph, D; Roland, C; Roland, G; Stephans, G S F; Stöckli, F; Sumorok, K; Velicanu, D; Veverka, J; Wyslouch, B; Yang, M; Zanetti, M; Zhukova, V; Dahmes, B; Gude, A; Kao, S C; Klapoetke, K; Kubota, Y; Mans, J; Pastika, N; Rusack, R; Singovsky, A; Tambe, N; Turkewitz, J; Acosta, J G; Oliveros, S; Avdeeva, E; Bloom, K; Bose, S; Claes, D R; Dominguez, A; Gonzalez Suarez, R; Keller, J; Knowlton, D; Kravchenko, I; Lazo-Flores, J; Malik, S; Meier, F; Snow, G R; Zvada, M; Dolen, J; Godshalk, A; Iashvili, I; Kharchilava, A; Kumar, A; Rappoccio, S; Alverson, G; Barberis, E; Baumgartel, D; Chasco, M; Haley, J; Massironi, A; Morse, D M; Nash, D; Orimoto, T; Trocino, D; Wang, R J; Wood, D; Zhang, J; Hahn, K A; Kubik, A; Mucia, N; Odell, N; Pollack, B; Pozdnyakov, A; Schmitt, M; Stoynev, S; Sung, K; Velasco, M; Won, S; Brinkerhoff, A; Chan, K M; Drozdetskiy, A; Hildreth, M; Jessop, C; Karmgard, D J; Kellams, N; Lannon, K; Luo, W; Lynch, S; Marinelli, N; Pearson, T; Planer, M; Ruchti, R; Valls, N; Wayne, M; Wolf, M; Woodard, A; Antonelli, L; Brinson, J; Bylsma, B; Durkin, L S; Flowers, S; Hart, A; Hill, C; Hughes, R; Kotov, K; Ling, T Y; Puigh, D; Rodenburg, M; Smith, G; Winer, B L; Wolfe, H; Wulsin, H W; Driga, O; Elmer, P; Hardenbrook, J; Hebda, P; Hunt, A; Koay, S A; Lujan, P; Marlow, D; Medvedeva, T; Mooney, M; Olsen, J; Piroué, P; Quan, X; Saka, H; Stickland, D; Tully, C; Werner, J S; Zuranski, A; Brownson, E; Mendez, H; Ramirez Vargas, J E; Barnes, V E; Benedetti, D; Bortoletto, D; De Mattia, M; Gutay, L; Hu, Z; Jha, M K; Jones, M; Jung, K; Kress, M; Leonardo, N; Lopes Pegna, D; Maroussov, V; Miller, D H; Neumeister, N; Radburn-Smith, B C; Shi, X; Shipsey, I; Silvers, D; Svyatkovskiy, A; Wang, F; Xie, W; Xu, L; Yoo, H D; Zablocki, J; Zheng, Y; Parashar, N; Stupak, J; Adair, A; Akgun, B; Ecklund, K M; Geurts, F J M; Li, W; Michlin, B; Padley, B P; Redjimi, R; Roberts, J; Zabel, J; Betchart, B; Bodek, A; Covarelli, R; de Barbaro, P; Demina, R; Eshaq, Y; Ferbel, T; Garcia-Bellido, A; Goldenzweig, P; Han, J; Harel, A; Khukhunaishvili, A; Petrillo, G; Vishnevskiy, D; Ciesielski, R; Demortier, L; Goulianos, K; Lungu, G; Mesropian, C; Arora, S; Barker, A; Chou, J P; Contreras-Campana, C; Contreras-Campana, E; Duggan, D; Ferencek, D; Gershtein, Y; Gray, R; Halkiadakis, E; Hidas, D; Kaplan, S; Lath, A; Panwalkar, S; Park, M; Patel, R; Salur, S; Schnetzer, S; Somalwar, S; Stone, R; Thomas, S; Thomassen, P; Walker, M; Rose, K; Spanier, S; York, A; Bouhali, O; Castaneda Hernandez, A; Eusebi, R; Flanagan, W; Gilmore, J; Kamon, T; Khotilovich, V; Krutelyov, V; Montalvo, R; Osipenkov, I; Pakhotin, Y; Perloff, A; Roe, J; Rose, A; Safonov, A; Sakuma, T; Suarez, I; Tatarinov, A; Akchurin, N; Cowden, C; Damgov, J; Dragoiu, C; Dudero, P R; Faulkner, J; Kovitanggoon, K; Kunori, S; Lee, S W; Libeiro, T; Volobouev, I; Appelt, E; Delannoy, A G; Greene, S; Gurrola, A; Johns, W; Maguire, C; Mao, Y; Melo, A; Sharma, M; Sheldon, P; Snook, B; Tuo, S; Velkovska, J; Arenton, M W; Boutle, S; Cox, B; Francis, B; Goodell, J; Hirosky, R; Ledovskoy, A; Li, H; Lin, C; Neu, C; Wood, J; Clarke, C; Harr, R; Karchin, P E; Kottachchi Kankanamge Don, C; Lamichhane, P; Sturdy, J; Belknap, D A; Carlsmith, D; Cepeda, M; Dasu, S; Dodd, L; Duric, S; Friis, E; Hall-Wilton, R; Herndon, M; Hervé, A; Klabbers, P; Lanaro, A; Lazaridis, C; Levine, A; Loveless, R; Mohapatra, A; Ojalvo, I; Perry, T; Pierro, G A; Polese, G; Ross, I; Sarangi, T; Savin, A; Smith, W H; Taylor, D; Verwilligen, P; Vuosalo, C; Woods, N

The inclusive jet cross section for proton-proton collisions at a centre-of-mass energy of 7[Formula: see text] was measured by the CMS Collaboration at the LHC with data corresponding to an integrated luminosity of 5.0[Formula: see text]. The measurement covers a phase space up to 2[Formula: see text] in jet transverse momentum and 2.5 in absolute jet rapidity. The statistical precision of these data leads to stringent constraints on the parton distribution functions of the proton. The data provide important input for the gluon density at high fractions of the proton momentum and for the strong coupling constant at large energy scales. Using predictions from perturbative quantum chromodynamics at next-to-leading order, complemented with electroweak corrections, the constraining power of these data is investigated and the strong coupling constant at the Z boson mass [Formula: see text] is determined to be [Formula: see text], which is in agreement with the world average.

SYNCHROTRON RADIO FREQUENCY PHASE CONTROL SYSTEM

DOEpatents

Plotkin, M.; Raka, E.C.; Snyder, H.S.

1963-05-01

A system for canceling varying phase changes introduced by connecting cables and control equipment in an alternating gradient synchrotron is presented. In a specific synchrotron embodiment twelve spaced accelerating stations for the proton bunches are utilized. In order to ensure that the protons receive their boost or kick at the exact instant necessary it is necessary to compensate for phase changes occurring in the r-f circuitry over the wide range of frequencies dictated by the accelerated velocities of the proton bunches. A constant beat frequency is utilized to transfer the r-f control signals through the cables and control equipment to render the phase shift constant and readily compensable. (AEC)

Raman microspectroscopic study of effects of Na(I) and Mg(II) ions on low pH induced DNA structural changes.

PubMed

Muntean, C M; Segers-Nolten, G M J

2003-01-01

In this work a confocal Raman microspectrometer is used to investigate the influence of Na(+) and Mg(2+) ions on the DNA structural changes induced by low pH. Measurements are carried out on calf thymus DNA at neutral pH (7) and pH 3 in the presence of low and high concentrations of Na(+) and Mg(2+) ions, respectively. It is found that low concentrations of Na(+) ions do not protect DNA against binding of H(+). High concentrations of monovalent ions can prevent protonation of the DNA double helix. Our Raman spectra show that low concentrations of Mg(2+) ions partly protect DNA against protonation of cytosine (line at 1262 cm(-1)) but do not protect adenine and guanine N(7) against binding of H(+) (characteristic lines at 1304 and 1488 cm(-1), respectively). High concentrations of Mg(2+) can prevent protonation of cytosine and protonation of adenine (disruption of AT pairs). By analyzing the line at 1488 cm(-1), which obtains most of its intensity from a guanine vibration, high magnesium salt protect the N(7) of guanine against protonation. A high salt concentration can prevent protonation of guanine, cytosine, and adenine in DNA. Higher salt concentrations cause less DNA protonation than lower salt concentrations. Magnesium ions are found to be more effective in protecting DNA against binding of H(+) as compared with calcium ions presented in a previous study. Divalent metal cations (Mg(2+), Ca(2+)) are more effective in protecting DNA against protonation than monovalent ions (Na(+)). Copyright 2003 Wiley Periodicals, Inc. Biopolymers (Biospectroscopy) 72: 000-000, 2003

Dynamic Nuclear Polarization Study of Inhibitor Binding to the M218–60 Proton Transporter from Influenza A

PubMed Central

Andreas, Loren B.; Barnes, Alexander B.; Corzilius, Björn; Chou, James J.; Miller, Eric A.; Caporini, Marc; Rosay, Melanie; Griffin, Robert G

2013-01-01

We demonstrate the use of dynamic nuclear polarization (DNP) to elucidate ligand binding to a membrane protein using dipolar recoupling magic angle spinning (MAS) NMR. In particular, we detect drug binding in the proton transporter M218–60 from influenza A using recoupling experiments at room temperature and with cryogenic DNP. The results indicate that the pore binding site of rimantadine is correlated with previously reported widespread chemical shift changes, suggesting functional binding in the pore. Futhermore, the 15N labeled ammonium of rimantadine was observed near A30 13Cβ and G34 13Cα suggesting a possible hydrogen bond to A30 Carbonyl. Cryogenic DNP was required to observe the weaker external binding site(s) in a ZF-TEDOR spectrum. This approach is generally applicable, particularly for weakly bound ligands, in which case the application of MAS NMR dipolar recoupling requires the low temperatures to quench dynamic exchange processes. For the fully protonated samples investigated, we observed DNP signal enhancements of ~10 at 400 MHz using only 4–6 mM of the polarizing agent TOTAPOL. At 600 MHz and with DNP, we measured a distance between the drug and the protein to a precision of 0.2 Å. PMID:23480101

Substituent Effects on the Coordination Chemistry of Metal-Binding Pharmacophores

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

Craig, Whitney R.; Baker, Tessa W.; Marts, Amy R.

2017-09-12

A combination of XAS, UV–vis, NMR, and EPR was used to examine the binding of a series of α-hydroxythiones to CoCA. All three appear to bind preferentially in their neutral, protonated forms. Two of the three clearly bind in a monodentate fashion, through the thione sulfur alone. Thiomaltol (TM) appears to show some orientational preference, on the basis of the NMR, while it appears that thiopyromeconic acid (TPMA) retains rotational freedom. In contrast, allothiomaltol (ATM), after initially binding in its neutral form, presumably through the thione sulfur, forms a final complex that is five-coordinate via bidentate coordination of ATM. Onmore » the basis of optical titrations, we speculate that this may be due to the lower initial pKa of ATM (8.3) relative to those of TM (9.0) and TPMA (9.5). Binding through the thione is shown to reduce the hydroxyl pKa by ~0.7 pH unit on metal binding, bringing only ATM’s pKa close to the pH of the experiment, facilitating deprotonation and subsequent coordination of the hydroxyl. The data predict the presence of a solvent-exchangeable proton on TM and TPMA, and Q-band 2-pulse ESEEM experiments on CoCA + TM suggest that the proton is present. ESE-detected EPR also showed a surprising frequency dependence, giving only a subset of the expected resonances at X-band.« less

An experimental and modeling study of humic acid concentration effect on H(+) binding: Application of the NICA-Donnan model.

PubMed

Vidali, Roza; Remoundaki, Emmanouela; Tsezos, Marios

2009-11-15

Humic substances are the most abundant components of the colloidal and the dissolved fraction of natural organic matter (NOM) and they are characterized by a strong binding capacity for both metals and organic pollutants, affecting their mobility and bioavailability. The understanding of the humic acidic character is the first necessary step for the study of the mechanisms of binding of other positively charged soluble metal species by humic molecules. The present work, which constitutes part of the Ph.D. thesis of Roza Vidali, reports results on the influence of the concentration of humic acids on the binding of protons obtained through both an experimental and a modeling approach. A reference purified peat humic acid (PPHA) isolated by the International Humic Substances Society (IHSS) and a humic acid from a Greek soil (GHA) were experimentally studied at various humic acid concentrations, ranging from 20 to 200mgL(-1). The proton binding isotherms obtained at different humic acid concentrations have shown that proton binding is dependent on the concentration of both humic acids. Proton binding experimental data were fitted to the NICA-Donnan model and the model parameter values were calculated for humic acid concentrations of 20 and >or=100mgL(-1). The results obtained for the NICA-Donnan parameters at humic acid concentrations >or=100mgL(-1) are in excellent agreement with those reported in the literature. However, these model parameter values cannot be used for modeling and predicting cation binding in natural aquatic systems, where humic acid concentrations are much lower. Two sets of the NICA-Donnan parameters are reported: one for humic acid concentrations of >or=100mgL(-1) and one for humic acid concentration of 20mgL(-1). The significance of the parameters values for each concentration level is also discussed.

Predicted Structures of the Proton-Bound Membrane-Embedded Rotor Rings of the Saccharomyces cerevisiae and Escherichia coli ATP Synthases.

PubMed

Zhou, Wenchang; Leone, Vanessa; Krah, Alexander; Faraldo-Gómez, José D

2017-04-20

Recent years have witnessed a renewed interest in the ATP synthase as a drug target against human pathogens. Indeed, clinical, biochemical, and structural data indicate that hydrophobic inhibitors targeting the membrane-embedded proton-binding sites of the c-subunit ring could serve as last-resort antibiotics against multidrug resistant strains. However, because inhibition of the mitochondrial ATP synthase in humans is lethal, it is essential that these inhibitors be not only potent but also highly selective for the bacterial enzyme. To this end, a detailed understanding of the structure of this protein target is arguably instrumental. Here, we use computational methods to predict the atomic structures of the proton-binding sites in two prototypical c-rings: that of the ATP synthase from Saccharomyces cerevisiae, which is a model system for mitochondrial enzymes, and that from Escherichia coli, which can be pathogenic for humans. Our study reveals the structure of these binding sites loaded with protons and in the context of the membrane, that is, in the state that would mediate the recognition of a potential inhibitor. Both structures reflect a mode of proton coordination unlike those previously observed in other c-ring structures, whether experimental or modeled.

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%.

Mechanism of interaction of the antileukemic drug cytosine arabinoside with aromatic peptides: role of sugar conformation and peptide backbone.

PubMed

Datta, G; Hosur, R V; Verma, N C; Khetrapal, C L; Gurnani, S

1989-01-01

Interaction of the antileukemic drugs, cytosine-arabinoside (Ara-C) and adenosine-arabinoside (Ara-A) and a structural analogue, cytidine, with aromatic dipeptides has been studied by fluorescence and NMR spectroscopy. Ara-C and cytidine bind tryptophanyl and histidyl dipeptides but not tyrosyl dipeptides, while Ara-A does not bind to any of them. Both studies indicate association involving stacking of aromatic moieties. NMR spectra also indicate a protonation of the histidine moiety by Ara-C. In case of cytidine, the chemical shifts observed on binding to His-Phe imply that the backbone protons of the dipeptide participate in the binding. The conformation of the sugar and the base seem to play a very important role in the binding phenomenon as three similar molecules, Ara-C, Ara-A and cytidine bind in totally different ways.

Stability, speciation and spectral properties of NpO2+ complexes with pyridine monocarboxylates in aqueous solution

NASA Astrophysics Data System (ADS)

Dumpala, Rama Mohana Rao; Rawat, Neetika; Tomar, B. S.

2017-06-01

Neptunyl ion as NpO2+ is the least reacting and most mobile radioactive species among all the actinides. The picolinic acid used for decontamination is co-disposed along with the radioactive waste. Thus, in long term storage of HLW, there is high possibility of interaction of actinides and long lived fission products with the picolinate and can cause migration. The complexation of NpO2+ with the three structural isomers of pyridine monocarboxylates provides an insight to explore the role of hetero atom (nitrogen) with respect to key binding moiety (carboxylate). In the present study, the log β values, speciation and spectral properties of NpO2+ complexes with pyridine monocarboxylates viz. picolinate, nicotinate and isonicotinate, have been studied at 298 K in 0.1 M NaClO4 medium using spectrophotometry. The complexation reactions involving protonated ligands are always accompanied by protonation/deprotonation process; thus, the protonation constants of all the three pyridine monocarboxylates under same conditions were also determined by potentiometry. The spectrophotometric data analysis for complexation of NpO2+ with pyridine monocarboxylates indicated the presence of ML and ML2 complexes with log β values of 2.96 ± 0.04, 5.67 ± 0.08 for picolinate, 1.34 ± 0.09, 1.65 ± 0.12 for nicotinate and 1.52 ± 0.04, 2.39 ± 0.06 for isonicotinate. The higher values of log β for picolinate were attributed to chelation while in other two isomers, the binding is through carboxylate group only. Density Functional Theory (DFT) calculations were carried out to get optimized geometries and electrostatic charges on various atoms of the complexes and free pyridine monocarboxylates to support the experimental data. The higher stability of NpO2+ nicotinate and isonicotinate complexes compared to simple carboxylates and the difference in log β between the two is due to the charge polarization from unbound nitrogen to the bound carboxylate oxygen atoms.

2S-4S spectroscopy in hydrogen atom: The new value for the Rydberg constant and the proton charge radius

NASA Astrophysics Data System (ADS)

Kolachevsky, N.; Beyer, A.; Maisenbacher, L.; Matveev, A.; Pohl, R.; Khabarova, K.; Grinin, A.; Lamour, T.; Yost, D. C.; Haensch, T. W.; Udem, Th.

2018-02-01

The core of the "proton radius puzzle" is the discrepancy of four standard deviations between the proton root mean square charge radii (rp) determined from regular hydrogen (H), and the muonic hydrogen atom (μp). We have measured the 2S-4P transition frequency in H, utilizing a cryogenic beam of H and directly demonstrate that quantum interference of neighboring atomic resonances can lead to line shifts much larger than the proton radius discrepancy. Using an asymmetric fit function we obtain rp = 0.8335(95) fm and the Rydberg constant R∞ = 10 973 731.568 076 (96) m-1. The new value for rp is 3.3 combined standard deviations smaller than the latest CODATA value, but in good agreement with the value from μp.

pH-responsive self-assembly by molecular recognition on a macroscopic scale.

PubMed

Zheng, Yongtai; Hashidzume, Akihito; Harada, Akira

2013-07-12

Macroscopic pH-responsive self-assembly is successfully constructed by polyacrylamide(pAAm)-based gels carrying dansyl (Dns) and β-cyclodextrin (βCD) residues, which are represented as Dns-gel and βCD-gel, respectively. Dns-gel and βCD-gel assemble together at pH ≥ 4.0, but disassemble at pH ≤ 3.0. The adhesion strengths for pairs of Dns-gel/βCD-gel increase with increasing pH. The fluorescence study on the model system of pAAm modified with 1 mol% Dns moieties (pAAm/Dns) reveals that Dns residues are protonated at a lower pH, which results in the reduction in binding constant (K) for Dns residues and βCD. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Three new defined proton affinities for polybasic molecules in the gas-phase: Proton microaffinity, proton macroaffinity and proton overallaffinity

NASA Astrophysics Data System (ADS)

Salehzadeh, Sadegh; Bayat, Mehdi

2006-08-01

A theoretical study on complete protonation of a series of tetrabasic molecules with general formula N[(CH 2) nNH 2][(CH 2) mNH 2][(CH 2) pNH 2] (tren, pee, ppe, tpt, epb and ppb) is reported. For first time, three kinds of gas-phase proton affinities for each polybasic molecule are defined as: 'proton microaffinity (PA n, i)', 'proton macroaffinity (PA)' and 'proton overall affinity ( PA)'. The variations of calculated logPA in the series of these molecules is very similar to that of their measured log Kn. There is also a good correlation between the calculated gas-phase proton macroaffinities and proton overallaffinities with corresponding equilibrium macroconstants and overall protonation constants in solution.

Site-specific acid-base properties of pholcodine and related compounds.

PubMed

Kovács, Z; Hosztafi, S; Noszál, B

2006-11-01

The acid-base properties of pholcodine, a cough-depressant agent, and related compounds including metabolites were studied by 1H NMR-pH titrations, and are characterised in terms of macroscopic and microscopic protonation constants. New N-methylated derivatives were also synthesized in order to quantitate site- and nucleus-specific protonation shifts and to unravel microscopic acid-base equilibria. The piperidine nitrogen was found to be 38 and 400 times more basic than its morpholine counterpart in pholcodine and norpholcodine, respectively. The protonation data show that the molecule of pholcodine bears an average of positive charge of 1.07 at physiological pH, preventing it from entering the central nervous system, a plausible reason for its lack of analgesic or addictive properties. The protonation constants of pholcodine and its derivatives are interpreted by comparing with related molecules of pharmaceutical interest. The pH-dependent relative concentrations of the variously protonated forms of pholcodine and morphine are depicted in distribution diagrams.

Protonation-dependent conformational dynamics of the multidrug transporter EmrE

PubMed Central

Dastvan, Reza; Mishra, Smriti; Meiler, Jens; Mchaourab, Hassane S.

2016-01-01

The small multidrug transporter from Escherichia coli, EmrE, couples the energetically uphill extrusion of hydrophobic cations out of the cell to the transport of two protons down their electrochemical gradient. Although principal mechanistic elements of proton/substrate antiport have been described, the structural record is limited to the conformation of the substrate-bound state, which has been shown to undergo isoenergetic alternating access. A central but missing link in the structure/mechanism relationship is a description of the proton-bound state, which is an obligatory intermediate in the transport cycle. Here we report a systematic spin labeling and double electron electron resonance (DEER) study that uncovers the conformational changes of EmrE subsequent to protonation of critical acidic residues in the context of a global description of ligand-induced structural rearrangements. We find that protonation of E14 leads to extensive rotation and tilt of transmembrane helices 1–3 in conjunction with repacking of loops, conformational changes that alter the coordination of the bound substrate and modulate its access to the binding site from the lipid bilayer. The transport model that emerges from our data posits a proton-bound, but occluded, resting state. Substrate binding from the inner leaflet of the bilayer releases the protons and triggers alternating access between inward- and outward-facing conformations of the substrate-loaded transporter, thus enabling antiport without dissipation of the proton gradient. PMID:26787875

The Rydberg constant and proton size from atomic hydrogen

NASA Astrophysics Data System (ADS)

Beyer, Axel; Maisenbacher, Lothar; Matveev, Arthur; Pohl, Randolf; Khabarova, Ksenia; Grinin, Alexey; Lamour, Tobias; Yost, Dylan C.; Hänsch, Theodor W.; Kolachevsky, Nikolai; Udem, Thomas

2017-10-01

At the core of the “proton radius puzzle” is a four-standard deviation discrepancy between the proton root-mean-square charge radii (rp) determined from the regular hydrogen (H) and the muonic hydrogen (µp) atoms. Using a cryogenic beam of H atoms, we measured the 2S-4P transition frequency in H, yielding the values of the Rydberg constant R∞ = 10973731.568076(96) per meterand rp = 0.8335(95) femtometer. Our rp value is 3.3 combined standard deviations smaller than the previous H world data, but in good agreement with the µp value. We motivate an asymmetric fit function, which eliminates line shifts from quantum interference of neighboring atomic resonances.

Verification of the electron/proton coupled mechanism for phenolic H-atom transfer using a triplet π,π ∗ carbonyl

NASA Astrophysics Data System (ADS)

Yamaji, Minoru; Oshima, Juro; Hidaka, Motohiko

2009-06-01

Evidence for the coupled electron/proton transfer mechanism of the phenolic H-atom transfer between triplet π,π ∗ 3,3'-carbonylbis(7-diethylaminocoumarin) and phenol derivatives is obtained by using laser photolysis techniques. It was confirmed that the quenching rate constants of triplet CBC by phenols having positive Hammett constants do not follow the Rehm-Weller equation for electron transfer while those by phenols with negative Hammett constants do it. From the viewpoint of thermodynamic parameters for electron transfer, the crucial factors for phenolic H-atom transfer to π,π ∗ triplet are discussed.

Changes in the zero-point energy of the protons as the source of the binding energy of water to A-phase DNA.

PubMed

Reiter, G F; Senesi, R; Mayers, J

2010-10-01

The measured changes in the zero-point kinetic energy of the protons are entirely responsible for the binding energy of water molecules to A phase DNA at the concentration of 6  water molecules/base pair. The changes in kinetic energy can be expected to be a significant contribution to the energy balance in intracellular biological processes and the properties of nano-confined water. The shape of the momentum distribution in the dehydrated A phase is consistent with coherent delocalization of some of the protons in a double well potential, with a separation of the wells of 0.2 Å.

Rational design of a colorimetric pH sensor from a soluble retinoic acid chaperone.

PubMed

Berbasova, Tetyana; Nosrati, Meisam; Vasileiou, Chrysoula; Wang, Wenjing; Lee, Kin Sing Stephen; Yapici, Ipek; Geiger, James H; Borhan, Babak

2013-10-30

Reengineering of cellular retinoic acid binding protein II (CRABPII) to be capable of binding retinal as a protonated Schiff base is described. Through rational alterations of the binding pocket, electrostatic perturbations of the embedded retinylidene chromophore that favor delocalization of the iminium charge lead to exquisite control in the regulation of chromophoric absorption properties, spanning the visible spectrum (474-640 nm). The pKa of the retinylidene protonated Schiff base was modulated from 2.4 to 8.1, giving rise to a set of proteins of varying colors and pH sensitivities. These proteins were used to demonstrate a concentration-independent, ratiometric pH sensor.

Real-time electron transfer in respiratory complex I

PubMed Central

Verkhovskaya, Marina L.; Belevich, Nikolai; Euro, Liliya; Wikström, Mårten; Verkhovsky, Michael I.

2008-01-01

Electron transfer in complex I from Escherichia coli was investigated by an ultrafast freeze-quench approach. The reaction of complex I with NADH was stopped in the time domain from 90 μs to 8 ms and analyzed by electron paramagnetic resonance (EPR) spectroscopy at low temperatures. The data show that after binding of the first molecule of NADH, two electrons move via the FMN cofactor to the iron–sulfur (Fe/S) centers N1a and N2 with an apparent time constant of ≈90 μs, implying that these two centers should have the highest redox potential in the enzyme. The rate of reduction of center N2 (the last center in the electron transfer sequence) is close to that predicted by electron transfer theory, which argues for the absence of coupled proton transfer or conformational changes during electron transfer from FMN to N2. After fast reduction of N1a and N2, we observe a slow, ≈1-ms component of reduction of other Fe/S clusters. Because all elementary electron transfer rates between clusters are several orders of magnitude higher than this observed rate, we conclude that the millisecond component is limited by a single process corresponding to dissociation of the oxidized NAD+ molecule from its binding site, where it prevents entry of the next NADH molecule. Despite the presence of approximately one ubiquinone per enzyme molecule, no transient semiquinone formation was observed, which has mechanistic implications, suggesting a high thermodynamic barrier for ubiquinone reduction to the semiquinone radical. Possible consequences of these findings for the proton translocation mechanism are discussed. PMID:18316732

Web application for studying the free energy of binding and protonation states of protein-ligand complexes based on HINT

PubMed Central

Bayden, Alexander S.; Fornabaio, Micaela; Scarsdale, J. Neel

2009-01-01

A public web server performing computational titration at the active site in a protein-ligand complex has been implemented. This calculation is based on the Hydropathic INTeraction (HINT) noncovalent force field. From 3D coordinate data for the protein, ligand and bridging waters (if available), the server predicts the best combination of protonation states for each ionizable residue and/or ligand functional group as well as the Gibbs free energy of binding for the ionization-optimized protein-ligand complex. The 3D structure for the modified molecules is available as output. In addition, a graph depicting how this energy changes with acidity, i.e., as a function of added protons, can be obtained. This data may prove to be of use in preparing models for virtual screening and molecular docking. A few illustrative examples are presented. In β secretase (2va7) computational titration flipped the amide groups of Gln12 and Asn37 and protonated a ligand amine yielding an improvement of 6.37 kcal mol−1 in the protein-ligand binding score. Protonation of Glu139 in mutant HIV-1 reverse transcriptase (2opq) allows a water bridge between the protein and inhibitor that increases the protein-ligand interaction score by 0.16 kcal mol−1. In human sialidase NEU2 complexed with an isobutyl ether mimetic inhibitor (2f11) computational titration suggested that protonating Glu218, deprotonating Arg237, flipping the amide bond on Tyr334, and optimizing the positions of several other polar protons would increase the protein-ligand interaction score by 0.71 kcal mol−1. PMID:19554265

Influence of the Selectivity Filter Properties on Proton Selectivity in the Influenza A M2 Channel.

PubMed

Dudev, Todor; Grauffel, Cédric; Lim, Carmay

2016-10-05

The homotetrameric M2 proton channel of influenza A plays a crucial role in the viral life cycle and is thus an important therapeutic target. It selectively conducts protons against a background of other competing cations whose concentrations are up to a million times greater than the proton concentration. Its selectivity is largely determined by a constricted region of its open pore known as the selectivity filter, which is lined by four absolutely conserved histidines. While the mechanism of proton transport through the channel has been studied, the physical principles underlying the selectivity for protons over other cations in the channel's His 4 selectivity filter remain elusive. Furthermore, it is not known if proton selectivity absolutely requires all four histidines with two of the four histidines protonated and if other titratable amino acid residues in lieu of the histidines could bind protons and how they affect proton selectivity. Here, we elucidate how the competition between protons and rival cations such as Na + depends on the selectivity filter's (1) histidine protonation state, (2) solvent exposure, (3) oligomeric state (the number of protein chains and thus the number of His ligands), and (4) ligand composition by evaluating the free energies for replacing monovalent Na + with H 3 O + in various model selectivity filters. We show that tetrameric His 4 filters are more proton-selective than their trimeric His 3 counterparts, and a dicationic His 4 filter where two of the four histidines are protonated is more proton-selective than tetrameric filters with other charge states/composition (different combinations of His protonation states or different metal-ligating ligands). The [His 4 ] 2+ filter achieves proton selectivity by providing suboptimal binding conditions for rival cations such as Na + , which prefers a neutral or negatively charged filter instead of a dicationic one, and three rather than four ligands with oxygen-ligating atoms.

Ti(IV) and the Siderophore Desferrioxamine B: A Tight Complex Has Biological and Environmental Implications.

PubMed

Jones, Kayleigh E; Batchler, Kathleen L; Zalouk, Célia; Valentine, Ann M

2017-02-06

The siderophore desferrioxamine B (DFOB) binds Ti(IV) tightly and precludes its hydrolytic precipitation under biologically and environmentally relevant conditions. This interaction of DFOB with Ti(IV) is investigated by using spectro-potentiometric and spectro-photometric titrations, mass spectrometry, isothermal titration calorimetry (ITC), and computational modeling. The data from pH 2-10 suggest two one-proton equilibria among three species, with one species predominating below pH 3.5, a second from pH 3.5 to 8, and a third above pH 8. The latter species is prone to slow hydrolytic precipitation. Electrospray mass spectrometry allowed the detection of [Ti(IV) (HDFOB)] 2+ and [Ti(DFOB)] + ; these species were assigned as the pH < 3.5 and the 3.5 < pH < 8 species, respectively. The stability constant for Ti(IV)-DFOB was determined by using UV/vis-monitored competition with ethylenediaminetetraacetic acid (EDTA). Taking into consideration the available binding constant of Ti(IV) and EDTA, the data reveal values of log β 111 = 41.7, log β 110 = 38.1, and log β 11-1 = 30.1. The former value was supported by ITC, with the transfer of Ti(IV) from EDTA to DFOB determined to be both enthalpically and entropically favorable. Computational methods yielded a model of Ti-DFOB. The physiological and environmental implications of this tight interaction and the potential role of DFOB in solubilizing Ti(IV) are discussed.

Relative binding affinities of monolignols to horseradish peroxidase

DOE PAGES

Sangha, Amandeep K.; Petridis, Loukas; Cheng, Xiaolin; ...

2016-07-22

Monolignol binding to the peroxidase active site is the first step in lignin polymerization in plant cell walls. Using molecular dynamics, docking, and free energy perturbation calculations, we investigate the binding of monolignols to horseradish peroxidase C. Our results suggest that p-coumaryl alcohol has the strongest binding affinity followed by sinapyl and coniferyl alcohol. Stacking interactions between the monolignol aromatic rings and nearby phenylalanine residues play an important role in determining the calculated relative binding affinities. p-Coumaryl and coniferyl alcohols bind in a pose productive for reaction in which a direct H-bond is formed between the phenolic –OH group andmore » a water molecule (W2) that may facilitate proton transfer during oxidation. In contrast, in the case of sinapyl alcohol there is no such direct interaction, the phenolic –OH group instead interacting with Pro139. Furthermore, since proton and electron transfer is the rate-limiting step in monolignol oxidation by peroxidase, the binding pose (and thus the formation of near attack conformation) appears to play a more important role than the overall binding affinity in determining the oxidation rate.« less

Crystal Structure of Phosphatidylglycerophosphatase (PGPase), a Putative Membrane-Bound Lipid Phosphatase, Reveals a Novel Binuclear Metal Binding Site and Two Proton Wires

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

Kumaran,D.; Bonnano, J.; Burley, S.

2006-01-01

Phosphatidylglycerophosphatase (PGPase), an enzyme involved in lipid metabolism, catalyzes formation of phosphatidylglycerol from phosphatidylglycerophosphate. Phosphatidylglycerol is a multifunctional phospholipid, found in the biological membranes of many organisms. Here, we report the crystal structure of Listeria monocytogenes PGPase at 1.8 Angstroms resolution. PGPase, an all-helical molecule, forms a homotetramer. Each protomer contains an independent active site with two metal ions, Ca{sup 2+} and Mg{sup 2+}, forming a hetero-binuclear center located in a hydrophilic cavity near the surface of the molecule. The binuclear center, conserved ligands, metal-bound water molecules, and an Asp-His dyad form the active site. The catalytic mechanism of thismore » enzyme is likely to proceed via binuclear metal activated nucleophilic water. The binuclear metal-binding active-site environment of this structure should provide insights into substrate binding and metal-dependent catalysis. A long channel with inter-linked linear water chains, termed 'proton wires', is observed at the tetramer interface. Comparison of similar water chain structures in photosynthetic reaction centers (RCs), Cytochrome f, gramicidin, and bacteriorhodopsin, suggests that PGPase may conduct protons via proton wires.« less

Double proton transfer in the complex of acetic acid with methanol: Theory versus experiment

NASA Astrophysics Data System (ADS)

Fernández-Ramos, Antonio; Smedarchina, Zorka; Rodríguez-Otero, Jesús

2001-01-01

To test the approximate instanton approach to intermolecular proton-transfer dynamics, we report multidimensional ab initio bimolecular rate constants of HH, HD, and DD exchange in the complex of acetic acid with methanol in tetrahydrofuran-d8, and compare them with the NMR (nuclear magnetic resonance) experiments of Gerritzen and Limbach. The bimolecular rate constants are evaluated as products of the exchange rates and the equilibrium rate constants of complex formation in solution. The two molecules form hydrogen-bond bridges and the exchange occurs via concerted transfer of two protons. The dynamics of this transfer is evaluated in the complete space of 36 vibrational degrees of freedom. The geometries of the two isolated molecules, the complex, and the transition states corresponding to double proton transfer are fully optimized at QCISD (quadratic configuration interaction including single and double substitutions) level of theory, and the normal-mode frequencies are calculated at MP2 (Møller-Plesset perturbation theory of second order) level with the 6-31G (d,p) basis set. The presence of the solvent is taken into account via single-point calculations over the gas phase geometries with the PCM (polarized continuum model). The proton exchange rate constants, calculated with the instanton method, show the effect of the structure and strength of the hydrogen bonds, reflected in the coupling between the tunneling motion and the other vibrations of the complex. Comparison with experiment, which shows substantial kinetic isotopic effects (KIE), indicates that tunneling prevails over classic exchange for the whole temperature range of observation. The unusual behavior of the experimental KIE upon single and double deuterium substitution is well reproduced and is related to the synchronicity of two-atom tunneling.

Simulation of electron-proton coupling with a Monte Carlo method: application to cytochrome c3 using continuum electrostatics.

PubMed Central

Baptista, A M; Martel, P J; Soares, C M

1999-01-01

A new method is presented for simulating the simultaneous binding equilibrium of electrons and protons on protein molecules, which makes it possible to study the full equilibrium thermodynamics of redox and protonation processes, including electron-proton coupling. The simulations using this method reflect directly the pH and electrostatic potential of the environment, thus providing a much closer and realistic connection with experimental parameters than do usual methods. By ignoring the full binding equilibrium, calculations usually overlook the twofold effect that binding fluctuations have on the behavior of redox proteins: first, they affect the energy of the system by creating partially occupied sites; second, they affect its entropy by introducing an additional empty/occupied site disorder (here named occupational entropy). The proposed method is applied to cytochrome c3 of Desulfovibrio vulgaris Hildenborough to study its redox properties and electron-proton coupling (redox-Bohr effect), using a continuum electrostatic method based on the linear Poisson-Boltzmann equation. Unlike previous studies using other methods, the full reduction order of the four hemes at physiological pH is successfully predicted. The sites more strongly involved in the redox-Bohr effect are identified by analysis of their titration curves/surfaces and the shifts of their midpoint redox potentials and pKa values. Site-site couplings are analyzed using statistical correlations, a method much more realistic than the usual analysis based on direct interactions. The site found to be more strongly involved in the redox-Bohr effect is propionate D of heme I, in agreement with previous studies; other likely candidates are His67, the N-terminus, and propionate D of heme IV. Even though the present study is limited to equilibrium conditions, the possible role of binding fluctuations in the concerted transfer of protons and electrons under nonequilibrium conditions is also discussed. The occupational entropy contributions to midpoint redox potentials and pKa values are computed and shown to be significant. PMID:10354425

Deletion mutation analysis on C-terminal domain of plant vacuolar H(+)-pyrophosphatase.

PubMed

Lin, Hsin Hung; Pan, Yih Jiuan; Hsu, Shen Hsing; Van, Ru Chuan; Hsiao, Yi Yuong; Chen, Jiun Hsien; Pan, Rong Long

2005-10-15

Vacuolar H(+)-translocating inorganic pyrophosphatase (V-PPase; EC 3.6.1.1) is a homodimeric proton-translocase; it contains a single type of polypeptide of approximately 81kDa. A line of evidence demonstrated that the carboxyl terminus of V-PPase is relatively conserved in various plant V-PPases and presumably locates in the vicinity of the catalytic site. In this study, we attempt to identify the roles of the C-terminus of V-PPase by generating a series of C-terminal deletion mutants over-expressed in Saccharomyces cerevisiae, and determining their enzymatic and proton translocating reactions. Our results showed that the deletion mutation at last 5 amino acids in the C-terminus (DeltaC5) induced a dramatic decline in enzymatic activity, proton translocation, and coupling efficiency of V-PPase; but the mutant lacking last 10 amino acids (DeltaC10) retained about 60-70% of the enzymatic activity of wild-type. Truncation of the C-terminus by more than 10 amino acids completely abolished the enzymatic activity and proton translocation of V-PPase. Furthermore, the DeltaC10 mutant displayed a shift in T(1/2) (pretreatment temperature at which half enzymatic activity is observed) but not the optimal pH for PP(i) hydrolytic activity. The deletion of the C-terminus substantially modified apparent K(+) binding constant, but exert no significant changes in the Na(+)-, F(-)-, and Ca(2+)-inhibition of the enzymatic activity of V-PPase. Taken together, we speculate that the C-terminus of V-PPase may play a crucial role in sustaining enzymatic activity and is likely involved in the K(+)-regulation of the enzyme in an indirect manner.

A VARIABLE REACTIVITY MODEL FOR ION BINDING TO ENVIRONMENTAL SORBENTS

EPA Science Inventory

The conceptual and mathematical basis for a new general-composite modeling approach for ion binding to environmental sorbents is presented. The work extends the Simple Metal Sorption (SiMS) model previously presented for metal and proton binding to humic substances. A surface com...

Acid-induced exchange of the imino proton in G.C pairs.

PubMed Central

Nonin, S; Leroy, J L; Gueron, M

1996-01-01

Acid-induced catalysis of imino proton exchange in G.C pairs of DNA duplexes is surprisingly fast, being nearly as fast as for the isolated nucleoside, despite base-pair dissociation constants in the range of 10(-5) at neutral or basic pH. It is also observed in terminal G.C pairs of duplexes and in base pairs of drug-DNA complexes. We have measured imino proton exchange in deoxyguanosine and in the duplex (ATATAGATCTATAT) as a function of pH. We show that acid-induced exchange can be assigned to proton transfer from N7-protonated guanosine to cytidine in the open state of the pair. This is faster than transfer from neutral guanosine (the process of intrinsic catalysis previously characterized at neutral ph) due to the lower imino proton pK of the protonated form, 7.2 instead of 9.4. Other interpretations are excluded by a study of exchange catalysis by formiate and cytidine as exchange catalysts. The cross-over pH between the regimes of pH-independent and acid-induced exchange rates is more basic in the case of base pairs than in the mononucleoside, suggestive of an increase by one to two decades in the dissociation constant of the base pair upon N7 protonation of G. Acid-induced catalysis is much weaker in A.T base pairs, as expected in view of the low pK for protonation of thymidine. PMID:8604298

Acid-induced exchange of the imino proton in G.C pairs.

PubMed

Nonin, S; Leroy, J L; Gueron, M

1996-02-15

Acid-induced catalysis of imino proton exchange in G.C pairs of DNA duplexes is surprisingly fast, being nearly as fast as for the isolated nucleoside, despite base-pair dissociation constants in the range of 10(-5) at neutral or basic pH. It is also observed in terminal G.C pairs of duplexes and in base pairs of drug-DNA complexes. We have measured imino proton exchange in deoxyguanosine and in the duplex (ATATAGATCTATAT) as a function of pH. We show that acid-induced exchange can be assigned to proton transfer from N7-protonated guanosine to cytidine in the open state of the pair. This is faster than transfer from neutral guanosine (the process of intrinsic catalysis previously characterized at neutral ph) due to the lower imino proton pK of the protonated form, 7.2 instead of 9.4. Other interpretations are excluded by a study of exchange catalysis by formiate and cytidine as exchange catalysts. The cross-over pH between the regimes of pH-independent and acid-induced exchange rates is more basic in the case of base pairs than in the mononucleoside, suggestive of an increase by one to two decades in the dissociation constant of the base pair upon N7 protonation of G. Acid-induced catalysis is much weaker in A.T base pairs, as expected in view of the low pK for protonation of thymidine.

Equilibrium constants and protonation site for N-methylbenzenesulfonamides

PubMed Central

Rosa da Costa, Ana M; García-Río, Luis; Pessêgo, Márcia

2011-01-01

Summary The protonation equilibria of four substituted N-methylbenzenesulfonamides, X-MBS: X = 4-MeO (3a), 4-Me (3b), 4-Cl (3c) and 4-NO2 (3d), in aqueous sulfuric acid were studied at 25 °C by UV–vis spectroscopy. As expected, the values for the acidity constants are highly dependent on the electron-donor character of the substituent (the pK BH+ values are −3.5 ± 0.2, −4.2 ± 0.2, −5.2 ± 0.3 and −6.0 ± 0.3 for 3a, 3b, 3c and 3d, respectively). The solvation parameter m* is always higher than 0.5 and points to a decrease in the importance of solvation on the cation stabilization as the electron-donor character of the substituent increases. Hammett plots of the equilibrium constants showed a better correlation with the σ+ substituent parameter than with σ, which indicates that the initial protonation site is the oxygen atom of the sulfonyl group. PMID:22238552

Rate constants measured for hydrated electron reactions with peptides and proteins

NASA Technical Reports Server (NTRS)

Braams, R.

1968-01-01

Effects of ionizing radiation on the amino acids of proteins and the reactivity of the protonated amino group depends upon the pK subscript a of the group. Estimates of the rate constants for reactions involving the amino acid side chains are presented. These rate constants gave an approximate rate constant for three different protein molecules.

Direct Measurement of Equilibrium Constants for High-Affinity Hemoglobins

PubMed Central

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

2003-01-01

The biological functions of heme proteins are linked to their rate and affinity constants for ligand binding. Kinetic experiments are commonly used to measure equilibrium constants for traditional hemoglobins comprised of pentacoordinate ligand binding sites and simple bimolecular reaction schemes. However, kinetic methods do not always yield reliable equilibrium constants with more complex hemoglobins for which reaction mechanisms are not clearly understood. Furthermore, even where reaction mechanisms are clearly understood, it is very difficult to directly measure equilibrium constants for oxygen and carbon monoxide binding to high-affinity (KD ≪ 1 μM) hemoglobins. This work presents a method for direct measurement of equilibrium constants for high-affinity hemoglobins that utilizes a competition for ligands between the "target" protein and an array of "scavenger" hemoglobins with known affinities. This method is described for oxygen and carbon monoxide binding to two hexacoordinate hemoglobins: rice nonsymbiotic hemoglobin and Synechocystis hemoglobin. Our results demonstrate that although these proteins have different mechanisms for ligand binding, their affinities for oxygen and carbon monoxide are similar. Their large affinity constants for oxygen, 285 and ∼100 μM−1 respectively, indicate that they are not capable of facilitating oxygen transport. PMID:12770899

The pressure distribution inside the proton

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

Burkert, V. D.; Elouadrhiri, L.; Girod, F. X.

The proton, one of the components of atomic nuclei, is composed of fundamental particles called quarks and gluons. Gluons are the carriers of the force that binds quarks together, and free quarks are never found in isolation—that is, they are confined within the composite particles in which they reside. The origin of quark confinement is one of the most important questions in modern particle and nuclear physics because confinement is at the core of what makes the proton a stable particle and thus provides stability to the Universe. The internal quark structure of the proton is revealed by deeply virtualmore » Compton scattering1,2, a process in which electrons are scattered off quarks inside the protons, which subsequently emit high-energy photons, which are detected in coincidence with the scattered electrons and recoil protons. Here we report a measurement of the pressure distribution experienced by the quarks in the proton. We find a strong repulsive pressure near the centre of the proton (up to 0.6 femtometres) and a binding pressure at greater distances. The average peak pressure near the centre is about 1035 pascals, which exceeds the pressure estimated for the most densely packed known objects in the Universe, neutron stars3. This work opens up a new area of research on the fundamental gravitational properties of protons, neutrons and nuclei, which can provide access to their physical radii, the internal shear forces acting on the quarks and their pressure distributions.« less

The pressure distribution inside the proton.

PubMed

Burkert, V D; Elouadrhiri, L; Girod, F X

2018-05-01

The proton, one of the components of atomic nuclei, is composed of fundamental particles called quarks and gluons. Gluons are the carriers of the force that binds quarks together, and free quarks are never found in isolation-that is, they are confined within the composite particles in which they reside. The origin of quark confinement is one of the most important questions in modern particle and nuclear physics because confinement is at the core of what makes the proton a stable particle and thus provides stability to the Universe. The internal quark structure of the proton is revealed by deeply virtual Compton scattering 1,2 , a process in which electrons are scattered off quarks inside the protons, which  subsequently emit high-energy photons, which are detected in coincidence with the scattered electrons and recoil protons. Here we report a measurement of the pressure distribution experienced by the quarks in the proton. We find a strong repulsive pressure near the centre of the proton (up to 0.6 femtometres) and a binding pressure at greater distances. The average peak pressure near the centre is about 10 35 pascals, which exceeds the pressure estimated for the most densely packed known objects in the Universe, neutron stars 3 . This work opens up a new area of research on the fundamental gravitational properties of protons, neutrons and nuclei, which can provide access to their physical radii, the internal shear forces acting on the quarks and their pressure distributions.

Constant-pH molecular dynamics using stochastic titration

NASA Astrophysics Data System (ADS)

Baptista, António M.; Teixeira, Vitor H.; Soares, Cláudio M.

2002-09-01

A new method is proposed for performing constant-pH molecular dynamics (MD) simulations, that is, MD simulations where pH is one of the external thermodynamic parameters, like the temperature or the pressure. The protonation state of each titrable site in the solute is allowed to change during a molecular mechanics (MM) MD simulation, the new states being obtained from a combination of continuum electrostatics (CE) calculations and Monte Carlo (MC) simulation of protonation equilibrium. The coupling between the MM/MD and CE/MC algorithms is done in a way that ensures a proper Markov chain, sampling from the intended semigrand canonical distribution. This stochastic titration method is applied to succinic acid, aimed at illustrating the method and examining the choice of its adjustable parameters. The complete titration of succinic acid, using constant-pH MD simulations at different pH values, gives a clear picture of the coupling between the trans/gauche isomerization and the protonation process, making it possible to reconcile some apparently contradictory results of previous studies. The present constant-pH MD method is shown to require a moderate increase of computational cost when compared to the usual MD method.

Time-resolved absorbance changes induced by fast acidification of bacteriorhodopsin in vesicle systems.

PubMed Central

Druckmann, S; Ottolenghi, M; Korenstein, R

1985-01-01

The direction of the accessibility to protons of the binding site in bacteriorhodopsin is of primary importance in elucidating the proton-pump mechanism. The problem is approached via the pH-dependent equilibrium bR560 in equilibrium bR605 in vesicles with preferentially oriented purple membranes. Fast acidification (stopped-flow) experiments with inside-out, monomeric, bR vesicles were carried out with and without a buffer enclosed in the vesicle interior. The results, showing a buffer-induced delay in the formation of bR605, indicate that the binding site is accessible to protons from the inside of the vesicles. We arrive at this conclusion also by working with inside-out trimeric vesicles in the presence and in the absence of H+ (and K+) ionophores. The results suggest that in Halobacterium halobium, the binding site and thus the retinal Schiff base are exposed to the outside of the cell. This conclusion is consistent with a pumping mechanism based on a light-induced pK change. PMID:3978185

Ortho effects in quantitative structure-activity relationships for acetylcholinesterase inhibition by aryl carbamates.

PubMed

Lin, Gialih; Liu, Yu-Chen; Lin, Yan-Fu; Wu, Yon-Gi

2004-10-01

Ortho-substituted phenyl-N-butyl carbamates (1-9) are characterized as "pseudo-pseudo-substrate" inhibitors of acetylcholinesterase. Since the inhibitors protonate at pH 7.0 buffer solution, the virtual inhibition constants (K'is) of the protonated inhibitors are calculated from the equation, - logK'i = - logKi - logKb. The logarithms of the inhibition constant (Ki), the carbamylation constant (k(c)), and the bimolecular inhibition constant (k(i)) for the enzyme inhibitions by carbamates 1-9 are multiply linearly correlated with the Hammett para-substituent constant (sigma(p)), the Taft-Kutter-Hansch ortho steric constant (E(S)), and the Swan-Lupton ortho polar constant (F). Values of rho, delta, and f for the - logKi-, logk(c)-, and logk(i)-correlations are -0.6, -0.16, 0.7; 0.11, 0.03, -0.3; and - 0.5, - 0.12, 0.4, respectively. The Ki step further divides into two steps: 1) the pre-equilibrium protonation of the inhibitors, Kb step and 2) formation of a negatively charged enzyme-inhibitor Michaelis-Menten complex--virtual inhibition, K'i step. The Ki step has little ortho steric enhancement effect; moreover, the k(c)step is insensitive to the ortho steric effect. The f value of 0.7 for the Ki step indicates that ortho electron-withdrawing substituents of the inhibitors accelerate the inhibition reactions from the ortho polar effect; however, the f value of -0.3 for the k(c)step implies that ortho electron-withdrawing substituents of the inhibitors lessen the inhibition reactions from the ortho polar effect.

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.

The second Cu(II)-binding site in a proton-rich environment interferes with the aggregation of amyloid-beta(1-40) into amyloid fibrils.

PubMed

Jun, Sangmi; Gillespie, Joel R; Shin, Byong-kyu; Saxena, Sunil

2009-11-17

The overall morphology and Cu(II) ion coordination for the aggregated amyloid-beta(1-40) [Abeta(1-40)] in N-ethylmorpholine (NEM) buffer are affected by Cu(II) ion concentration. This effect is investigated by transmission electron microscopy (TEM), atomic force microscopy (AFM), and electron spin echo envelope modulation (ESEEM) spectroscopy. At lower than equimolar concentrations of Cu(II) ions, fibrillar aggregates of Abeta(1-40) are observed. At these concentrations of Cu(II), the monomeric and fibrillar Abeta(1-40) ESEEM data indicate that the Cu(II) ion is coordinated by histidine residues. For aggregated Abeta(1-40) at a Cu(II):Abeta molar ratio of 2:1, TEM and AFM images show both linear fibrils and granular amorphous aggregates. The ESEEM spectra show that the multi-histidine coordination for Cu(II) ion partially breaks up and becomes exposed to water or exchangeable protons of the peptide at a higher Cu(II) concentration. Since the continuous-wave electron spin resonance results also suggest two copper-binding sites in Abeta(1-40), the proton ESEEM peak may arise from the second copper-binding site, which may be significantly involved in the formation of granular amorphous aggregates. Thioflavin T fluorescence and circular dichroism experiments also show that Cu(II) inhibits the formation of fibrils and induces a nonfibrillar beta-sheet conformation. Therefore, we propose that Abeta(1-40) has a second copper-binding site in a proton-rich environment and the second binding Cu(II) ion interferes with a conformational transition into amyloid fibrils, inducing the formation of granular amorphous aggregates.

Nuclear magnetic resonance studies of phenylalanine analog interactions with normal and sicklen hemoglobin

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

Lee, Y.H.

Several phenylalanine derivatives have been found to inhibit the gelation of deoxygenated sickle hemoglobin (deoxy HbS). Proton and /sup 19/F-NMR techniques were used to monitor the interaction of selected phenylalanine derivatives with the Hb molecule by using fluorine containing phenylalanine derivatives, Hb labeled at the ..beta..93 position with N-(2,2,2-trifluoroethyl) iodoacetamide (IA-F/sub 3/), and by monitoring the relaxation rates of the C2 and C4 histidine protons. The results show that the /sup 19/F spin-spin relaxation times of L-phenylalanin-4-fluorobenzylamide (PheNBz1-F), which has a deoxy HbS antigelling activity comparable to that of the amino acid, tryptophan, are affected much more strongly by interactionmore » with Hb than are those of glycin-4-fluorobenzylamide (GlyNBz1-F). In contrast, it is shown that N-(2,2,5,5-tetramethylpyrrolidin-1-oxy-3-carboxyl)-L-phenylalanine t-butyl ester (SL-Phe) exhibits specific binding to Hb, and an antigelling activity more than two orders of magnitude greater than that of phenylalanine. These results indicate that the fluorine nuclei strongly influenced by the presence of spin label nitroxide are located in a conformation within a few angstroms of the SL-Phe binding site. Proton NMR relaxation measurements of the C2 and C4 proton resonances from the ..beta..2, 4b143 and ..beta..146 histidine residues show significant and selective effects from the binding of SL-Phe to Hb, indicating that the SL-Phe binding site must be close to the side chains of these three residues. The strong antigelation activity of SL-Phe suggests that this binding site may be one of the intermolecular contact sites of importance to the deoxy HbS aggregation process.« less

Proton and non-proton activation of ASIC channels

PubMed Central

Gautschi, Ivan; van Bemmelen, Miguel Xavier; Schild, Laurent

2017-01-01

The Acid-Sensing Ion Channels (ASIC) exhibit a fast desensitizing current when activated by pH values below 7.0. By contrast, non-proton ligands are able to trigger sustained ASIC currents at physiological pHs. To analyze the functional basis of the ASIC desensitizing and sustained currents, we have used ASIC1a and ASIC2a mutants with a cysteine in the pore vestibule for covalent binding of different sulfhydryl reagents. We found that ASIC1a and ASIC2a exhibit two distinct currents, a proton-induced desensitizing current and a sustained current triggered by sulfhydryl reagents. These currents differ in their pH dependency, their sensitivity to the sulfhydryl reagents, their ionic selectivity and their relative magnitude. We propose a model for ASIC1 and ASIC2 activity where the channels can function in two distinct modes, a desensitizing mode and a sustained mode depending on the activating ligands. The pore vestibule of the channel represents a functional site for binding non-proton ligands to activate ASIC1 and ASIC2 at neutral pH and to prevent channel desensitization. PMID:28384246

Binding of pyrimidin-2-one ribonucleoside by cytidine deaminase as the transition-state analogue 3,4-dihydrouridine and the contribution of the 4-hydroxyl group to its binding affinity.

PubMed

Frick, L; Yang, C; Marquez, V E; Wolfenden, R

1989-11-28

Cytidine deaminase, purified to homogeneity from constitutive mutants of Escherichia coli, was found to bind the competitive inhibitors pyrimidin-2-one ribonucleoside (apparent Ki = 3.6 x 10(-7) M) and 5-fluoropyrimidin-2-one ribonucleoside (apparent Ki = 3.5 x 10(-8) M). Enzyme binding resulted in a change of the lambda max of pyrimidin-2-one ribonucleoside from 303 nm for the free species to 239 nm for the bound species. The value for the bound species was identical with that of an oxygen adduct formed by combination of hydroxide ion with 1,3-dimethyl-2-oxopyrimidinium (239 nm), but lower than that of a sulfur adduct formed by combination of the thiolate anion of N-acetylcysteamine with 1,3-dimethyl-2-oxopyrimidinium (259 nm). The results suggest that pyrimidin-2-one ribonucleoside is bound by cytidine deaminase as an oxygen adduct, probably the covalent hydrate 3,4-dihydrouridine, rather than intact or as an adduct involving a thiol group of the enzyme. In dilute solution at 25 degrees C, the equilibrium constant for formation of a single diastereomer of 3,4-dihydrouridine from pyrimidin-2-one ribonucleoside was estimated as approximately 4.7 x 10(-6), from equilibria of dissociation of water, protonation of 1-methylpyrimidin-2-one, and combination of the 1,3-dimethylpyrimidinium cation with the hydroxide ion.(ABSTRACT TRUNCATED AT 250 WORDS)

The Binding Constant of Estradiol to Bovine Serum Albumin: An Upper-Level Experiment Utilizing Tritium-Labeled Estradiol and Liquid Scintillation Counting

ERIC Educational Resources Information Center

Peihong Liang; Adhyaru, Bhavin; Pearson, Wright L.; Williams, Kathryn R.

2006-01-01

The experiment used [to the third power]H-labeled estradiol to determine the binding constant of estradiol to bovine serum albumin. Estradiol must complex with serum proteins for the transport in the blood stream because of its low solubility in aqueous systems and estradiol-protein binding constant, where K[subscript B] is important to understand…

Elucidating the mechanism for the reduction of nitrite by copper nitrite reductase--a contribution from quantum chemical studies.

PubMed

De Marothy, S A; Blomberg, M R A; Siegbahn, P E M

2007-01-30

Density functional methods have been applied to investigate the properties of the active site of copper-containing nitrite reductases and possible reaction mechanisms for the enzyme catalysis. The results for a model of the active site indicate that a hydroxyl intermediate is not formed during the catalytic cycle, but rather a state with a protonated nitrite bound to the reduced copper. Electron affinity calculations indicate that reduction of the T2 copper site does not occur immediately after nitrite binding. Proton affinity calculations are indicative of substantial pK(a) differences between different states of the T2 site. The calculations further suggest that the reaction does not proceed until uptake of a second proton from the bulk solution. They also indicate that Asp-92 may play both a key role as a proton donor to the substrate, and a structural role in promoting catalysis. In the D92N mutant another base, presumably a nearby histidine (His-249) may take the role as the proton donor. On the basis of these model calculations and available experimental evidence, an ordered reaction mechanism for the reduction of nitrite is suggested. An investigation of the binding modes of the nitric oxide product and the nitrite substrate to the model site has also been made, indicating that nitric oxide prefers to bind in an end-on fashion to the reduced T2 site.

Detailed Analysis of the Binding Mode of Vanilloids to Transient Receptor Potential Vanilloid Type I (TRPV1) by a Mutational and Computational Study

PubMed Central

Mori, Yoshikazu; Ogawa, Kazuo; Warabi, Eiji; Yamamoto, Masahiro; Hirokawa, Takatsugu

2016-01-01

Transient receptor potential vanilloid type 1 (TRPV1) is a non-selective cation channel and a multimodal sensor protein. Since the precise structure of TRPV1 was obtained by electron cryo-microscopy, the binding mode of representative agonists such as capsaicin and resiniferatoxin (RTX) has been extensively characterized; however, detailed information on the binding mode of other vanilloids remains lacking. In this study, mutational analysis of human TRPV1 was performed, and four agonists (capsaicin, RTX, [6]-shogaol and [6]-gingerol) were used to identify amino acid residues involved in ligand binding and/or modulation of proton sensitivity. The detailed binding mode of each ligand was then simulated by computational analysis. As a result, three amino acids (L518, F591 and L670) were newly identified as being involved in ligand binding and/or modulation of proton sensitivity. In addition, in silico docking simulation and a subsequent mutational study suggested that [6]-gingerol might bind to and activate TRPV1 in a unique manner. These results provide novel insights into the binding mode of various vanilloids to the channel and will be helpful in developing a TRPV1 modulator. PMID:27606946

Moving protons with pendant amines: proton mobility in a nickel catalyst for oxidation of hydrogen.

PubMed

O'Hagan, Molly; Shaw, Wendy J; Raugei, Simone; Chen, Shentan; Yang, Jenny Y; Kilgore, Uriah J; DuBois, Daniel L; Bullock, R Morris

2011-09-14

Proton transport is ubiquitous in chemical and biological processes, including the reduction of dioxygen to water, the reduction of CO(2) to formate, and the production/oxidation of hydrogen. In this work we describe intramolecular proton transfer between Ni and positioned pendant amines for the hydrogen oxidation electrocatalyst [Ni(P(Cy)(2)N(Bn)(2)H)(2)](2+) (P(Cy)(2)N(Bn)(2) = 1,5-dibenzyl-3,7-dicyclohexyl-1,5-diaza-3,7-diphosphacyclooctane). Rate constants are determined by variable-temperature one-dimensional NMR techniques and two-dimensional EXSY experiments. Computational studies provide insight into the details of the proton movement and energetics of these complexes. Intramolecular proton exchange processes are observed for two of the three experimentally observable isomers of the doubly protonated Ni(0) complex, [Ni(P(Cy)(2)N(Bn)(2)H)(2)](2+), which have N-H bonds but no Ni-H bonds. For these two isomers, with pendant amines positioned endo to the Ni, the rate constants for proton exchange range from 10(4) to 10(5) s(-1) at 25 °C, depending on isomer and solvent. No exchange is observed for protons on pendant amines positioned exo to the Ni. Analysis of the exchange as a function of temperature provides a barrier for proton exchange of ΔG(‡) = 11-12 kcal/mol for both isomers, with little dependence on solvent. Density functional theory calculations and molecular dynamics simulations support the experimental observations, suggesting metal-mediated intramolecular proton transfers between nitrogen atoms, with chair-to-boat isomerizations as the rate-limiting steps. Because of the fast rate of proton movement, this catalyst may be considered a metal center surrounded by a cloud of exchanging protons. The high intramolecular proton mobility provides information directly pertinent to the ability of pendant amines to accelerate proton transfers during catalysis of hydrogen oxidation. These results may also have broader implications for proton movement in homogeneous catalysts and enzymes in general, with specific implications for the proton channel in the Ni-Fe hydrogenase enzyme.

Calculations of proton-binding thermodynamics in proteins.

PubMed

Beroza, P; Case, D A

1998-01-01

Computational models of proton binding can range from the chemically complex and statistically simple (as in the quantum calculations) to the chemically simple and statistically complex. Much progress has been made in the multiple-site titration problem. Calculations have improved with the inclusion of more flexibility in regard to both the geometry of the proton binding and the larger scale protein motions associated with titration. This article concentrated on the principles of current calculations, but did not attempt to survey their quantitative performance. This is (1) because such comparisons are given in the cited papers and (2) because continued developments in understanding conformational flexibility and interaction energies will be needed to develop robust methods with strong predictive power. Nevertheless, the advances achieved over the past few years should not be underestimated: serious calculations of protonation behavior and its coupling to conformational change can now be confidently pursued against a backdrop of increasing understanding of the strengths and limitations of such models. It is hoped that such theoretical advances will also spur renewed experimental interest in measuring both overall titration curves and individual pKa values or pKa shifts. Exploration of the shapes of individual titration curves (as measured by Hill coefficients and other parameters) would also be useful in assessing the accuracy of computations and in drawing connections to functional behavior.

Influence of Proton Acceptors on the Proton-Coupled Electron Transfer Reaction Kinetics of a Ruthenium-Tyrosine Complex.

PubMed

Lennox, J Christian; Dempsey, Jillian L

2017-11-22

A polypyridyl ruthenium complex with fluorinated bipyridine ligands and a covalently bound tyrosine moiety was synthesized, and its photo-induced proton-coupled electron transfer (PCET) reactivity in acetonitrile was investigated with transient absorption spectroscopy. Using flash-quench methodology with methyl viologen as an oxidative quencher, a Ru 3+ species is generated that is capable of initiating the intramolecular PCET oxidation of the tyrosine moiety. Using a series of substituted pyridine bases, the reaction kinetics were found to vary as a function of proton acceptor concentration and identity, with no significant H/D kinetic isotope effect. Through analysis of the kinetics traces and comparison to a control complex without the tyrosine moiety, PCET reactivity was found to proceed through an equilibrium electron transfer followed by proton transfer (ET-PT) pathway in which irreversible deprotonation of the tyrosine radical cation shifts the ET equilibrium, conferring a base dependence on the reaction. Comprehensive kinetics modeling allowed for deconvolution of complex kinetics and determination of rate constants for each elementary step. Across the five pyridine bases explored, spanning a range of 4.2 pK a units, a linear free-energy relationship was found for the proton transfer rate constant with a slope of 0.32. These findings highlight the influence that proton transfer driving force exerts on PCET reaction kinetics.

Precise Measurement of Long-Range Heteronuclear Coupling Constants by a Novel Broadband Proton–Proton-Decoupled CPMG-HSQMBC Method

PubMed Central

Timári, István; Illyés, Tünde Z; Adams, Ralph W; Nilsson, Mathias; Szilágyi, László; Morris, Gareth A; Kövér, Katalin E

2015-01-01

A broadband proton–proton-decoupled CPMG-HSQMBC method for the precise and direct measurement of long-range heteronuclear coupling constants is presented. The Zangger–Sterk-based homodecoupling scheme reported herein efficiently removes unwanted proton–proton splittings from the heteronuclear multiplets, so that the desired heteronuclear couplings can be determined simply by measuring frequency differences between singlet maxima in the resulting spectra. The proposed pseudo-1D/2D pulse sequences were tested on nucleotides, a metal complex incorporating P heterocycles, and diglycosyl (di)selenides, as well as on other carbohydrate derivatives, for the extraction of nJ(1H,31P), nJ(1H,77Se), and nJ(1H,13C) values, respectively. PMID:25573660

Halorhodopsin pumps Cl– and bacteriorhodopsin pumps protons by a common mechanism that uses conserved electrostatic interactions

PubMed Central

Gunner, M. R.

2014-01-01

Key mutations differentiate the functions of homologous proteins. One example compares the inward ion pump halorhodopsin (HR) and the outward proton pump bacteriorhodopsin (BR). Of the nine essential buried ionizable residues in BR, six are conserved in HR. However, HR changes three BR acids, D85 in a central cluster of ionizable residues, D96, nearer the intracellular, and E204, nearer the extracellular side of the membrane to the small, neutral amino acids T111, V122, and T230, respectively. In BR, acidic amino acids are stationary anions whose proton affinity is modulated by conformational changes, establishing a sequence of directed binding and release of protons. Multiconformation continuum electrostatics calculations of chloride affinity and residue protonation show that, in reaction intermediates where an acid is ionized in BR, a Cl– is bound to HR in a position near the deleted acid. In the HR ground state, Cl– binds tightly to the central cluster T111 site and weakly to the extracellular T230 site, recovering the charges on ionized BR-D85 and neutral E204 in BR. Imposing key conformational changes from the BR M intermediate into the HR structure results in the loss of Cl– from the central T111 site and the tight binding of Cl– to the extracellular T230 site, mirroring the changes that protonate BR-D85 and ionize E204 in BR. The use of a mobile chloride in place of D85 and E204 makes HR more susceptible to the environmental pH and salt concentrations than BR. These studies shed light on how ion transfer mechanisms are controlled through the interplay of protein and ion electrostatics. PMID:25362051

Characterization of Receptor Binding Profiles of Influenza A Viruses Using An Ellipsometry-Based Label-Free Glycan Microarray Assay Platform

PubMed Central

Fei, Yiyan; Sun, Yung-Shin; Li, Yanhong; Yu, Hai; Lau, Kam; Landry, James P.; Luo, Zeng; Baumgarth, Nicole; Chen, Xi; Zhu, Xiangdong

2015-01-01

A key step leading to influenza viral infection is the highly specific binding of a viral spike protein, hemagglutinin (HA), with an extracellular glycan receptor of a host cell. Detailed and timely characterization of virus-receptor binding profiles may be used to evaluate and track the pandemic potential of an influenza virus strain. We demonstrate a label-free glycan microarray assay platform for acquiring influenza virus binding profiles against a wide variety of glycan receptors. By immobilizing biotinylated receptors on a streptavidin-functionalized solid surface, we measured binding curves of five influenza A virus strains with 24 glycans of diverse structures and used the apparent equilibrium dissociation constants (avidity constants, 10–100 pM) as characterizing parameters of viral receptor profiles. Furthermore by measuring binding kinetic constants of solution-phase glycans to immobilized viruses, we confirmed that the glycan-HA affinity constant is in the range of 10 mM and the reaction is enthalpy-driven. PMID:26193329

Characterization of Receptor Binding Profiles of Influenza A Viruses Using An Ellipsometry-Based Label-Free Glycan Microarray Assay Platform.

PubMed

Fei, Yiyan; Sun, Yung-Shin; Li, Yanhong; Yu, Hai; Lau, Kam; Landry, James P; Luo, Zeng; Baumgarth, Nicole; Chen, Xi; Zhu, Xiangdong

2015-07-16

A key step leading to influenza viral infection is the highly specific binding of a viral spike protein, hemagglutinin (HA), with an extracellular glycan receptor of a host cell. Detailed and timely characterization of virus-receptor binding profiles may be used to evaluate and track the pandemic potential of an influenza virus strain. We demonstrate a label-free glycan microarray assay platform for acquiring influenza virus binding profiles against a wide variety of glycan receptors. By immobilizing biotinylated receptors on a streptavidin-functionalized solid surface, we measured binding curves of five influenza A virus strains with 24 glycans of diverse structures and used the apparent equilibrium dissociation constants (avidity constants, 10-100 pM) as characterizing parameters of viral receptor profiles. Furthermore by measuring binding kinetic constants of solution-phase glycans to immobilized viruses, we confirmed that the glycan-HA affinity constant is in the range of 10 mM and the reaction is enthalpy-driven.

Spectroscopic analyses and studies on respective interaction of cyanuric acid and uric acid with bovine serum albumin and melamine

NASA Astrophysics Data System (ADS)

Chen, Dandan; Wu, Qiong; Wang, Jun; Wang, Qi; Qiao, Heng

2015-01-01

In this work, the fluorescence quenching was used to study the interaction of cyanuric acid (CYA) and uric acid (UA) with bovine serum albumin (BSA) at two different temperatures (283 K and 310 K). The bimolecular quenching constant (Kq), apparent quenching constant (Ksv), effective binding constant (KA) and corresponding dissociation constant (KD), binding site number (n) and binding distance (r) were calculated by adopting Stern-Volmer, Lineweaver-Burk, Double logarithm and overlap integral equations. The results show that CYA and UA are both able to obviously bind to BSA, but the binding strength order is BSA + CYA < BSA + UA. And then, the interactions of CYA and UA with melamine (MEL) under the same conditions were also studied by using similar methods. The results indicates that both CYA and UA can bind together closely with melamine (MEL). It is wished that these research results would facilitate the understanding the formation of kidney stones and gout in the body after ingesting excess MEL.

Molecular recognition of nucleotides in micelles and the development and expansion of a chemistry outreach program

NASA Astrophysics Data System (ADS)

Schechinger, Linda Sue

I. To investigate the delivery of nucleotide-based drugs, we are studying molecular recognition of nucleotide derivatives in environments that are similar to cell membranes. The Nowick group previously discovered that membrane-like surfactant micelles tetradecyltrimethylammonium bromide (TTAB) micelle facilitate molecular of adenosine monophosphate (AMP) recognition. The micelles bind nucleotides by means of electrostatic interactions and hydrogen bonding. We observed binding by following 1H NMR chemical shift changes of unique hexylthymine protons upon addition of AMP. Cationic micelles are required for binding. In surfactant-free or sodium dodecylsulfate solutions, no hydrogen bonding is observed. These observations suggest that the cationic surfactant headgroups bind the nucleotide phosphate group, while the intramicellar base binds the nucleotide base. The micellar system was optimized to enhance binding and selectivity for adenosine nucleotides. The selectivity for adenosine and the number of phosphate groups attached to the adenosine were both investigated. Addition of cytidine, guanidine, or uridine monophosphates, results in no significant downfield shifting of the NH resonance. Selectivity for the phosphate is limited, since adenosine mono-, di-, and triphosphates all have similar binding constants. We successfully achieved molecular recognition of adenosine nucleotides in micellar environments. There is significant difference in the binding interactions between the adenosine nucleotides and three other natural nucleotides. II. The UCI Chemistry Outreach Program (UCICOP) addresses the declining interest of the nations youth for science. UCICOP brings fun and exciting chemistry experiments to local high schools, to remind students that science is fun and has many practical uses. Volunteer students and alumni of UCI perform the demonstrations using scripts and material provided by UCICOP. The preparation of scripts and materials is done by two coordinators. These coordinators organize the program and provide continuity to the program. The success of UCICOP can be measured by the high praise and gratitude expressed by the teachers, students and volunteers.

WE-AB-207B-06: Dose and Biological Uncertainties in Sarcoma

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

Marteinsdottir, M; University of Iceland, Reykjavik; Schuemann, J

2016-06-15

Purpose: To understand the clinical impact of key uncertainties in proton therapy potentially affecting the analysis of clinical trials, namely the assumption of using a constant relative biological effectiveness (RBE) of 1.1 compared to variable RBE for proton therapy and the use of analytical dose calculation (ADC) methods. Methods: Proton dose distributions were compared for analytical and Monte Carlo (TOPAS) dose calculations. In addition, differences between using a constant RBE of 1.1 (RBE-constant) were compared with four different RBE models (to assess model variations). 10 patients were selected from an ongoing clinical trial on IMRT versus scanned protons for sarcoma.more » Comparisons were performed using dosimetric indices based on dose-volume histogram analyses and γ-index analyses. Results: For three of the RBE-models the mean dose, D95, D50 and D02 (dose values covering 95%, 50% and 2% of the target volume, respectively) were up to 5% lower than for RBE-constant. The dosimetric indices for one of the RBE-models were around 9% lower than for the RBE-constant model. The differences for V90 (the percentage of the target volume covered by 90% of the prescription dose) were up to 40% for three RBE-models, whereas for one the difference was around 95%. All ADC dosimetric indices were up to 5% larger than for RBE-constant. The γ-index passing rate for the target volume with a 3%/3mm criterion was above 97% for all models except for one, which was below 24%. Conclusion: Interpretation of clinical trials on sarcoma may depend on dose calculation uncertainties (as assessed by Monte Carlo). In addition, the biological dose distribution depends notably on which RBE model is utilized. The current practice of using a constant RBE of 1.1 may overestimate the target dose by as much as 5% for biological dose calculations. Performing an RBE uncertainty analysis is recommended for trial analysis. U19 projects - U19 CA 021239. PI: Delaney.« less

Cation Hydration Constants by Proton NMR: A Physical Chemistry Experiment.

ERIC Educational Resources Information Center

Smith, Robert L.; And Others

1988-01-01

Studies the polarization effect on water by cations and anions. Describes an experiment to illustrate the polarization effect of sodium, lithium, calcium, and strontium ions on the water molecule in the hydration spheres of the ions. Analysis is performed by proton NMR. (MVL)

Molecular switching behavior in isosteric DNA base pairs.

PubMed

Jissy, A K; Konar, Sukanya; Datta, Ayan

2013-04-15

The structures and proton-coupled behavior of adenine-thymine (A-T) and a modified base pair containing a thymine isostere, adenine-difluorotoluene (A-F), are studied in different solvents by dispersion-corrected density functional theory. The stability of the canonical Watson-Crick base pair and the mismatched pair in various solvents with low and high dielectric constants is analyzed. It is demonstrated that A-F base pairing is favored in solvents with low dielectric constant. The stabilization and conformational changes induced by protonation are also analyzed for the natural as well as the mismatched base pair. DNA sequences capable of changing their sequence conformation on protonation are used in the construction of pH-based molecular switches. An acidic medium has a profound influence in stabilizing the isostere base pair. Such a large gain in stability on protonation leads to an interesting pH-controlled molecular switch, which can be incorporated in a natural DNA tract. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Protonation states of histidine and other key residues in deoxy normal human adult hemoglobin by neutron protein crystallography

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

Kovalevsky, Andrey, E-mail: ayk@lanl.gov; Chatake, Toshiyuki; Shibayama, Naoya

2010-11-01

Using neutron diffraction analysis, the protonation states of 35 of 38 histidine residues were determined for the deoxy form of normal human adult hemoglobin. Distal and buried histidines may contribute to the increased affinity of the deoxy state for hydrogen ions and its decreased affinity for oxygen compared with the oxygenated form. The protonation states of the histidine residues key to the function of deoxy (T-state) human hemoglobin have been investigated using neutron protein crystallography. These residues can reversibly bind protons, thereby regulating the oxygen affinity of hemoglobin. By examining the OMIT F{sub o} − F{sub c} and 2F{sub o}more » − F{sub c} neutron scattering maps, the protonation states of 35 of the 38 His residues were directly determined. The remaining three residues were found to be disordered. Surprisingly, seven pairs of His residues from equivalent α or β chains, αHis20, αHis50, αHis58, αHis89, βHis63, βHis143 and βHis146, have different protonation states. The protonation of distal His residues in the α{sub 1}β{sub 1} heterodimer and the protonation of αHis103 in both subunits demonstrates that these residues may participate in buffering hydrogen ions and may influence the oxygen binding. The observed protonation states of His residues are compared with their ΔpK{sub a} between the deoxy and oxy states. Examination of inter-subunit interfaces provided evidence for interactions that are essential for the stability of the deoxy tertiary structure.« less

Comparison of solute-binding properties of plastic materials used as pharmaceutical product containers.

PubMed

Jenke, Dennis; Couch, Tom; Gillum, Amy

2010-01-01

Material/water equilibrium binding constants (E(b)) were determined for 11 organic solutes and 2 plastic materials commonly used in pharmaceutical product containers (plasticized polyvinyl chloride and polyolefin). In general, solute binding by the plasticized polyvinyl chloride material was greater, by nearly an order of magnitude, than the binding by the polyolefin (on an equal weight basis). The utilization of the binding constants to facilitate container compatibility assessments (e.g., drug loss by container binding) for drug-containing products is discussed.

A novel methodological approach for the analysis of host-ligand interactions.

PubMed

Strat, Daniela; Missailidis, Sotiris; Drake, Alex F

2007-02-02

Traditional analysis of drug-binding data relies upon the Scatchard formalism. These methods rely upon the fitting of a linear equation providing intercept and gradient data that relate to physical properties, such as the binding constant, cooperativity coefficients and number of binding sites. However, the existence of different binding modes with different binding constants makes the implementation of these models difficult. This article describes a novel approach to the binding model of host-ligand interactions by using a derived analytical function describing the observed signal. The benefit of this method is that physically significant parameters, that is, binding constants and number of binding sites, are automatically derived by the use of a minimisation routine. This methodology was utilised to analyse the interactions between a novel antitumour agent and DNA. An optical spectroscopy study confirms that the pentacyclic acridine derivative (DH208) binds to nucleic acids. Two binding modes can be identified: a stronger one that involves intercalation and a weaker one that involves oriented outer-sphere binding. In both cases the plane of the bound acridine ring is parallel to the nucleic acid bases, orthogonal to the phosphate backbone. Ultraviolet (UV) and circular dichroism (CD) data were fitted using the proposed model. The binding constants and the number of binding sites derived from the model remained consistent across the different techniques used. The different wavelengths at which the measurements were made maintained the coherence of the results.

The Mg2+-containing Water Cluster of Mammalian Cytochrome c Oxidase Collects Four Pumping Proton Equivalents in Each Catalytic Cycle*

PubMed Central

Yano, Naomine; Muramoto, Kazumasa; Shimada, Atsuhiro; Takemura, Shuhei; Baba, Junpei; Fujisawa, Hidenori; Mochizuki, Masao; Shinzawa-Itoh, Kyoko; Yamashita, Eiki; Tsukihara, Tomitake; Yoshikawa, Shinya

2016-01-01

Bovine heart cytochrome c oxidase (CcO) pumps four proton equivalents per catalytic cycle through the H-pathway, a proton-conducting pathway, which includes a hydrogen bond network and a water channel operating in tandem. Protons are transferred by H3O+ through the water channel from the N-side into the hydrogen bond network, where they are pumped to the P-side by electrostatic repulsion between protons and net positive charges created at heme a as a result of electron donation to O2 bound to heme a3. To block backward proton movement, the water channel remains closed after O2 binding until the sequential four-proton pumping process is complete. Thus, the hydrogen bond network must collect four proton equivalents before O2 binding. However, a region with the capacity to accept four proton equivalents was not discernable in the x-ray structures of the hydrogen bond network. The present x-ray structures of oxidized/reduced bovine CcO are improved from 1.8/1.9 to 1.5/1.6 Å resolution, increasing the structural information by 1.7/1.6 times and revealing that a large water cluster, which includes a Mg2+ ion, is linked to the H-pathway. The cluster contains enough proton acceptor groups to retain four proton equivalents. The redox-coupled x-ray structural changes in Glu198, which bridges the Mg2+ and CuA (the initial electron acceptor from cytochrome c) sites, suggest that the CuA-Glu198-Mg2+ system drives redox-coupled transfer of protons pooled in the water cluster to the H-pathway. Thus, these x-ray structures indicate that the Mg2+-containing water cluster is the crucial structural element providing the effective proton pumping in bovine CcO. PMID:27605664

The Mg2+-containing Water Cluster of Mammalian Cytochrome c Oxidase Collects Four Pumping Proton Equivalents in Each Catalytic Cycle.

PubMed

Yano, Naomine; Muramoto, Kazumasa; Shimada, Atsuhiro; Takemura, Shuhei; Baba, Junpei; Fujisawa, Hidenori; Mochizuki, Masao; Shinzawa-Itoh, Kyoko; Yamashita, Eiki; Tsukihara, Tomitake; Yoshikawa, Shinya

2016-11-11

Bovine heart cytochrome c oxidase (CcO) pumps four proton equivalents per catalytic cycle through the H-pathway, a proton-conducting pathway, which includes a hydrogen bond network and a water channel operating in tandem. Protons are transferred by H 3 O + through the water channel from the N-side into the hydrogen bond network, where they are pumped to the P-side by electrostatic repulsion between protons and net positive charges created at heme a as a result of electron donation to O 2 bound to heme a 3 To block backward proton movement, the water channel remains closed after O 2 binding until the sequential four-proton pumping process is complete. Thus, the hydrogen bond network must collect four proton equivalents before O 2 binding. However, a region with the capacity to accept four proton equivalents was not discernable in the x-ray structures of the hydrogen bond network. The present x-ray structures of oxidized/reduced bovine CcO are improved from 1.8/1.9 to 1.5/1.6 Å resolution, increasing the structural information by 1.7/1.6 times and revealing that a large water cluster, which includes a Mg 2+ ion, is linked to the H-pathway. The cluster contains enough proton acceptor groups to retain four proton equivalents. The redox-coupled x-ray structural changes in Glu 198 , which bridges the Mg 2+ and Cu A (the initial electron acceptor from cytochrome c) sites, suggest that the Cu A -Glu 198 -Mg 2+ system drives redox-coupled transfer of protons pooled in the water cluster to the H-pathway. Thus, these x-ray structures indicate that the Mg 2+ -containing water cluster is the crucial structural element providing the effective proton pumping in bovine CcO. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Interaction of cinnamic acid derivatives with serum albumins: A fluorescence spectroscopic study

NASA Astrophysics Data System (ADS)

Singh, T. Sanjoy; Mitra, Sivaprasad

2011-03-01

Cinnamic acid (CA) derivatives are known to possess broad therapeutic applications including anti-tumor activity. The present study was designed to determine the underlying mechanism and thermodynamic parameters for the binding of two CA based intramolecular charge transfer (ICT) fluorescent probes, namely, 4-(dimethylamino) cinnamic acid (DMACA) and trans-ethyl p-(dimethylamino) cinnamate (EDAC), with albumins by fluorescence spectroscopy. Stern-Volmer analysis of the tryptophan fluorescence quenching data in presence of the added ligand reveals fluorescence quenching constant ( κq), Stern-Volmer constant ( KSV) and also the ligand-protein association constant ( Ka). The thermodynamic parameters like enthalpy (Δ H) and entropy (Δ S) change corresponding to the ligand binding process were also estimated. The results show that the ligands bind into the sub-domain IIA of the proteins in 1:1 stoichiometry with an apparent binding constant value in the range of 10 4 dm 3 mol -1. In both the cases, the spontaneous ligand binding to the proteins occur through entropy driven mechanism, although the interaction of DMACA is relatively stronger in comparison with EDAC. The temperature dependence of the binding constant indicates the induced change in protein secondary structure.

SU-E-T-353: Verification of Water Equivalent Thickness (WET) and Water Equivalent Spreadness (WES) of Proton Beam

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

Demez, N; Lee, T; Keppel, Cynthia

Purpose: To verify calculated water equivalent thickness (WET) and water equivalent spreadness (WES) in various tissue equivalent media for proton therapy Methods: Water equivalent thicknesses (WET) of tissue equivalent materials have been calculated using the Bragg-Kleeman rule. Lateral spreadness and fluence reduction of proton beams both in those media were calculated using proton loss model (PLM) algorithm. In addition, we calculated lateral spreadness ratios with respect to that in water at the same WET depth and so the WES was defined. The WETs of those media for different proton beam energies were measured using MLIC (Multi-Layered Ionization Chamber). Also, fluencemore » and field sizes in those materials of various thicknesses were measured with ionization chambers and films Results: Calculated WETs are in agreement with measured WETs within 0.5%. We found that water equivalent spreadness (WES) is constant and the fluence and field size measurements verify that fluence can be estimated using the concept of WES. Conclusions: Calculation of WET based on the Bragg-Kleeman rule as well as the constant WES of proton beams for tissue equivalent phantoms can be used to predict fluence and field sizes at the depths of interest both in tissue equivalent media accurately for clinically available protonenergies.« less

The Use of Hammett Constants to Understand the Non-Covalent Binding of Aromatics

PubMed Central

Lewis, Michael; Bagwill, Christina; Hardebeck, Laura K. E.; Wireduaah, Selina

2012-01-01

Non-covalent interactions of aromatics are important in a wide range of chemical and biological applications. The past two decades have seen numerous reports of arene-arene binding being understood in terms Hammett substituent constants, and similar analyses have recently been extended to cation-arene and anion-arene binding. It is not immediately clear why electrostatic Hammett parameters should work so well in predicting the binding for all three interactions, given that different intermolecular forces dominate each interaction. This review explores such anomalies, and summarizes how Hammett substituent constants have been employed to understand the non-covalent binding in arene-arene, cation-arene and anion-arene interactions. PMID:24688634

Influence of Ca and pH on the uptake and effects of Cd in Folsomia candida exposed to simplified soil solutions.

PubMed

Ardestani, Masoud M; Ortiz, Maria Diez; van Gestel, Cornelis A M

2013-08-01

The present study sought to quantify the components of a biotic ligand model (BLM) for the effects of Cd on Folsomia candida (Collembola). Assuming that soil porewater is the main route of exposure and to exclude the effects of soil particles on metal availability, animals were exposed for 7 d to different Cd concentrations between 0.1 mM and 100 mM in simplified soil solutions at different Ca concentrations (0.2 mM, 0.8 mM, 3.2 mM, and 12.8 mM) or at different pH (5.0, 6.0, and 7.0). Higher Ca concentrations decreased the toxicity of Cd (adult survival) in test solutions, whereas toxicity was slightly lower at pH 7 and 6 than at pH 5, suggesting a mitigating effect of Ca and to a lesser extent pH on Cd toxicity to F. candida. Internal Cd concentrations in the animals increased with increasing exposure level but were significantly reduced by increasing Ca concentrations and were not significantly affected by pH. By using Langmuir isotherms, binding constants for Cd, Ca, and protons and the fraction of binding sites occupied by Cd were calculated and used to predict effects of Cd on survival. Predicted toxicity showed a good agreement with measured responses when Ca and pH were used as separate factors or combined together. The present study shows indications of protective effects of Ca but less of protons on the toxicity and uptake of Cd in F. candida on exposure to simplified soil solutions, which can be described using the principles of a biotic ligand model. Copyright © 2013 SETAC.

Substrate-bound structure of the E. coli multidrug resistance transporter MdfA

PubMed Central

Heng, Jie; Zhao, Yan; Liu, Ming; Liu, Yue; Fan, Junping; Wang, Xianping; Zhao, Yongfang; Zhang, Xuejun C

2015-01-01

Multidrug resistance is a serious threat to public health. Proton motive force-driven antiporters from the major facilitator superfamily (MFS) constitute a major group of multidrug-resistance transporters. Currently, no reports on crystal structures of MFS antiporters in complex with their substrates exist. The E. coli MdfA transporter is a well-studied model system for biochemical analyses of multidrug-resistance MFS antiporters. Here, we report three crystal structures of MdfA-ligand complexes at resolutions up to 2.0 Å, all in the inward-facing conformation. The substrate-binding site sits proximal to the conserved acidic residue, D34. Our mutagenesis studies support the structural observations of the substrate-binding mode and the notion that D34 responds to substrate binding by adjusting its protonation status. Taken together, our data unveil the substrate-binding mode of MFS antiporters and suggest a mechanism of transport via this group of transporters. PMID:26238402

Direct detection of formate ligation in cytochrome c oxidase by ATR-FTIR spectroscopy.

PubMed

Iwaki, Masayo; Rich, Peter R

2004-03-03

The IR signature of binding of formate to the heme a(3-)Cu(B) binuclear site of bovine cytochrome c oxidase has been obtained by perfusion ATR-FTIR spectroscopy. The data show unequivocally that formate binds in its anionic form despite its binding being electroneutral overall. The bound formate can be distinguished from free ligand by the binding-induced sharpening and downshifting of vibrational bands. Formate ligation also causes shifts of vibrational modes of heme a(3) and its substituents and perturbation of histidine residues. The association of the accompanying protonation change with a carboxylate or tyrosine can be ruled out and may involve a histidine metal ligand or, more likely, a simple displacement into the bulk phase of a hydroxide ligand to heme a(3) or CU(B), a reaction which would account for stoichiometric proton uptake and maintenance of net charge within the binuclear center domain.

Mannobiose Binding Induces Changes in Hydrogen Bonding and Protonation States of Acidic Residues in Concanavalin A As Revealed by Neutron Crystallography

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

Gerlits, Oksana O.; Coates, Leighton; Woods, Robert J.

Plant lectins are carbohydrate-binding proteins with various biomedical applications. Concanavalin A (Con A) holds promise in treating cancerous tumors. To better understand the Con A carbohydrate binding specificity, we obtained a room-temperature neutron structure of this legume lectin in complex with a disaccharide Manα1–2Man, mannobiose. The neutron structure afforded direct visualization of the hydrogen bonding between the protein and ligand, showing that the ligand is able to alter both protonation states and interactions for residues located close to and distant from the binding site. An unprecedented low-barrier hydrogen bond was observed forming between the carboxylic side chains of Asp28 andmore » Glu8, with the D atom positioned equidistant from the oxygen atoms having an O···D···O angle of 101.5°.« less

Ph-Dependent Inhibition of Voltage-Gated H+ Currents in Rat Alveolar Epithelial Cells by Zn2+ and Other Divalent Cations

PubMed Central

Cherny, Vladimir V.; DeCoursey, Thomas E.

1999-01-01

Inhibition by polyvalent cations is a defining characteristic of voltage-gated proton channels. The mechanism of this inhibition was studied in rat alveolar epithelial cells using tight-seal voltage clamp techniques. Metal concentrations were corrected for measured binding to buffers. Externally applied ZnCl2 reduced the H+ current, shifted the voltage-activation curve toward positive potentials, and slowed the turn-on of H+ current upon depolarization more than could be accounted for by a simple voltage shift, with minimal effects on the closing rate. The effects of Zn2+ were inconsistent with classical voltage-dependent block in which Zn2+ binds within the membrane voltage field. Instead, Zn2+ binds to superficial sites on the channel and modulates gating. The effects of extracellular Zn2+ were strongly pHo dependent but were insensitive to pHi, suggesting that protons and Zn2+ compete for external sites on H+ channels. The apparent potency of Zn2+ in slowing activation was ∼10× greater at pHo 7 than at pHo 6, and ∼100× greater at pHo 6 than at pHo 5. The pHo dependence suggests that Zn2+, not ZnOH+, is the active species. Evidently, the Zn2+ receptor is formed by multiple groups, protonation of any of which inhibits Zn2+ binding. The external receptor bound H+ and Zn2+ with pK a 6.2–6.6 and pK M 6.5, as described by several models. Zn2+ effects on the proton chord conductance–voltage (g H–V) relationship indicated higher affinities, pK a 7 and pK M 8. CdCl2 had similar effects as ZnCl2 and competed with H+, but had lower affinity. Zn2+ applied internally via the pipette solution or to inside-out patches had comparatively small effects, but at high concentrations reduced H+ currents and slowed channel closing. Thus, external and internal zinc-binding sites are different. The external Zn2+ receptor may be the same modulatory protonation site(s) at which pHo regulates H+ channel gating. PMID:10578017

Modeling solubility, acid-base properties and activity coefficients of amoxicillin, ampicillin and (+)6-aminopenicillanic acid, in NaCl(aq) at different ionic strengths and temperatures.

PubMed

Crea, Francesco; Cucinotta, Daniela; De Stefano, Concetta; Milea, Demetrio; Sammartano, Silvio; Vianelli, Giuseppina

2012-11-20

The total solubility of three penicillin derivatives was determined, in pure water and NaCl aqueous solutions at different salt concentrations (from ∼0.15 to 1.0 mol L(-1) for ampicillin and amoxicillin, and from ∼0.05 to 2.0 mol L(-1) for (+)6-aminopenicillanic acid), using the shake-flask method for generating the saturated solutions, followed by potentiometric analysis. The knowledge of the pH of solubilization and of the protonation constants determined in the same experimental conditions, allowed us to calculate, by means of the mass balance equations, the solubility of the neutral species at different ionic strength values, to model its dependence on the salt concentration and to determine the corresponding values at infinite dilution. The salting parameter and the activity coefficients of the neutral species were calculated by the Setschenow equation. The protonation constants of ampicillin and amoxicillin, determined at different temperatures (from T=288.15 to 318.15K), from potentiometric and spectrophotometric measurements, were used to calculate, by means of the Van't Hoff equation, the temperature coefficients at different ionic strength values and the corresponding protonation entropies. The protonation enthalpies of the (+)6-aminopenicillanic acid were determined by isoperibol calorimetric titrations at T=298.15K and up to I=2.0 mol L(-1). The dependence of the protonation constants on ionic strength was modeled by means of the Debye-Hückel and SIT (Specific ion Interaction Theory) approaches, and the specific interaction parameters of the ionic species were determined. The hydrolysis of the β-lactam ring was studied by spectrophotometric and H NMR investigations as a function of pH, ionic strength and time. Potentiometric measurements carried out on the hydrolyzed (+)6-aminopenicillanic acid allowed us to highlight that the opened and the closed β-lactam forms of the (+)6-aminopenicillanic acid have quite different acid-base properties. An analysis of literature solubility, protonation constants, enthalpies and activity coefficients is reported too. Copyright © 2012 Elsevier B.V. All rights reserved.

Lack of conventional oxygen-linked proton and anion binding sites does not impair allosteric regulation of oxygen binding in dwarf caiman hemoglobin

PubMed Central

Fago, Angela; Malte, Hans; Storz, Jay F.; Gorr, Thomas A.

2013-01-01

In contrast to other vertebrate hemoglobins (Hbs) whose high intrinsic O2 affinities are reduced by red cell allosteric effectors (mainly protons, CO2, organic phosphates, and chloride ions), crocodilian Hbs exhibit low sensitivity to organic phosphates and high sensitivity to bicarbonate (HCO3−), which is believed to augment Hb-O2 unloading during diving and postprandial alkaline tides when blood HCO3− levels and metabolic rates increase. Examination of α- and β-globin amino acid sequences of dwarf caiman (Paleosuchus palpebrosus) revealed a unique combination of substitutions at key effector binding sites compared with other vertebrate and crocodilian Hbs: β82Lys→Gln, β143His→Val, and β146His→Tyr. These substitutions delete positive charges and, along with other distinctive changes in residue charge and polarity, may be expected to disrupt allosteric regulation of Hb-O2 affinity. Strikingly, however, P. palpebrosus Hb shows a strong Bohr effect, and marked deoxygenation-linked binding of organic phosphates (ATP and DPG) and CO2 as carbamate (contrasting with HCO3− binding in other crocodilians). Unlike other Hbs, it polymerizes to large complexes in the oxygenated state. The highly unusual properties of P. palpebrosus Hb align with a high content of His residues (potential sites for oxygenation-linked proton binding) and distinctive surface Cys residues that may form intermolecular disulfide bridges upon polymerization. On the basis of its singular properties, P. palpebrosus Hb provides a unique opportunity for studies on structure-function coupling and the evolution of compensatory mechanisms for maintaining tissue O2 delivery in Hbs that lack conventional effector-binding residues. PMID:23720132

Mechanistic Insight from Calorimetric Measurements of the Assembly of the Binuclear Metal Active Site of Glycerophosphodiesterase (GpdQ) from Enterobacter aerogenes.

PubMed

Pedroso, Marcelo M; Ely, Fernanda; Carpenter, Margaret C; Mitić, Nataša; Gahan, Lawrence R; Ollis, David L; Wilcox, Dean E; Schenk, Gerhard

2017-07-05

Glycerophosphodiesterase (GpdQ) from Enterobacter aerogenes is a binuclear metallohydrolase with a high affinity for metal ions at its α site but a lower affinity at its β site in the absence of a substrate. Isothermal titration calorimetry (ITC) has been used to quantify the Co(II) and Mn(II) binding affinities and thermodynamics of the two sites in wild-type GpdQ and two mutants, both in the absence and in the presence of phosphate. Metal ions bind to the six-coordinate α site in an entropically driven process with loss of a proton, while binding at the β site is not detected by ITC. Phosphate enhances the metal affinity of the α site by increasing the binding entropy and the metal affinity of the β site by enthalpic (Co) or entropic (Mn) contributions, but no additional loss of protons. Mutations of first- and second-coordination sphere residues at the β site increase the metal affinity of both sites by enhancing the binding enthalpy. In particular, loss of the hydrogen bond from second-sphere Ser127 to the metal-coordinating Asn80 has a significant effect on the metal binding thermodynamics that result in a resting binuclear active site with high catalytic activity. While structural and spectroscopic data with excess metal ions have indicated a bridging hydroxide in the binuclear GpdQ site, analysis of ITC data here reveals the loss of a single proton in the assembly of this site, indicating that the metal-bound hydroxide nucleophile is formed in the resting inactive mononuclear form, which becomes catalytically competent upon binding the second metal ion.

Energetics and kinetics of cooperative cofilin-actin filament interactions.

PubMed

Cao, Wenxiang; Goodarzi, Jim P; De La Cruz, Enrique M

2006-08-11

We have evaluated the thermodynamic parameters associated with cooperative cofilin binding to actin filaments, accounting for contributions of ion-linked equilibria, and determined the kinetic basis of cooperative cofilin binding. Ions weaken non-contiguous (isolated, non-cooperative) cofilin binding to an actin filament without affecting cooperative filament interactions. Non-contiguous cofilin binding is coupled to the dissociation of approximately 1.7 thermodynamically bound counterions. Counterion dissociation contributes approximately 40% of the total cofilin binding free energy (in the presence of 50 mM KCl). The non-contiguous and cooperative binding free energies are driven entirely by large, positive entropy changes, consistent with a cofilin-mediated increase in actin filament structural dynamics. The rate constant for cofilin binding to an isolated site on an actin filament is slow and likely to be limited by filament breathing. Cooperative cofilin binding arises from an approximately tenfold more rapid association rate constant and an approximately twofold slower dissociation rate constant. The more rapid association rate constant is presumably a consequence of cofilin-dependent changes in the average orientation of subdomain 2, subunit angular disorder and filament twist, which increase the accessibility of a neighboring cofilin-binding site on an actin filament. Cooperative association is more rapid than binding to an isolated site, but still slow for a second-order reaction, suggesting that cooperative binding is limited also by binding site accessibility. We suggest that the dissociation of actin-associated ions weakens intersubunit interactions in the actin filament lattice that enhance cofilin-binding site accessibility, favor cooperative binding and promote filament severing.

A Cr(VI) selective probe based on a quinoline-amide calix[4]arene

NASA Astrophysics Data System (ADS)

Ferreira, Juliane F.; Bagatin, Izilda A.

2018-01-01

A new quinoline-amide calix[4]arene 3-receptor for detection of hazardous anions and cations have been synthesized. The 3-receptor was examined for its sensing properties towards several different anions (Cr2O72 -, SCN-, F-, Cl-, NO3-) and metal ions (Hg2+, Cd2+, Ag+) by UV-vis and fluorescence spectroscopies. It was detected that the 3-receptor has only sensing ability for Cr2O72 - and Hg2+ ions, resulting in the association constants higher for Cr2O72 - than to the Hg2+ ions. High selectivity towards Cr2O72 - were also observed by fluorescence measurement among other ions (F-, Cl-, SCN-, Hg2+, Cd2+, Ag+) with a low limit of detection (7.36 × 10-6 mol dm-3). Proton NMR anion-binding investigations revealed a strong interaction of Cr2O72 - anion with NH and CH groups of the receptor, showing that the combination with hydrogen-bonds donor groups strengthened the anion receptor association. Furthermore, remarkable association constants for dichromate anion obtained by all techniques strongly suggest the 3-receptor as a selective Cr(VI) sensor.

Binding the diproton in stars: anthropic limits on the strength of gravity

NASA Astrophysics Data System (ADS)

Barnes, Luke A.

2015-12-01

We calculate the properties and investigate the stability of stars that burn via strong (and electromagnetic) interactions, and compare their properties with those that, as in our Universe, include a rate-limiting weak interaction. It has been suggested that, if the diproton were bound, stars would burn ~1018 times brighter and faster via strong interactions, resulting in a universe that would fail to support life. By considering the representative case of a star in our Universe with initially equal numbers of protons and deuterons, we find that stable, "strong-burning" stars adjust their central densities and temperatures to have familiar surface temperatures, luminosities and lifetimes. There is no "diproton disaster". In addition, strong-burning stars are stable in a much larger region of the parameter space of fundamental constants, specifically the strength of electromagnetism and gravity. The strongest anthropic bound on stars in such universes is not their stability, as is the case for stars limited by the weak interaction, but rather their lifetime. Regardless of the strength of electromagnetism, all stars burn out in mere millions of years unless the gravitational coupling constant is extremely small, αGlesssim 10-30.

A Cr(VI) selective probe based on a quinoline-amide calix[4]arene.

PubMed

Ferreira, Juliane F; Bagatin, Izilda A

2018-01-15

A new quinoline-amide calix[4]arene 3-receptor for detection of hazardous anions and cations have been synthesized. The 3-receptor was examined for its sensing properties towards several different anions (Cr 2 O 7 2- , SCN - , F - , Cl - , NO 3 - ) and metal ions (Hg 2+ , Cd 2+ , Ag + ) by UV-vis and fluorescence spectroscopies. It was detected that the 3-receptor has only sensing ability for Cr 2 O 7 2- and Hg 2+ ions, resulting in the association constants higher for Cr 2 O 7 2- than to the Hg 2+ ions. High selectivity towards Cr 2 O 7 2- were also observed by fluorescence measurement among other ions (F - , Cl - , SCN - , Hg 2+ , Cd 2+ , Ag + ) with a low limit of detection (7.36×10 -6 moldm -3 ). Proton NMR anion-binding investigations revealed a strong interaction of Cr 2 O 7 2- anion with NH and CH groups of the receptor, showing that the combination with hydrogen-bonds donor groups strengthened the anion receptor association. Furthermore, remarkable association constants for dichromate anion obtained by all techniques strongly suggest the 3-receptor as a selective Cr(VI) sensor. Copyright © 2017 Elsevier B.V. All rights reserved.

Spectroscopic, potentiometric and theoretical studies on the binding properties of a novel tripodal polycatechol-imine ligand towards iron(III)

NASA Astrophysics Data System (ADS)

Kanungo, B. K.; Sahoo, Suban K.; Baral, Minati

2008-12-01

A novel multidentate tripodal ligand, cis, cis-1,3,5-tris[(2,3-dihydroxybenzylidene)aminomethyl]cyclohexane (TDBAC, L) containing one catechol unit in each arms of a tripodal amine, cis, cis-1,3,5-tris(aminomethyl)cyclohexane was investigated as a chelator for iron(III) through potentiometric and spectrophotometric methods in an aqueous medium of 0.1N ionic strength and 25 ± 1 °C as well as in ethanol by continuous variation method. From pH metric in water, three protonation constants characterized for the three-hydroxyl groups of the catechol units at ortho were used as input data to evaluate the stability constants of the complexes. Formation of monomeric complexes FeLH 3, FeLH 2, FeLH and FeL were depicted. In ethanol, formation of complexes FeL, Fe 2L and Fe 3L were characterized. Structures of the complexes were explained by using the experimental evidences and predicted through molecular modeling calculations. The ligand showed potential to coordinate iron(III) through three imine nitrogens and three catecholic oxygens at ortho to form a tris(iminocatecholate) type complex.

ENDOR/ESR of Mn atoms and MnH molecules in solid argon

NASA Astrophysics Data System (ADS)

van Zee, R. J.; Garland, D. A.; Weltner, W., Jr.

1986-09-01

Mn atoms and MnH molecules, the latter formed by reaction between metal and hydrogen atoms, were trapped in solid argon and their ESR/ENDOR spectra measured at 4 K. At each pumping magnetic field two ENDOR lines were observed for 55Mn(I=5/2) atoms, corresponding to hyperfine transitions within the MS =±1/2 levels. Values of the hyperfine interaction constant and nuclear moment of 55Mn were derived from the six sets of data. For MnH, three sets of signals were detected: a proton ``matrix ENDOR'' line, transitions in the MS =0,±1 levels involving MI (55Mn)=1/2, 3/2, 5/2 levels, and proton transitions corresponding to νH and νH±aH. Analysis yielded the hyperfine constant aH =6.8(1) MHz and the nuclear quadrupole coupling constant Q'(55Mn)=-11.81(2) MHz. The latter compared favorably with a theoretical value derived earlier by Bagus and Schaefer. A higher term in the spin Hamiltonian appeared to be necessary to fit the proton hyperfine data.

Spin relaxation measurements of electrostatic bias in intermolecular exploration

NASA Astrophysics Data System (ADS)

Teng, Ching-Ling; Bryant, Robert G.

2006-04-01

We utilize the paramagnetic contribution to proton spin-lattice relaxation rate constants induced by freely diffusing charged paramagnetic centers to investigate the effect of charge on the intermolecular exploration of a protein by the small molecule. The proton NMR spectrum provided 255 resolved resonances that report how the explorer molecule local concentration varies with position on the surface. The measurements integrate over local dielectric constant variations, and, in principle, provide an experimental characterization of the surface free energy sampling biases introduced by the charge distribution on the protein. The experimental results for ribonuclease A obtained using positive, neutral, and negatively charged small nitroxide radicals are qualitatively similar to those expected from electrostatic calculations. However, while systematic electrostatic trends are apparent, the three different combinations of the data sets do not yield internally consistent values for the electrostatic contribution to the intermolecular free energy. We attribute this failure to the weakness of the electrostatic sampling bias for charged nitroxides in water and local variations in effective translational diffusion constant at the water-protein interface, which enters the nuclear spin relaxation equations for the nitroxide-proton dipolar coupling.

Zinc sorption to three gram-negative bacteria: combined titration, modeling, and EXAFS study.

PubMed

Guiné, V; Spadini, L; Sarret, G; Muris, M; Delolme, C; Gaudet, J P; Martins, J M F

2006-03-15

The acid-base and Zn sorption properties of three bacteria, Cupriavidus metallidurans CH34, Pseudomonas putida ATCC12633, and Escherichia coli K12DH5alpha, were investigated through an original combination of extended X-ray absorption fine structure (EXAFS) spectroscopy and equilibrium titration studies. Acid-base titration curves of the three strains were fitted with a model accounting for three conceptual reactive sites: an acidic (carboxyl and/or phosphodiester), a neutral (phosphomonoester), and a basic (amine and/or hydroxyl) group. Calculated proton and Zn equilibrium constants and site densities compare with literature data. The nature of Zn binding sites was studied by EXAFS spectroscopy. Phosphoester, carboxyl, and unexpectedly sulfhydryl ligands were identified. Their proportions depended on Zn loading and bacterial strain and were consistent with the titration results. These findings were compared to the structure and site density of the major cell wall components. It appeared that the cumulated theoretical site density of these structures (<2 Zn nm(-2)) was much lower than the total site density of the investigated strains (16-56 Zn nm(-2)). These results suggest a dominant role of extracellular polymeric substances in Zn retention processes, although Zn binding to inner cell components cannot be excluded.

Dense proton injection into phosphate glasses using corona discharge treatment

NASA Astrophysics Data System (ADS)

Kinoshita, Takuya; Miyazaki, Atsushi; Kawaguchi, Keiga; Sakai, Daisuke; Yamaguchi, Takuya; Omata, Takahisa; Ishiyama, Tomohiro; Fujioka, Masaya; Kaiju, Hideo; Nishii, Junji

2018-01-01

Sodium ions in 25NaO1/2-6LaO3/2-6GeO2-63PO5/2 (mol%) glasses were substituted with protons using corona discharge treatment (CDT) under a H2 atmosphere. The substitution of sodium ion to proton proceeded from the anode side to the cathode side with constant current flow during the CDT. A crystalline free and transparent glass plate of 0.3 mm thickness was obtained after CDT for 96 h. The maximum decrease rate from sodium ion to proton was 78 ± 10%. The proton conductivity of 8.5 × 10-4 S/cm was attained at 400 °C.

Fine structure in the transition region: reaction force analyses of water-assisted proton transfers.

PubMed

Yepes, Diana; Murray, Jane S; Santos, Juan C; Toro-Labbé, Alejandro; Politzer, Peter; Jaque, Pablo

2013-07-01

We have analyzed the variation of the reaction force F(ξ) and the reaction force constant κ(ξ) along the intrinsic reaction coordinates ξ of the water-assisted proton transfer reactions of HX-N = Y (X,Y = O,S). The profile of the force constant of the vibration associated with the reactive mode, k ξ (ξ), was also determined. We compare our results to the corresponding intramolecular proton transfers in the absence of a water molecule. The presence of water promotes the proton transfers, decreasing the energy barriers by about 12 - 15 kcal mol(-1). This is due in part to much smaller bond angle changes being needed than when water is absent. The κ(ξ) profiles along the intrinsic reaction coordinates for the water-assisted processes show striking and intriguing differences in the transition regions. For the HS-N = S and HO-N = S systems, two κ(ξ) minima are obtained, whereas for HO-N = O only one minimum is found. The k ξ (ξ) show similar behavior in the transition regions. We propose that this fine structure reflects the degree of synchronicity of the two proton migrations in each case.

A bimodal activation mechanism underlies scorpion toxin–induced pain

PubMed Central

Yang, Shilong; Yang, Fan; Zhang, Bei; Lee, Bo Hyun; Li, Bowen; Luo, Lei; Zheng, Jie; Lai, Ren

2017-01-01

Venomous animals use peptide toxins for hunting and self-defense. To achieve these goals, toxins need to bind to their targets with high affinity due to the small amount that a single bite or sting can deliver. The scorpion toxin BmP01 is linked to sting-induced excruciating pain; however, the reported minimum concentrations for activating TRPV1 channel or inhibiting voltage-gated potassium (Kv) channels (both in the micromolar range) appear too high to be biologically relevant. We show that the effective concentration of BmP01 is highly pH-dependent—it increases by about 10-fold in inhibiting Kv channels upon a 1-U drop in pH but decreases more than 100-fold in activating TRPV1. Mechanistic investigation revealed that BmP01 binds to one of the two proton-binding sites on TRPV1 and, together with a proton, uses a one-two punch approach to strongly activate the nociceptive channel. Because most animal venoms are acidic, proton-facilitated synergistic action may represent a general strategy for maximizing toxin potency. PMID:28782041

Synthesis of β-arabinofuranoside glycolipids, studies of their binding to surfactant protein-A and effect on sliding motilities of M. smegmatis.

PubMed

Naresh, Kottari; Avaji, Prakash Gouda; Maiti, Krishnagopal; Bharati, Binod K; Syal, Kirtimaan; Chatterji, Dipankar; Jayaraman, Narayanaswamy

2012-04-01

Surfactant protein A (SP-A), which is a lung innate immune system component, is known to bind glycolipids present at the cell surface of a mycobacterial pathogen. Lipoarabinomannan (LAM), a component of mycobacterial thick, waxy cell wall, is one of the glycolipid ligands for SP-A. In order to assess binding of synthetic glycolipids with SP-A and the glycosidic linkage preferences for the interaction, β-arabinofuranoside trisaccharide glycolipids constituted with β-(1→2), β-(1→3) and β-(1→2), β-(1→5) linkages relevant to LAM were synthesized through chemical glycosylations. The efficacies of synthetic glycolipids to interact with SP-A were assessed by using the surface plasmon resonance (SPR) technique, from which association-dissociation rate constants and equilibrium binding constants were derived. The equilibrium binding constants of the interaction of two constitutionally varying β-arabinofuranoside glycolipids with SP-A were found to be in the millimolar range. A comparison of the results with few α-anomeric arabinofuranoside glycolipids showed that glycolipids with β-anomeric linkages were having relatively lower equilibrium binding constants than those with α-anomeric linkages in binding to the protein, whereas oligosaccharides alone, without lipidic chains, exhibited higher equilibrium binding constants. Further, the synthetic compounds inhibited the growth of mycobacteria and affected sliding motilities of the bacteria, although to an extent relatively lesser than that of synthetic compounds constituted with α-anomeric linkages.

Constant pH Molecular Dynamics in Explicit Solvent with Enveloping Distribution Sampling and Hamiltonian Exchange

PubMed Central

2015-01-01

We present a new computational approach for constant pH simulations in explicit solvent based on the combination of the enveloping distribution sampling (EDS) and Hamiltonian replica exchange (HREX) methods. Unlike constant pH methods based on variable and continuous charge models, our method is based on discrete protonation states. EDS generates a hybrid Hamiltonian of different protonation states. A smoothness parameter s is used to control the heights of energy barriers of the hybrid-state energy landscape. A small s value facilitates state transitions by lowering energy barriers. Replica exchange between EDS potentials with different s values allows us to readily obtain a thermodynamically accurate ensemble of multiple protonation states with frequent state transitions. The analysis is performed with an ensemble obtained from an EDS Hamiltonian without smoothing, s = ∞, which strictly follows the minimum energy surface of the end states. The accuracy and efficiency of this method is tested on aspartic acid, lysine, and glutamic acid, which have two protonation states, a histidine with three states, a four-residue peptide with four states, and snake cardiotoxin with eight states. The pKa values estimated with the EDS-HREX method agree well with the experimental pKa values. The mean absolute errors of small benchmark systems range from 0.03 to 0.17 pKa units, and those of three titratable groups of snake cardiotoxin range from 0.2 to 1.6 pKa units. This study demonstrates that EDS-HREX is a potent theoretical framework, which gives the correct description of multiple protonation states and good calculated pKa values. PMID:25061443

The chemical and dynamical influence of the anti-viral drug amantadine on the M2 proton channel transmembrane domain.

PubMed

Hu, Jun; Fu, Riqiang; Cross, Timothy A

2007-07-01

The M(2) proton channel plays a vital role in the life cycle of the influenza A virus. His(37), the key residue in the M(2) transmembrane domain (M(2)-TMD), plays a central role in the proton conductance mechanism. The anti-influenza drug, amantadine, inhibits the channel activity through binding to the pore of the M(2) channel. The nuclear spin relaxation data and polarization inversion spin exchange at the magic angle spectra show that both the polypeptide backbone and His(37) side chain are more constrained in the presence of amantadine. Using (15)N cross polarization magic-angle spinning NMR spectroscopy, the protonation of His(37) of M(2)-TMD in lipid bilayers was monitored in the absence and presence of amantadine as a function of pH. Binding amantadine lowers the His(37) pK(a) values by approximately three orders of magnitude compared with the first pK(a) of histidine in amantadine-free M(2)-TMD. Amantadine's influence on the His(37) chemical properties suggests a novel mechanism by which amantadine may inhibit proton conductance.

Proton-coupled electron transfer in the catalytic cycle of Alcaligenes xylosoxidans copper-dependent nitrite reductase.

PubMed

Leferink, Nicole G H; Han, Cong; Antonyuk, Svetlana V; Heyes, Derren J; Rigby, Stephen E J; Hough, Michael A; Eady, Robert R; Scrutton, Nigel S; Hasnain, S Samar

2011-05-17

We demonstrated recently that two protons are involved in reduction of nitrite to nitric oxide through a proton-coupled electron transfer (ET) reaction catalyzed by the blue Cu-dependent nitrite reductase (Cu NiR) of Alcaligenes xylosoxidans (AxNiR). Here, the functionality of two putative proton channels, one involving Asn90 and the other His254, is studied using single (N90S, H254F) and double (N90S--H254F) mutants. All mutants studied are active, indicating that protons are still able to reach the active site. The H254F mutation has no effect on the catalytic activity, while the N90S mutation results in ~70% decrease in activity. Laser flash-photolysis experiments show that in H254F and wild-type enzyme electrons enter at the level of the T1Cu and then redistribute between the two Cu sites. Complete ET from T1Cu to T2Cu occurs only when nitrite binds at the T2Cu site. This indicates that substrate binding to T2Cu promotes ET from T1Cu, suggesting that the enzyme operates an ordered mechanism. In fact, in the N90S and N90S--H254F variants, where the T1Cu site redox potential is elevated by ∼60 mV, inter-Cu ET is only observed in the presence of nitrite. From these results it is evident that the Asn90 channel is the main proton channel in AxNiR, though protons can still reach the active site if this channel is disrupted. Crystallographic structures provide a clear structural rationale for these observations, including restoration of the proton delivery via a significant movement of the loop connecting the T1Cu ligands Cys130 and His139 that occurs on binding of nitrite. Notably, a role for this loop in facilitating interaction of cytochrome c(551) with Cu NiR has been suggested previously based on a crystal structure of the binary complex.

Microwave measurements of proton tunneling and structural parameters for the propiolic acid-formic acid dimer.

PubMed

Daly, Adam M; Douglass, Kevin O; Sarkozy, Laszlo C; Neill, Justin L; Muckle, Matt T; Zaleski, Daniel P; Pate, Brooks H; Kukolich, Stephen G

2011-10-21

Microwave spectra of the propiolic acid-formic acid doubly hydrogen bonded complex were measured in the 1 GHz to 21 GHz range using four different Fourier transform spectrometers. Rotational spectra for seven isotopologues were obtained. For the parent isotopologue, a total of 138 a-dipole transitions and 28 b-dipole transitions were measured for which the a-dipole transitions exhibited splittings of a few MHz into pairs of lines and the b-type dipole transitions were split by ~580 MHz. The transitions assigned to this complex were fit to obtain rotational and distortion constants for both tunneling levels: A(0+) = 6005.289(8), B(0+) = 930.553(8), C(0+) = 803.9948(6) MHz, Δ(0+)(J) = 0.075(1), Δ(0+)(JK) = 0.71(1), and δ(0+)(j) = -0.010(1) kHz and A(0-) = 6005.275(8), B(0-) = 930.546(8), C(0-) = 803.9907(5) MHz, Δ(0-)(J) = 0.076(1), Δ(0-)(JK) = 0.70(2), and δ(0-)(j) = -0.008(1) kHz. Double resonance experiments were used on some transitions to verify assignments and to obtain splittings for cases when the b-dipole transitions were difficult to measure. The experimental difference in energy between the two tunneling states is 291.428(5) MHz for proton-proton exchange and 3.35(2) MHz for the deuterium-deuterium exchange. The vibration-rotation coupling constant between the two levels, F(ab), is 120.7(2) MHz for the proton-proton exchange. With one deuterium atom substituted in either of the hydrogen-bonding protons, the tunneling splittings were not observed for a-dipole transitions, supporting the assignment of the splitting to the concerted proton tunneling motion. The spectra were obtained using three Flygare-Balle type spectrometers and one chirped-pulse machine at the University of Virginia. Rotational constants and centrifugal distortion constants were obtained for HCOOH···HOOCCCH, H(13)COOH···HOOCCCH, HCOOD···HOOCCCH, HCOOH···DOOCCCH, HCOOD···DOOCCCH, DCOOH···HOOCCCH, and DCOOD···HOOCCCH. High-level ab initio calculations provided initial rotational constants for the complex, structural parameters, and some details of the proton tunneling potential energy surface. A least squares fit to the isotopic data reveals a planar structure that is slightly asymmetric in the OH distances. The formic OH···O propiolic hydrogen bond length is 1.8 Å and the propiolic OH···O formic hydrogen bond length is 1.6 Å, for the equilibrium configuration. The magnitude of the dipole moment was experimentally determined to be 1.95(3) × 10(-30) C m (0.584(8) D) for the 0(+) states and 1.92(5) × 10(-30) C m (0.576(14) D) for the 0(-) states. © 2011 American Institute of Physics

Testing Electrostatic Complementarity in Enzyme Catalysis: Hydrogen Bonding in the Ketosteroid Isomerase Oxyanion Hole

PubMed Central

Kraut, Daniel A; Sigala, Paul A; Pybus, Brandon; Liu, Corey W; Ringe, Dagmar; Petsko, Gregory A

2006-01-01

A longstanding proposal in enzymology is that enzymes are electrostatically and geometrically complementary to the transition states of the reactions they catalyze and that this complementarity contributes to catalysis. Experimental evaluation of this contribution, however, has been difficult. We have systematically dissected the potential contribution to catalysis from electrostatic complementarity in ketosteroid isomerase. Phenolates, analogs of the transition state and reaction intermediate, bind and accept two hydrogen bonds in an active site oxyanion hole. The binding of substituted phenolates of constant molecular shape but increasing p K a models the charge accumulation in the oxyanion hole during the enzymatic reaction. As charge localization increases, the NMR chemical shifts of protons involved in oxyanion hole hydrogen bonds increase by 0.50–0.76 ppm/p K a unit, suggesting a bond shortening of ˜0.02 Å/p K a unit. Nevertheless, there is little change in binding affinity across a series of substituted phenolates (ΔΔG = −0.2 kcal/mol/p K a unit). The small effect of increased charge localization on affinity occurs despite the shortening of the hydrogen bonds and a large favorable change in binding enthalpy (ΔΔH = −2.0 kcal/mol/p K a unit). This shallow dependence of binding affinity suggests that electrostatic complementarity in the oxyanion hole makes at most a modest contribution to catalysis of ˜300-fold. We propose that geometrical complementarity between the oxyanion hole hydrogen-bond donors and the transition state oxyanion provides a significant catalytic contribution, and suggest that KSI, like other enzymes, achieves its catalytic prowess through a combination of modest contributions from several mechanisms rather than from a single dominant contribution. PMID:16602823

A Comparative Study of the Application of Fluorescence Excitation-Emission Matrices Combined with Parallel Factor Analysis and Nonnegative Matrix Factorization in the Analysis of Zn Complexation by Humic Acids

PubMed Central

Boguta, Patrycja; Pieczywek, Piotr M.; Sokołowska, Zofia

2016-01-01

The main aim of this study was the application of excitation-emission fluorescence matrices (EEMs) combined with two decomposition methods: parallel factor analysis (PARAFAC) and nonnegative matrix factorization (NMF) to study the interaction mechanisms between humic acids (HAs) and Zn(II) over a wide concentration range (0–50 mg·dm−3). The influence of HA properties on Zn(II) complexation was also investigated. Stability constants, quenching degree and complexation capacity were estimated for binding sites found in raw EEM, EEM-PARAFAC and EEM-NMF data using mathematical models. A combination of EEM fluorescence analysis with one of the proposed decomposition methods enabled separation of overlapping binding sites and yielded more accurate calculations of the binding parameters. PARAFAC and NMF processing allowed finding binding sites invisible in a few raw EEM datasets as well as finding totally new maxima attributed to structures of the lowest humification. Decomposed data showed an increase in Zn complexation with an increase in humification, aromaticity and molecular weight of HAs. EEM-PARAFAC analysis also revealed that the most stable compounds were formed by structures containing the highest amounts of nitrogen. The content of oxygen-functional groups did not influence the binding parameters, mainly due to fact of higher competition of metal cation with protons. EEM spectra coupled with NMF and especially PARAFAC processing gave more adequate assessments of interactions as compared to raw EEM data and should be especially recommended for modeling of complexation processes where the fluorescence intensities (FI) changes are weak or where the processes are interfered with by the presence of other fluorophores. PMID:27782078

Proton transfer from imidazole to chloranil studied by FTIR spectroscopy

NASA Astrophysics Data System (ADS)

Sharma, Amit

2018-05-01

Imidazole is incorporated into many important biological molecules. The most obvious is the amino acid histidine, which has an imidazole side chain. Histidine is present in many proteins and enzymes and plays a vital part in the structure and binding functions of hemoglobin. Therefore it is important to study its proton transfer property. In the present work proton transfer from imidazole to chloranil is investigated by Fourier Transform Infra red Spectroscopy.

Inclusion of a variable RBE into proton and photon plan comparison for various fractionation schedules in prostate radiation therapy.

PubMed

Ödén, Jakob; Eriksson, Kjell; Toma-Dasu, Iuliana

2017-03-01

A constant relative biological effectiveness (RBE) of 1.1 is currently used in proton radiation therapy to account for the increased biological effectiveness compared to photon therapy. However, there is increasing evidence that proton RBE vary with the linear energy transfer (LET), the dose per fraction, and the type of the tissue. Therefore, this study aims to evaluate the impact of disregarding variations in RBE when comparing proton and photon dose plans for prostate treatments for various fractionation schedules using published RBE models and several α/β assumptions. Photon and proton dose plans were created for three generic prostate cancer cases. Three BED 3Gy equivalent schedules were studied, 78, 57.2, and 42.8 Gy in 39, 15, and 7 fractions, respectively. The proton plans were optimized assuming a constant RBE of 1.1. By using the Monte Carlo calculated dose-averaged LET (LET d ) distribution and assuming α/β values on voxel level, three variable RBE models were applied to the proton dose plans. The impact of the variable RBE was studied in the plan comparison, which was based on the dose distribution, DVHs, and normal tissue complication probabilities (NTCP) for the rectum. Subsequently, the physical proton dose was reoptimized for each proton plan based on the LET d distribution, to achieve a homogeneous RBE-weighted target dose when applying a specific RBE model and still fulfill the clinical goals for the rectum and bladder. All the photon and proton plans assuming RBE = 1.1 met the clinical goals with similar target coverage. The proton plans fulfilled the robustness criteria in terms of range and setup uncertainty. Applying the variable RBE models generally resulted in higher target doses and rectum NTCP compared to the photon plans. The increase was most pronounced for the fractionation dose of 2 Gy(RBE), whereas it was of less magnitude and more dependent on model and α/β assumption for the hypofractionated schedules. The reoptimized proton plans proved to be robust and showed similar target coverage and doses to the organs at risk as the proton plans optimized with a constant RBE. Model predicted RBE values may differ substantially from 1.1. This is most pronounced for fractionation doses of around 2 Gy(RBE) with higher doses to the target and the OARs, whereas the effect seems to be of less importance for the hypofractionated schedules. This could result in misleading conclusions when comparing proton plans to photon plans. By accounting for a variable RBE in the optimization process, robust and clinically acceptable dose plans, with the potential of lowering rectal NTCP, may be generated by reoptimizing the physical dose. However, the direction and magnitude of the changes in the physical proton dose to the prostate are dependent on RBE model and α/β assumptions and should therefore be used conservatively. © 2017 American Association of Physicists in Medicine.

Calorimetry of heterogeneous systems: H+ binding to TiO2 in NaCl

USGS Publications Warehouse

Mehr, S.R.; Eatough, D.J.; Hansen, L.D.; Lewis, E.A.; Davis, J.A.

1989-01-01

A simultaneous calorimetric and potentiometric technique has been developed for measuring the thermodynamics of proton binding to mineral oxides in the presence of a supporting electrolyte. Modifications made to a commercial titration calorimeter to add a combination pH electrode and maintain an inert atmosphere in the calorimeter reaction vessel are described. A procedure to calibrate potentiometric measurements in heterogeneous systems to correct for the suspension effect on pH is given. The enthalpy change for proton dissociation from TiO2 in aqueous suspension as a function of pH is reported for 0.01, 0.1, and 0.5 M NaCl. The enthalpy change for proton dissociation is endothermic, ranging from 10.5 ?? 3.8 to 45.0 ?? 3.8 kJ mol-1 over the pH range from 4 to 10. ?? 1989.

Rotational spectroscopic study of carbonyl sulfide solvated with hydrogen molecules.

PubMed

Michaud, Julie M; Jäger, Wolfgang

2008-10-14

Rotational spectra of small-sized (H(2))(N)-OCS clusters with N = 2-7 were measured using a pulsed-jet Fourier transform microwave spectrometer. These include spectra of pure (para-H(2))(N)-OCS clusters, pure (ortho-H(2))(N)-OCS clusters, and mixed ortho-H(2) and para-H(2) containing clusters. The rotational lines of ortho-H(2) molecules containing clusters show proton spin-proton spin hyperfine structure, and the pattern evolves as the number of ortho-H(2) molecules in the cluster increases. Various isotopologues of the clusters were investigated, including those with O(13)CS, OC(33)S, OC(34)S, and O(13)C(34)S. Nuclear quadrupole hyperfine structures of rotational transitions were observed for (33)S (nuclear spin quantum number I = 3/2) containing isotopologues. The (33)S nuclear quadrupole coupling constants are compared to the corresponding constant of the OCS monomer and those of the He(N)-OCS clusters. The assignment of the number of solvating hydrogen molecules N is supported by the analyses of the proton spin-proton spin hyperfine structures of the mixed clusters, the dependence of line intensities on sample conditions (pressure and concentrations), and the agreement of the (para-H(2))(N)-OCS and (ortho-H(2))(N)-OCS rotational constants with those from a previous infrared study [J. Tang and A. R. W. McKellar, J. Chem. Phys. 121, 3087 (2004)].

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.

A Conserved Steroid Binding Site in Cytochrome c Oxidase

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

Qin, Ling; Mills, Denise A.; Buhrow, Leann

2010-09-02

Micromolar concentrations of the bile salt deoxycholate are shown to rescue the activity of an inactive mutant, E101A, in the K proton pathway of Rhodobacter sphaeroides cytochrome c oxidase. A crystal structure of the wild-type enzyme reveals, as predicted, deoxycholate bound with its carboxyl group at the entrance of the K path. Since cholate is a known potent inhibitor of bovine oxidase and is seen in a similar position in the bovine structure, the crystallographically defined, conserved steroid binding site could reveal a regulatory site for steroids or structurally related molecules that act on the essential K proton path.

Clarithromycin and Tetracycline Binding to Soil Humic Acid in the Absence and Presence of Calcium.

PubMed

Christl, Iso; Ruiz, Mercedes; Schmidt, J R; Pedersen, Joel A

2016-09-20

Numerous ionizable organic micropollutants contain positively charged moieties at pH values typical of environmental systems. Describing organic cation and zwitterion interaction with dissolved natural organic matter requires explicit consideration of the pH-dependent speciation of both sorbate and sorbent. We studied the pH-, ionic strength-, and concentration-dependent binding of relatively large, organic cations and zwitterions (viz., the antibiotics clarithromycin and tetracycline) to dissolved humic acid in the absence and presence of Ca(2+) and evaluated the ability of the NICA-Donnan model to describe the data. Clarithromycin interaction with dissolved humic acid was well described by the model including the competitive effect of Ca(2+) on clarithromycin binding over a wide range of solution conditions by considering only the binding of the cationic species to low proton-affinity sites in humic acid. Tetracycline possesses multiple ionizable moieties and forms complexes with Ca(2+). An excellent fit to experimental data was achieved by considering tetracycline cation interaction with both low and high proton-affinity sites of humic acid and zwitterion interaction with high proton-affinity sites. In contrast to clarithromycin, tetracycline binding to humic acid increased in the presence of Ca(2+), especially under alkaline conditions. Model calculations indicate that this increase is due to electrostatic interaction of positively charged tetracycline-Ca complexes with humic acid rather than due to the formation of ternary complexes, except at very low TC concentrations.

Investigation of flavonoids bearing different substituents on ring C and their cu2+ complex binding with bovine serum albumin: structure-affinity relationship aspects.

PubMed

Shi, Shuyun; Zhang, Yuping; Chen, Xiaoqin; Peng, Mijun

2011-10-12

The effects of 1:1 flavonoid-Cu(2+) complexes of four flavonoids with different C-ring substituents, quercetin (QU), luteolin (LU), taxifolin (TA), and (+)-catechin (CA), on bovine serum albumin (BSA) were investigated and compared with corresponding free flavonoids by spectroscopic analysis in an attempt to characterize the chemical association taking place. The results indicated that all of the quenching mechanisms were based on static quenching combined with nonradiative energy transfer. Cu(2+) chelation changed the binding constants for BSA depending on the structures of flavonoids and the detected concentrations. The reduced hydroxyl groups, increased steric hindrance, and hydrophilicity of Cu(2+) chelation may be the main reasons for the reduced binding constants, whereas the formation of stable flavonoid-Cu(2+) complexes and synergistic action could increase the binding constants. The changed trends of critical energy transfer distance (R(0)) for Cu(2+) chelation were contrary to those of binding constants.

Substitutions of S101 decrease proton and hydride transfers in the oxidation of betaine aldehyde by choline oxidase.

PubMed

Gadda, Giovanni; Yuan, Hongling

2017-11-15

Choline oxidase oxidizes choline to glycine betaine, with two flavin-mediated reactions to convert the alcohol substrate to the carbon acid product. Proton abstraction from choline or hydrated betaine aldehyde in the wild-type enzyme occurs in the mixing time of the stopped-flow spectrophotometer, thereby precluding a mechanistic investigation. Mutagenesis of S101 rendered the proton transfer reaction amenable to study. Here, we have investigated the aldehyde oxidation reaction catalyzed by the mutant enzymes using steady-state and rapid kinetics with betaine aldehyde. Stopped-flow traces for the reductive half-reaction of the S101T/V/C variants were biphasic, corresponding to the reactions of proton abstraction and hydride transfer. In contrast, the S101A enzyme yielded monophasic traces like wild-type choline oxidase. The rate constants for proton transfer in the S101T/C/V variants decreased logarithmically with increasing hydrophobicity of residue 101, indicating a behavior different from that seen previously with choline for which no correlation was determined. The rate constants for hydride transfer also showed a logarithmic decrease with increasing hydrophobicity at position 101, which was similar to previous results with choline as a substrate for the enzyme. Thus, the hydrophilic character of S101 is necessary not only for efficient hydride transfer but also for the proton abstraction reaction. Copyright © 2017. Published by Elsevier Inc.

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

Spectrophotometric study on the proton transfer reaction between 2-amino-4-methylpyridine with 2,6-dichloro-4-nitrophenol in methanol, acetonitrile and the binary mixture 50% methanol + 50% acetonitrile

NASA Astrophysics Data System (ADS)

Al-Ahmary, Khairia M.; Habeeb, Moustafa M.; Al-Obidan, Areej H.

2016-02-01

Proton transfer reaction between 2-amino-4-methylpyridine (2AMP) as the proton acceptor with 2,6-dichloro-4-nitrophenol (DCNP) as the proton donor has been investigated spectrophotometrically in methanol (MeOH), acetonitrile (AN) and a binary mixture composed of 50% MeOH and 50% AN (AN-Me). The composition of the complex has been investigated utilizing Job's and photometric titration methods to be 1:1. Minimum-maximum absorbance equation has been applied to estimate the formation constant of the proton transfer reaction (KPT) where it reached high values in the investigated solvent confirming its high stability. The formation constant recorded higher value in AN compared with MeOH and mixture of AN-Me. Based on the formation of stable proton transfer complex, a sensitive spectrophotometric method was suggested for quantitative determination of 2AMP. The Lambert-Beer's law was obeyed in the concentration range 0.5-8 μg mL- 1 with small values of limits of detection and quantification. The solid complex between 2AMP with DCNP has been synthesized and characterized by elemental analysis to be 1:1 in concordant with the molecular stoichiometry in solution. Further analysis of the solid complex was carried out using infrared and 1H NMR spectroscopy.

1H NMR studies of the 5-(hydroxymethyl)-2'-deoxyuridine containing TF1 binding site.

PubMed

Pasternack, L B; Bramham, J; Mayol, L; Galeone, A; Jia, X; Kearns, D R

1996-07-15

The pyrimidine base 5-(hydroxymethyl)-2'-deoxyuridine (HmU) is a common nucleotide in SPO1 phage DNA. Numerous transcriptional proteins bind HmU-containing DNA preferentially implicating a regulatory function of HmU. We have investigated the conformation and dynamics of d-(5'-CHmUCHmUACACGHmUGHmUAGAG-OH-3')2 (HmU-DNA). This oligonucleotide mimics the consensus sequence of Transcription Factor 1 (TF1). The HmU-DNA was compared to the thymine-containing oligonucleotide. NOESY and DQF COSY spectroscopy provided resonance assignments of nonexchangeable and exchangeable protons, intranucleotide, internucleotide and intrastrand proton-proton distances, and dihedral angle constraints. Methylene protons of the hydroxymethyl group are nonequivalent protons and the hydroxymethyl group is not freely rotating. The hydroxymethyl group adopts a specific orientation with the OH group oriented on the 3' side of the plane of the base. Analysis of imino proton resonances and NOEs indicates additional end base pair fraying and a temperature-induced transition to a conformation in which the internal HmU-A base pairs are disrupted or have reduced lifetimes. Orientation of the hydroxymethyl group indicates the presence of internucleotide intrastrand hydrogen bonding between the HmU12C5 hydroxyl group and A13. All sugars in both DNAs show a C2'endo conformation (typical of B-DNA).

Calcium ion binding to a soil fulvic acid using a donnan potential model

USGS Publications Warehouse

Marinsky, J.A.; Mathuthu, A.; Ephraim, J.H.; Reddy, M.M.

1999-01-01

Calcium ion binding to a soil fulvic acid (Armadale Bh Horizon) was evaluated over a range of calcium ion concentrations, from pH 3.8 to 7.3, using potentiometric titrations and calcium ion electrode measurements. Fulvic acid concentration was constant (100 milligrams per liter) and calcium ion concentration varied up to 8 X 10-4 moles per liter. Experiments discussed here included: (1) titrations of fulvic acid-calcium ion containing solutions with sodium hydroxide; and (2) titrations of fully neutralized fulvic acid with calcium chloride solutions. Apparent binding constants (expressed as the logarithm of the value, log ??app) vary with solution pH, calcium ion concentration, degree of acid dissociation, and ionic strength (from log ??app = 2.5 to 3.9) and are similar to those reported by others. Fulvic acid charge, and the associated Donnan Potential, influences calcium ion-fulvic acid ion pair formation. A Donnan Potential corrrection term allowed calculation of intrinsic calcium ion-fulvic acid binding constants. Intrinsic binding constants vary from 1.2 to 2.5 (the average value is about log??= 1.6) and are similar to, but somewhat higher than, stability constants for calcium ion-carboxylic acid monodentate complexes. ?? by Oldenbourg Wissenschaftsverlag, Mu??nchen.

Cation Binding to Xanthorhodopsin: Electron Paramagnetic Resonance and Magnetic Studies.

PubMed

Smolensky Koganov, Elena; Leitus, Gregory; Rozin, Rinat; Weiner, Lev; Friedman, Noga; Sheves, Mordechai

2017-05-04

Xanthorhodopsin (xR) is a member of the retinal protein family and acts as a proton pump in the cell membranes of the extremely halophilic eubacterium Salinibacter ruber. In addition to the retinal chromophore, xR contains a carotenoid, which acts as a light-harvesting antenna as it transfers 40% of the quanta it absorbs to the retinal. Our previous studies have shown that the CD and absorption spectra of xR are dramatically affected due to the protonation of two different residues. It is still unclear whether xR can bind cations. Electron paramagnetic resonance (EPR) spectroscopy used in the present study revealed that xR can bind divalent cations, such as Mn 2+ and Ca 2+ , to deionized xR (DI-xR). We also demonstrate that xR can bind 1 equiv of Mn 2+ to a high-affinity binding site followed by binding of ∼40 equiv in cooperative manner and ∼100 equiv of Mn 2+ that are weakly bound. SQUID magnetic studies suggest that the high cooperative binding of Mn 2+ cations to xR is due to the formation of Mn 2+ clusters. Our data demonstrate that Ca 2+ cations bind to DI-xR with a lower affinity than Mn 2+ , supporting the assumption that binding of Mn 2+ occurs through cluster formation, because Ca 2+ cations cannot form clusters in contrast to Mn 2+ .

Acid-base properties of humic and fulvic acids formed during composting.

PubMed

Plaza, César; Senesi, Nicola; Polo, Alfredo; Brunetti, Gennaro

2005-09-15

The soil acid-base buffering capacity and the biological availability, mobilization, and transport of macro- and micronutrients, toxic metal ions, and xenobiotic organic cations in soil are strongly influenced by the acid-base properties of humic substances, of which humic and fulvic acids are the major fractions. For these reasons, the proton binding behavior of the humic acid-like (HA) and fulvic acid-like (FA) fractions contained in a compost are believed to be instrumental in its successful performance in soil. In this work, the acid-base properties of the HAs and FAs isolated from a mixture of the sludge residue obtained from olive oil mill wastewater (OMW) evaporated in an open-air pond and tree cuttings (TC) at different stages of composting were investigated by a current potentiometric titration method and the nonideal competitive adsorption (NICA)-Donnan model. The NICA-Donnan model provided an excellent description of the acid-base titration data, and pointed out substantial differences in site density and proton-binding affinity between the HAs and FAs examined. With respect to FAs, HAs were characterized by a smaller content of carboxylic- and phenolic-type groups and their larger affinities for proton binding. Further, HAs featured a greater heterogeneity in carboxylic-type groups than FAs. The composting process increased the content and decreased the proton affinity of carboxylic- and phenolic-type groups of HAs and FAs, and increased the heterogeneity of phenolic-type groups of HAs. As a whole, these effects indicated that the composting process could produce HA and FA fractions with greater cation binding capacities. These results suggest that composting of organic materials improves their agronomic and environmental value by increasing their potential to retain and exchange macro- and micronutrients, and to reduce the bioavailability of organic and inorganic pollutants.

The role of the AT pairs in the acid denaturation of DNA.

PubMed Central

Hermann, P; Fredericq, E

1977-01-01

It has been determined previously that the protonation of the GC pairs induces a DNA conformation change which leads to a "metastable" structure. The role of the AT pairs, however, is no well known because the protonation does not modify their spectral properties. By means of an indirect method based on the binding of proflavine, it has been determined that the AT pairs are protonated before the acid-induced denaturation and that they seem to be unable to assume a conformation change when protonated. These results would indicate that the protonated AT pairs may be responsible for the induction of the acid denaturation and not the GC pairs as it was thought previously. PMID:20604

Electronic wave function and binding effects in M-shell ionization of gold by protons

NASA Astrophysics Data System (ADS)

Pajek, M.; Banaś, D.; Jabłoński, Ł.; Mukoyama, T.

2018-02-01

The measured M-X-ray production cross sections for protons, which are used in the particle induced X-ray emission (PIXE) technique, are systematically underestimated for low impact energies by the ECPSSR and ECUSAR theories. These theories, which are based on the plane wave Born approximation (PWBA) and use the screened hydrogenic wave functions, include corrections for the projectile Coulomb deflection and electron relativistic and binding effects. In the present paper, in order to interpret the observed disagreement at low impact energies, the systematic calculations of the M-shell ionization cross sections for gold were performed using the semiclassical (SCA) and the binary encounter (BEA) approximations in order to identify a role of the electronic wave function and electron binding effects. In these calculations the different wave functions, from nonrelativistic hydrogenic to selfconsistent Dirac-Hartree-Fock, were considered and the binding effect was treated within extreme separated- (SA) and united-atoms (UA) limits. The results are discussed in details and the observed discrepancies are attributed to inadequate description of the electron binding effect at the lowest impact energies for which the molecular approach is required.

The selectivity of the Na+/K+-pump is controlled by binding site protonation and self-correcting occlusion

PubMed Central

Rui, Huan; Artigas, Pablo; Roux, Benoît

2016-01-01

The Na+/K+-pump maintains the physiological K+ and Na+ electrochemical gradients across the cell membrane. It operates via an 'alternating-access' mechanism, making iterative transitions between inward-facing (E1) and outward-facing (E2) conformations. Although the general features of the transport cycle are known, the detailed physicochemical factors governing the binding site selectivity remain mysterious. Free energy molecular dynamics simulations show that the ion binding sites switch their binding specificity in E1 and E2. This is accompanied by small structural arrangements and changes in protonation states of the coordinating residues. Additional computations on structural models of the intermediate states along the conformational transition pathway reveal that the free energy barrier toward the occlusion step is considerably increased when the wrong type of ion is loaded into the binding pocket, prohibiting the pump cycle from proceeding forward. This self-correcting mechanism strengthens the overall transport selectivity and protects the stoichiometry of the pump cycle. DOI: http://dx.doi.org/10.7554/eLife.16616.001 PMID:27490484

Ab initio estimates of the size of the observable universe

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

Page, Don N., E-mail: profdonpage@gmail.com

2011-09-01

When one combines multiverse predictions by Bousso, Hall, and Nomura for the observed age and size of the universe in terms of the proton and electron charge and masses with anthropic predictions of Carter, Carr, and Rees for these masses in terms of the charge, one gets that the age of the universe should be roughly the inverse 64th power, and the cosmological constant should be around the 128th power, of the proton charge. Combining these with a further renormalization group argument gives a single approximate equation for the proton charge, with no continuous adjustable or observed parameters, and withmore » a solution that is within 8% of the observed value. Using this solution gives large logarithms for the age and size of the universe and for the cosmological constant that agree with the observed values within 17%.« less

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

Unveiling N-protonation and anion-binding effects on Fe/N/C-catalysts for O2 reduction in PEM fuel cells

PubMed Central

Herranz, Juan; Jaouen, Frédéric; Lefèvre, Michel; Kramm, Ulrike I.; Proietti, Eric; Dodelet, Jean-Pol; Bogdanoff, Peter; Fiechter, Sebastian; Abs-Wurmbach, Irmgard; Bertrand, Patrick; Arruda, Thomas M.; Mukerjee, Sanjeev

2013-01-01

The high cost of proton-exchange-membrane fuel cells would be considerably reduced if platinumbased catalysts were replaced by iron-based substitutes, which have recently demonstrated comparable activity for oxygen reduction, but whose cause of activity decay in acidic medium has been elusive. Here, we reveal that the activity of Fe/N/C-catalysts prepared through a pyrolysis in NH3 is mostly imparted by acid-resistant FeN4-sites whose turnover frequency for the O2 reduction can be regulated by fine chemical changes of the catalyst surface. We show that surface N-groups protonate at pH 1 and subsequently bind anions. This results in decreased activity for the O2 reduction. The anions can be removed chemically or thermally, which restores the activity of acid-resistant FeN4-sites. These results are interpreted as an increased turnover frequency of FeN4-sites when specific surface N-groups protonate. These unprecedented findings provide new perspective for stabilizing the most active Fe/N/C-catalysts known to date. PMID:24179561

Unveiling N-protonation and anion-binding effects on Fe/N/C-catalysts for O2 reduction in PEM fuel cells.

PubMed

Herranz, Juan; Jaouen, Frédéric; Lefèvre, Michel; Kramm, Ulrike I; Proietti, Eric; Dodelet, Jean-Pol; Bogdanoff, Peter; Fiechter, Sebastian; Abs-Wurmbach, Irmgard; Bertrand, Patrick; Arruda, Thomas M; Mukerjee, Sanjeev

2011-11-18

The high cost of proton-exchange-membrane fuel cells would be considerably reduced if platinumbased catalysts were replaced by iron-based substitutes, which have recently demonstrated comparable activity for oxygen reduction, but whose cause of activity decay in acidic medium has been elusive. Here, we reveal that the activity of Fe/N/C-catalysts prepared through a pyrolysis in NH 3 is mostly imparted by acid-resistant FeN 4 -sites whose turnover frequency for the O 2 reduction can be regulated by fine chemical changes of the catalyst surface. We show that surface N-groups protonate at pH 1 and subsequently bind anions. This results in decreased activity for the O 2 reduction. The anions can be removed chemically or thermally, which restores the activity of acid-resistant FeN 4 -sites. These results are interpreted as an increased turnover frequency of FeN 4 -sites when specific surface N-groups protonate. These unprecedented findings provide new perspective for stabilizing the most active Fe/N/C-catalysts known to date.

Binding constant of cell adhesion receptors and substrate-immobilized ligands depends on the distribution of ligands

NASA Astrophysics Data System (ADS)

Li, Long; Hu, Jinglei; Xu, Guangkui; Song, Fan

2018-01-01

Cell-cell adhesion and the adhesion of cells to tissues and extracellular matrix, which are pivotal for immune response, tissue development, and cell locomotion, depend sensitively on the binding constant of receptor and ligand molecules anchored on the apposing surfaces. An important question remains of whether the immobilization of ligands affects the affinity of binding with cell adhesion receptors. We have investigated the adhesion of multicomponent membranes to a flat substrate coated with immobile ligands using Monte Carlo simulations of a statistical mesoscopic model with biologically relevant parameters. We find that the binding of the adhesion receptors to ligands immobilized on the substrate is strongly affected by the ligand distribution. In the case of ligand clusters, the receptor-ligand binding constant can be significantly enhanced due to the less translational entropy loss of lipid-raft domains in the model cell membranes upon the formation of additional complexes. For ligands randomly or uniformly immobilized on the substrate, the binding constant is rather decreased since the receptors localized in lipid-raft domains have to pay an energetic penalty in order to bind ligands. Our findings help to understand why cell-substrate adhesion experiments for measuring the impact of lipid rafts on the receptor-ligand interactions led to contradictory results.

Excited state proton transfer in strongly enhanced GFP (sGFP2).

PubMed

van Oort, Bart; ter Veer, Mirelle J T; Groot, Marie Louise; van Stokkum, Ivo H M

2012-07-07

Proton transfer is an elementary process in biology. Green fluorescent protein (GFP) has served as an important model system to elucidate the mechanistic details of this reaction, because in GFP proton transfer can be induced by light absorption. We have used pump-dump-probe spectroscopy to study how proton transfer through the 'proton-wire' around the chromophore is affected by a combination of mutations in a modern GFP variety (sGFP2). The results indicate that in H(2)O, after absorption of a photon, a proton is transferred (A* → I*) in 5 ps, and back-transferred from a ground state intermediate (I → A) in 0.3 ns, similar to time constants found with GFPuv, although sGFP2 shows less heterogeneous proton transfer. This suggests that the mutations left the proton-transfer largely unchanged, indicating the robustness of the proton-wire. We used pump-dump-probe spectroscopy in combination with target analysis to probe suitability of the sGFP2 fluorophore for super-resolution microscopy.

Determination of the strong coupling constant from jet rates in deep inelastic scattering

NASA Astrophysics Data System (ADS)

Ahmed, T.; Aid, S.; Andreev, V.; Andrieu, B.; Appuhn, R.-D.; Arpagaus, M.; Babaev, A.; Baehr, J.; Bán, J.; Baranov, P.; Barrelet, E.; Bartel, W.; Barth, M.; Bassler, U.; Beck, H. P.; Behrend, H.-J.; Belousov, A.; Berger, Ch.; Bergstein, H.; Bernardi, G.; Bernet, R.; Bertrand-Coremans, G.; Besançon, M.; Beyer, R.; Biddulph, P.; Bizot, J. C.; Blobel, V.; Borras, K.; Botterweck, F.; Boudry, V.; Braemer, A.; Brasse, F.; Braunschweig, W.; Brisson, V.; Bruncko, D.; Brune, C.; Buchholz, R.; Büngener, L.; Bürger, J.; Büsser, F. W.; Buniatian, A.; Burke, S.; Buschhorn, G.; Campbell, A. J.; Carli, T.; Charles, F.; Clarke, D.; Clegg, A. B.; Clerbaux, B.; Colombo, M.; Contreras, J. G.; Cormack, C.; Coughlan, J. A.; Courau, A.; Coutures, Ch.; Cozzika, G.; Criegee, L.; Cussans, D. G.; Cvach, J.; Dagoret, S.; Dainton, J. B.; Danilov, M.; Dau, W. D.; Daum, K.; David, M.; Deffur, E.; Delcourt, B.; Del Buono, L.; De Roeck, A.; De Wolf, E. A.; Di Nezza, P.; Dollfus, C.; Dowell, J. D.; Dreis, H. B.; Droutskoi, A.; Duboc, J.; Düllmann, D.; Dünger, O.; Duhm, H.; Ebert, J.; Ebert, T. R.; Eckerlin, G.; Efremenko, V.; Egli, S.; Ehrlichmann, H.; Eichenberger, S.; Eichler, R.; Eisele, F.; Eisenhandler, E.; Ellison, R. J.; Elsen, E.; Erdmann, M.; Erdmann, W.; Evrard, E.; Favart, L.; Fedotov, A.; Feeken, D.; Felst, R.; Feltesse, J.; Ferencei, J.; Ferrarotto, F.; Flamm, K.; Fleischer, M.; Flieser, M.; Flügge, G.; Fomenko, A.; Fominykh, B.; Forbush, M.; Formánek, J.; Foster, J. M.; Franke, G.; Fretwurst, E.; Gabathuler, E.; Gabathuler, K.; Gamerdinger, K.; Garvey, J.; Gayler, J.; Gebauer, M.; Gellrich, A.; Genzel, H.; Gerhards, R.; Goerlach, U.; Goerlich, L.; Gogitidze, N.; Goldberg, M.; Goldner, D.; Gonzalez-Pineiro, B.; Gorelov, I.; Goritchev, P.; Grab, C.; Grässler, H.; Grässler, R.; Greenshaw, T.; Grindhammer, G.; Gruber, A.; Gruber, C.; Haack, J.; Haidt, D.; Hajduk, L.; Hamon, O.; Hampel, M.; Hanlon, E. M.; Hapke, M.; Haynes, W. J.; Heatherington, J.; Heinzelmann, G.; Henderson, R. C. W.; Henschel, H.; Herma, R.; Herynek, I.; Hess, M. F.; Hildesheim, W.; Hill, P.; Hill, K. H.; Hilton, C. D.; Hladký, J.; Hoeger, K. C.; Höppner, M.; Horisberger, R.; Huet, Ph.; Hufnagel, H.; Ibbotson, M.; Itterbeck, H.; Jabiol, M.-A.; Jacholkowska, A.; Jacobsson, C.; Jaffre, M.; Janoth, J.; Jansen, T.; Jönsson, L.; Johannsen, K.; Johnson, D. P.; Johnson, L.; Jung, H.; Kalmus, P. I. P.; Kant, D.; Kaschowitz, R.; Kasselmann, P.; Kathage, U.; Kaufmann, H. H.; Kazarian, S.; Kenyon, I. R.; Kermiche, S.; Keuler, C.; Kiesling, C.; Klein, M.; Kleinwort, C.; Knies, G.; Ko, W.; Köhler, T.; Köhne, J. H.; Kolanoski, H.; Kole, F.; Kolya, S. D.; Korbel, V.; Korn, M.; Kostka, P.; Kotelnikov, S. K.; Krämerkämper, T.; Krasny, M. W.; Krehbiel, H.; Krücker, D.; Krüger, U.; Krüner-Marquis, U.; Kubenka, J. P.; Küster, H.; Kuhlen, M.; Kurča, T.; Kurzhöfer, J.; Kuznik, B.; Lacour, D.; Lamarche, F.; Lander, R.; Landon, M. P. J.; Lange, W.; Lanius, P.; Laporte, J.-F.; Lebedev, A.; Leverenz, C.; Levonian, S.; Ley, Ch.; Lindner, A.; Lindström, G.; Linsel, F.; Lipinski, J.; List, B.; Loch, P.; Lohmander, H.; Lopez, G. C.; Lubimov, V.; Lüke, D.; Magnussen, N.; Malinovski, E.; Mani, S.; Maraček, R.; Marage, P.; Marks, J.; Marshall, R.; Martens, J.; Martin, R.; Martyn, H.-U.; Martyniak, J.; Masson, S.; Mavroidis, T.; Maxfield, S. J.; McMahon, S. J.; Mehta, A.; Meier, K.; Mercer, D.; Merz, T.; Meyer, C. A.; Meyer, H.; Meyer, J.; Mikocki, S.; Milstead, D.; Moreau, F.; Morris, J. V.; Müller, G.; Müller, K.; Murín, P.; Nagovizin, V.; Nahnhauer, R.; Naroska, B.; Naumann, Th.; Newman, P. R.; Newton, D.; Neyret, D.; Nguyen, H. K.; Niebergall, F.; Niebuhr, C.; Nisius, R.; Nowak, G.; Noyes, G. W.; Nyberg-Werther, M.; Oakden, M.; Oberlack, H.; Obrock, U.; Olsson, J. E.; Panaro, E.; Panitch, A.; Pascaud, C.; Patel, G. D.; Peppel, E.; Perez, E.; Phillips, J. P.; Pichler, Ch.; Pitzl, D.; Pope, G.; Prell, S.; Prosi, R.; Rädel, G.; Raupach, F.; Reimer, P.; Reinshagen, S.; Ribarics, P.; Rick, H.; Riech, V.; Riedlberger, J.; Riess, S.; Rietz, M.; Robertson, S. M.; Robmann, P.; Roloff, H. E.; Roosen, R.; Rosenbauer, K.; Rostovtsev, A.; Rouse, F.; Royon, C.; Rüter, K.; Rusakov, S.; Rybicki, K.; Rylko, R.; Sahlmann, N.; Sanchez, E.; Sankey, D. P. C.; Savitsky, M.; Schacht, P.; Schiek, S.; Schleper, P.; von Schlippe, W.; Schmidt, C.; Schmidt, D.; Schmidt, G.; Schöning, A.; Schröder, V.; Schuhmann, E.; Schwab, B.; Schwind, A.; Seehausen, U.; Sefkow, F.; Seidel, M.; Sell, R.; Semenov, A.; Shekelyan, V.; Sheviakov, I.; Shooshtari, H.; Shtarkov, L. N.; Siegmon, G.; Siewert, U.; Sirois, Y.; Skillicorn, I. O.; Smirnov, P.; Smith, J. R.; Soloviev, Y.; Spitzer, H.; Starosta, R.; Steenbock, M.; Steffen, P.; Steinberg, R.; Stella, B.; Stephens, K.; Stier, J.; Stiewe, J.; Stösslein, U.; Strachota, J.; Straumann, U.; Struczinski, W.; Sutton, J. P.; Tapprogge, S.; Taylor, R. E.; Tchernyshov, V.; Thiebaux, C.; Thompson, G.; Truöl, P.; Turnau, J.; Tutas, J.; Uelkes, P.; Usik, A.; Valkár, S.; Valkárová, A.; Vallée, C.; Van Esch, P.; Van Mechelen, P.; Vartapetian, A.; Vazdik, Y.; Vecko, M.; Verrecchia, P.; Villet, G.; Wacker, K.; Wagener, A.; Wagener, M.; Walker, I. W.; Walther, A.; Weber, G.; Weber, M.; Wegener, D.; Wegner, A.; Wellisch, H. P.; West, L. R.; Willard, S.; Winde, M.; Winter, G.-G.; Wright, A. E.; Wünsch, E.; Wulff, N.; Yiou, T. P.; Žáček, J.; Zarbock, D.; Zhang, Z.; Zhokin, A.; Zimmer, M.; Zimmermann, W.; Zomer, F.; Zuber, K.; H1 Collaboration

1995-02-01

Jet rates in deep inelastic electron proton scattering are studied with the H1 detector at HERA for momentum transfers squared between 10 and 4000 GeV 2. It is shown that they can be quantitatively described by perturbative QCD in next to leading order making use of the parton densities of the proton and with the strong coupling constant αs as a free parameter. The measured value, αs( MZ2) = 0.123 ± 0.018, is in agreement both with determinations from e+e- annihilation at LEP using the same observable and with the world average.

QED is not endangered by the proton's size

NASA Astrophysics Data System (ADS)

De Rújula, A.

2010-10-01

Pohl et al. have reported a very precise measurement of the Lamb-shift in muonic hydrogen (Pohl et al., 2010) [1], from which they infer the radius characterizing the proton's charge distribution. The result is 5 standard deviations away from the one of the CODATA compilation of physical constants. This has been interpreted (Pohl et al., 2010) [1] as possibly requiring a 4.9 standard-deviation modification of the Rydberg constant, to a new value that would be precise to 3.3 parts in 1013, as well as putative evidence for physics beyond the standard model (Flowers, 2010) [2]. I demonstrate that these options are unsubstantiated.

Constraints on parton distribution functions and extraction of the strong coupling constant from the inclusive jet cross section in pp collisions at $$\\sqrt{s} = 7$$ $$\\,\\text {TeV}$$

DOE PAGES

Khachatryan, Vardan

2015-06-26

The inclusive jet cross section for proton–proton collisions at a centre-of-mass energy of 7TeVwas measured by the CMS Collaboration at the LHC with data corresponding to an integrated luminosity of 5.0fb -1. The measurement covers a phase space up to 2TeV in jet transverse momentum and 2.5 in absolute jet rapidity. The statistical precision of these data leads to stringent constraints on the parton distribution functions of the proton. The data provide important input for the gluon density at high fractions of the proton momentum and for the strong coupling constant at large energy scales. Using predictions from perturbative quantummore » chromodynamics at next-to-leading order, complemented with electroweak corrections, the constraining power of these data is investigated and the strong coupling constant at the Z boson mass M Z is determined to be α S(M Z)=0.1185±0.0019(exp) +0.0060 -0.0037(theo), which is in agreement with the world average.« less

Zinc chelation with hydroxamate in histone deacetylases modulated by water access to the linker binding channel.

PubMed

Wu, Ruibo; Lu, Zhenyu; Cao, Zexing; Zhang, Yingkai

2011-04-27

It is of significant biological interest and medical importance to develop class- and isoform-selective histone deacetylase (HDAC) modulators. The impact of the linker component on HDAC inhibition specificity has been revealed but is not understood. Using Born-Oppenheimer ab initio QM/MM MD simulations, a state-of-the-art approach to simulating metallo-enzymes, we have found that the hydroxamic acid remains to be protonated upon its binding to HDAC8, and thus disproved the mechanistic hypothesis that the distinct zinc-hydroxamate chelation modes between two HDAC subclasses come from different protonation states of the hydroxamic acid. Instead, our simulations suggest a novel mechanism in which the chelation mode of hydroxamate with the zinc ion in HDACs is modulated by water access to the linker binding channel. This new insight into the interplay between the linker binding and the zinc chelation emphasizes its importance and gives guidance regarding linker design for the development of new class-IIa-specific HDAC inhibitors.

Modeling the protonation states of β-secretase binding pocket by molecular dynamics simulations and docking studies.

PubMed

Sabbah, Dima A; Zhong, Haizhen A

2016-07-01

β-secretase (BACE1) is an aspartyl protease that processes the β-amyloid peptide in the human brain in patients with Alzheimer's disease. There are two catalytic aspartates (ASP32 and ASP228) in the active domain of BACE1. Although it is believed that the net charge of the Asp dyad is -1, the exact protonation state still remains a matter of debate. We carried out molecular dynamic (MD) simulations for the four protonation states of BACE1 proteins. We applied Glide docking studies to 21 BACE1 inhibitors against the MD extracted conformations. The dynamic results infer that the protein/ligand complex remains stable during the entire simulation course for HD32D228 model. The results show that the hydrogen bonds between the inhibitor and the Asp dyad are maintained in the 10,000th ps snapshot of HD32D228 model. Our results also reveal the significant loop residues in maintaining the active binding conformation in the HD32D228 model. Molecular docking results show that the HD32D228 model provided the best enrichment factor score, suggesting that this model was able to recognize the most active compounds. Our observations provide an evidence for the preference of the anionic state (HD32D228) in BACE1 binding site and are in accord with reported computational data. The protonation state study would provide significant information to assign the correct protonation state for structure-based drug design and docking studies targeting the BACE1 proteins as a tactic to develop potential AD inhibitors. Copyright © 2016 Elsevier Inc. All rights reserved.

Effect of temperature on the protonation of N-(2-hydroxyethyl)ethylenediamine-N,N',N'-triacetic acid in aqueous solutions: Potentiometric and calorimetric studies

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

Li, Xingliang; Zhang, Zhicheng; Endrizzi, Francesco

2015-06-01

The TALSPEAK process (Trivalent Actinide Lanthanide Separations by Phosphorus-reagent Extraction from Aqueous Komplexes) has been demonstrated in several pilot-scale operations to be effective at separating trivalent actinides (An 3+) from trivalent lanthanides (Ln 3+). However, fundamental studies have revealed undesired aspects of TALSPEAK, such as the significant partitioning of Na +, lactic acid, and water into the organic phase, thermodynamically unpredictable pH dependence, and the slow extraction kinetics. In the modified TALSPEAK process, the combination of the aqueous holdback complexant HEDTA (N-(2-hydroxyethyl)ethylenediamine-N,N',N'-triacetic acid) with the extractant HEH[EHP] (2-ethyl(hexyl) phosphonic acid mono-2-ethylhexyl ester) in the organic phase has been found tomore » exhibit a nearly flat pH dependence between 2.5 and 4.5 and more rapid phase transfer kinetics for the heavier lanthanides. To help understand the speciation of Ln 3+ and An 3+ in the modified TALSPEAK, systematic studies are underway on the thermodynamics of major reactions in the HEDTA system under conditions relevant to the process (e.g., higher temperatures). Thermodynamics of the protonation and complexation of HEDTA with Ln 3+ were studied at variable temperatures. Equilibrium constants and enthalpies were determined by a combination of techniques including potentiometry and calorimetry. This paper presents the protonation constants of HEDTA at T = (25 to 70) °C. The potentiometric titrations have demonstrated that, stepwise, the first two protonation constants decrease and the third one slightly increases with the increase of temperature. This trend is in good agreement with the enthalpy of protonation directly determined by calorimetry. The results of NMR analysis further confirm that the first two protonation reactions occur on the diamine nitrogen atoms, while the third protonation reaction occurs on the oxygen of a carboxylate group. These data, in conjunction with the thermodynamic parameters of Ln 3+/An 3+ complexes with HEDTA at different temperatures, will help to predict the speciation and temperature-dependent behavior of Ln 3+/An 3+ in the modified TALSPEAK process.« less

Characterization of complexes between phenethylamine enantiomers and β-cyclodextrin derivatives by capillary electrophoresis-Determination of binding constants and complex mobilities.

PubMed

Wahl, Joachim; Furuishi, Takayuki; Yonemochi, Etsuo; Meinel, Lorenz; Holzgrabe, Ulrike

2017-04-01

To optimize chiral separation conditions and to improve the knowledge of enantioseparation, it is important to know the binding constants K between analytes and cyclodextrins and the electrophoretic mobilities of the temporarily formed analyte-cyclodextrin-complexes. K values for complexes between eight phenethylamine enantiomers, namely ephedrine, pseudoephedrine, methylephedrine and norephedrine, and four different β-cyclodextrin derivatives were determined by affinity capillary electrophoresis. The binding constants were calculated from the electrophoretic mobility values of the phenethylamine enantiomers at increasing concentrations of cyclodextrins in running buffer. Three different linear plotting methods (x-reciprocal, y-reciprocal, double reciprocal) and nonlinear regression were used for the determination of binding constants with β-cyclodextrin, (2-hydroxypropyl)-β-cyclodextrin, methyl-β-cyclodextrin and 6-O-α-maltosyl-β-cyclodextrin. The cyclodextrin concentration in a 50 mM phosphate buffer pH 3.0 was varied from 0 to 12 mM. To investigate the influence of the binding constant values on the enantioseparation the observed electrophoretic selectivities were compared with the obtained K values and the calculated enantiomer-cyclodextrin-complex mobilities. The different electrophoretic mobilities of the temporarily formed complexes were crucial factors for the migration order and enantioseparation of ephedrine derivatives. To verify the apparent binding constants determined by capillary electrophoresis, a titration process using ephedrine enantiomers and β-cyclodextrin was carried out. Furthermore, the isothermal titration calorimetry measurements gave information about the thermal properties of the complexes. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Toward Quantitatively Accurate Calculation of the Redox-Associated Acid–Base and Ligand Binding Equilibria of Aquacobalamin

DOE PAGES

Johnston, Ryne C.; Zhou, Jing; Smith, Jeremy C.; ...

2016-07-08

In redox processes in complex transition metal-containing species are often intimately associated with changes in ligand protonation states and metal coordination number. Moreover, a major challenge is therefore to develop consistent computational approaches for computing pH-dependent redox and ligand dissociation properties of organometallic species. Reduction of the Co center in the vitamin B12 derivative aquacobalamin can be accompanied by ligand dissociation, protonation, or both, making these properties difficult to compute accurately. We examine this challenge here by using density functional theory and continuum solvation to compute Co ligand binding equilibrium constants (Kon/off), pKas and reduction potentials for models of aquacobalaminmore » in aqueous solution. We consider two models for cobalamin ligand coordination: the first follows the hexa, penta, tetra coordination scheme for Co III, Co II, and Co I species, respectively, and the second model features saturation of each vacant axial coordination site on Co II and Co I species with a single, explicit water molecule to maintain six directly interacting ligands or water molecules in each oxidation state. Comparing these two coordination schemes in combination with five dispersion-corrected density functionals, we find that the accuracy of the computed properties is largely independent of the scheme used, but including only a continuum representation of the solvent yields marginally better results than saturating the first solvation shell around Co throughout. PBE performs best, displaying balanced accuracy and superior performance overall, with RMS errors of 80 mV for seven reduction potentials, 2.0 log units for five pK as and 2.3 log units for two log K on/off values for the aquacobalamin system. Furthermore, we find that the BP86 functional commonly used in corrinoid studies suffers from erratic behavior and inaccurate descriptions of Co axial ligand binding, leading to substantial errors in predicted pK as and K on/off values. Finally, these findings demonstrate the effectiveness of the present approach for computing electrochemical and thermodynamic properties of a complex transition metal-containing cofactor.« less

Light-Activated Reversible Imine Isomerization: Towards a Photochromic Protein Switch.

PubMed

Berbasova, Tetyana; Santos, Elizabeth M; Nosrati, Meisam; Vasileiou, Chrysoula; Geiger, James H; Borhan, Babak

2016-03-02

Mutants of cellular retinoic acid-binding protein II (CRABPII), engineered to bind all-trans-retinal as an iminium species, demonstrate photochromism upon irradiation with light at different wavelengths. UV light irradiation populates the cis-imine geometry, which has a high pKa , leading to protonation of the imine and subsequent "turn-on" of color. Yellow light irradiation yields the trans-imine isomer, which has a depressed pKa , leading to loss of color because the imine is not protonated. The protein-bound retinylidene chromophore undergoes photoinduced reversible interconversion between the colored and uncolored species, with excellent fatigue resistance. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Binding Site Configurations Probe the Structure and Dynamics of the Zinc Finger of NEMO (NF-κB Essential Modulator).

PubMed

Godwin, Ryan C; Melvin, Ryan L; Gmeiner, William H; Salsbury, Freddie R

2017-01-31

Zinc-finger proteins are regulators of critical signaling pathways for various cellular functions, including apoptosis and oncogenesis. Here, we investigate how binding site protonation states and zinc coordination influence protein structure, dynamics, and ultimately function, as these pivotal regulatory proteins are increasingly important for protein engineering and therapeutic discovery. To better understand the thermodynamics and dynamics of the zinc finger of NEMO (NF-κB essential modulator), as well as the role of zinc, we present results of 20 μs molecular dynamics trajectories, 5 μs for each of four active site configurations. Consistent with experimental evidence, the zinc ion is essential for mechanical stabilization of the functional, folded conformation. Hydrogen bond motifs are unique for deprotonated configurations yet overlap in protonated cases. Correlated motions and principal component analysis corroborate the similarity of the protonated configurations and highlight unique relationships of the zinc-bound configuration. We hypothesize a potential mechanism for zinc binding from results of the thiol configurations. The deprotonated, zinc-bound configuration alone predominantly maintains its tertiary structure throughout all 5 μs and alludes rare conformations potentially important for (im)proper zinc-finger-related protein-protein or protein-DNA interactions.

Multiparameter flow cytometric analysis of a pH sensitive formyl peptide with application to receptor structure and processing kinetics

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

Fay, S.P.; Domalewski, M.D.; Houghton, T.G.

1994-02-01

Environmentally sensitive molecules have many potential cellular applications. The authors have investigated the utility of a pH sensitive ligand for the formyl peptide receptor, CHO-Met-Leu-Phe-Phe-Lys (SNAFL)-OH (SNAFL-seminaphthofluorescein), because in previous studies protonation has been used to explain the quenching when the fluorescinated formyl pentapeptide ligand binds to this receptor. Moreover, acidification in intracellular compartments is a general mechanism occurring in cells during processing of ligand-receptor complexes. Because the protonated form of SNAFL is excited at 488 nm with emission at 530 nm and the unprotonated form is excited at 568 nm with emission at 650 nm, the ratio of protonatedmore » and unprotonated forms can be examined by multiparameter flow cytometry. The authors found that the receptor-bound ligand is sensitive to both the extracellular and intracellular pH. There is a small increase in the pK[sub a] of the ligand upon binding to the receptor consistent with protonation in the binding pocket. Once internalized, spectral changes in the probe consistent with acidification and ligand dissociation from the receptor are observed. 22 refs., 4 figs.« less

Theoretical study of the adsorption of DNA bases on the acidic external surface of montmorillonite.

PubMed

Mignon, Pierre; Sodupe, Mariona

2012-01-14

In the present study, DFT periodic plane wave calculations, at the PBE-D level of theory, were carried out to investigate the interaction of DNA nucleobases with acidic montmorillonite. The surface model was considered in its octahedral (Osub) and tetrahedral (Tsub) substituted forms, known to have different acidic properties. The adsorption of adenine, guanine and cytosine was considered in both orthogonal and coplanar orientations with the surface, interacting with the proton via a given heteroatom. In almost all considered cases, adsorption involved the spontaneous proton transfer to the nucleobase, with a more pronounced character in the Osub structures. The binding energy is about 10 kcal mol(-1) larger for Osub than for Tsub complexes mainly due to the larger acidity in Osub surfaces and due to the better stabilization by H-bond contacts between the negatively charged surface and the protonated base. The binding energy of coplanar orientations of the base is observed to be as large as the orthogonal ones due to a balance between electrostatic and dispersion contributions. Finally the binding of guanine and adenine on the acidic surface amounts to 50 kcal mol(-1) while that of cytosine rises to 44 kcal mol(-1).

Molecular recognition of naphthalene diimide ligands by telomeric quadruplex-DNA: the importance of the protonation state and mediated hydrogen bonds.

PubMed

Spinello, A; Barone, G; Grunenberg, J

2016-01-28

In depth Monte Carlo conformational scans in combination with molecular dynamics (MD) simulations and electronic structure calculations were applied in order to study the molecular recognition process between tetrasubstituted naphthalene diimide (ND) guests and G-quadruplex (G4) DNA receptors. ND guests are a promising class of telomere stabilizers due to which they are used in novel anticancer therapeutics. Though several ND guests have been studied experimentally in the past, the protonation state under physiological conditions is still unclear. Based on chemical intuition, in the case of N-methyl-piperazine substitution, different protonation states are possible and might play a crucial role in the molecular recognition process by G4-DNA. Depending on the proton concentration, different nitrogen atoms of the N-methyl-piperazine might (or might not) be protonated. This fact was considered in our simulation in terms of a case by case analysis, since the process of molecular recognition is determined by possible donor or acceptor positions. The results of our simulations show that the electrostatic interactions between the ND ligands and the G4 receptor are maximized in the case of the protonation of the terminal nitrogen atoms, forming compact ND G4 complexes inside the grooves. The influence of different protonation states in terms of the ability to form hydrogen bonds with the sugar-phosphate backbone, as well as the importance of mediated vs. direct hydrogen bonding, was analyzed in detail by MD and relaxed force constant (compliance constant) simulations.

Proton conduction within the reaction centers of Rhodobacter capsulatus: the electrostatic role of the protein.

PubMed

Maróti, P; Hanson, D K; Baciou, L; Schiffer, M; Sebban, P

1994-06-07

Light-induced charge separation in the photosynthetic reaction center results in delivery of two electrons and two protons to the terminal quinone acceptor QB. In this paper, we have used flash-induced absorbance spectroscopy to study three strains that share identical amino acid sequences in the QB binding site, all of which lack the protonatable amino acids Glu-L212 and Asp-L213. These strains are the photosynthetically incompetent site-specific mutant Glu-L212/Asp-L213-->Ala-L212/Ala-L213 and two different photocompetent derivatives that carry both alanine substitutions and an intergenic suppressor mutation located far from QB (class 3 strain, Ala-Ala + Arg-M231-->Leu; class 4 strain, Ala-Ala + Asn-M43-->Asp). At pH 8 in the double mutant, we observe a concomitant decrease of nearly 4 orders of magnitude in the rate constants of second electron and proton transfer to QB compared to the wild type. Surprisingly, these rates are increased to about the same extent in both types of suppressor strains but remain > 2 orders of magnitude smaller than those of the wild type. In the double mutant, at pH 8, the loss of Asp-L213 and Glu-L212 leads to a substantial stabilization (> or = 60 meV) of the semiquinone energy level. Both types of compensatory mutations partially restore, to nearly the same level, the original free energy difference for electron transfer from primary quinone QA to QB. The pH dependence of the electron and proton transfer processes in the double-mutant and the suppressor strains suggests that when reaction centers of the double mutant are shifted to lower pH (1.5-2 units), they function like those of the suppressor strains at physiological pH. Our data suggest that the main effect of the compensatory mutations is to partially restore the negative electrostatic environment of QB and to increase an apparent "functional" pK of the system for efficient proton transfer to the active site. This emphasizes the role of the protein in tuning the electrostatic environment of its cofactors and highlights the possible long-range electrostatic effects.

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

Garbuio, Viviana; Pulci, Olivia; Cascella, Michele

The proton disorder in ice has a key role in several properties such as the growth mode, thermodynamical properties, and ferroelectricity. While structural phase transitions from proton disordered to proton ordered ices have been extensively studied, much less is known about their electronic and optical properties. Here, we present ab initio many body perturbation theory-based calculations of the electronic and optical properties of cubic ice at different levels of proton disorder. We compare our results with those from liquid water, that acts as an example of a fully (proton- and oxygen-)disordered system. We find that by increasing the proton disorder,more » a shrinking of the electronic gap occurs in ice, and it is smallest in the liquid water. Simultaneously, the excitonic binding energy decreases, so that the final optical gaps result to be almost independent on the degree of proton disorder. We explain these findings as an interplay between the local dipolar disorder and the electronic correlation.« less

Kinetic Effects Of Increased Proton Transfer Distance On Proton-Coupled Oxidations Of Phenol-Amines

PubMed Central

Rhile, Ian J.

2011-01-01

To test the effect of varying the proton donor-acceptor distance in proton-coupled electron transfer (PCET) reactions, the oxidation of a bicyclic amino-indanol (2) is compared with that of a closely related phenol with an ortho CPh2NH2 substituent (1). Spectroscopic, structural, thermochemical and computational studies show that the two amino-phenols are very similar, except that the O⋯N distance (dON) is >0.1 Å longer in 2 than in 1. The difference in dON is 0.13 ± 0.03 Å from X-ray crystallography and 0.165 Å from DFT calculations. Oxidations of these phenols by outer-sphere oxidants yield distonic radical cations •OAr–NH3+ by concerted proton-electron transfer (CPET). Simple tunneling and classical kinetic models both predict that the longer donor-acceptor distance in 2 should lead to slower reactions, by ca. two orders of magnitude, as well as larger H/D kinetic isotope effects (KIEs). However, kinetic studies show that the compound with the longer proton-transfer distance, 2, exhibits smaller KIEs and has rate constants that are quite close to those of 1. For example, the oxidation of 2 by the triarylamminium radical cation N(C6H4OMe)3•+ (3a+) occurs at (1.4 ± 0.1) × 104 M-1 s-1, only a factor of two slower than the closely related reaction of 1 with N(C6H4OMe)2(C6H4Br)•+ (3b+). This difference in rate constants is well accounted for by the slightly different free energies of reaction: ΔG°(2 + 3a+) = +0.078 V vs. ΔG°(1 + 3b+) = +0.04 V. The two phenol-amines do display some subtle kinetic differences: for instance, compound 2 has a shallower dependence of CPET rate constants on driving force (Brønsted α, Δln(k)/Δln(Keq)). These results show that the simple tunneling model is not a good predictor of the effect of proton donor-acceptor distance on concerted-electron transfer reactions involving strongly hydrogen-bonded systems. Computational analysis of the observed similarity of the two phenols emphasizes the importance of the highly anharmonic O⋯H⋯N potential energy surface and the influence of proton vibrational excited states. PMID:21919508

Observation of an energy dependence of the radiation damage on standard and oxygenated silicon diodes by 16, 21, and 27 MeV protons

NASA Astrophysics Data System (ADS)

Wyss, J.; Bisello, D.; Candelori, A.; Kaminsky, A.; Pantano, D.

2001-01-01

First measurement of the energy dependence of the radiation damage induced by low-energy protons on standard and oxygen enriched diodes is presented. The current damage constant α is always insensitive to the oxygen content and increases for lower energy protons, whereas the acceptor creation rate β for both types of diodes slowly decreases for lower proton energies, this effect being amplified when the fluences are normalized to their 1 MeV neutron equivalent values. The dependence from the proton energy of the normalized β values is in open disagreement with the currently accepted NIEL hypothesis. Irradiations and measurements have been performed at the INFN Laboratorio Nazionale di Legnaro.

Carotenoid Antenna Binding and Function in Retinal Proteins

DTIC Science & Technology

2012-08-13

REPORT Carotenoid antenna binding and function in retinal proteins 14. ABSTRACT 16. SECURITY CLASSIFICATION OF: Xanthorhodopsin, a proton pump from the...eubacterium Salinibacter ruber, is a unique dual chromophore system that contains, in addition to retinal, the carotenoid salinixanthin as a light... carotenoid ring near the retinal ring. Substitution of the small glycine with bulky tryptophan in this site eliminates binding. The second factor is the 4

Photodissociation pathways and lifetimes of protonated peptides and their dimers

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

Aravind, G.; Klaerke, B.; Rajput, J.

2012-01-07

Photodissociation lifetimes and fragment channels of gas-phase, protonated YA{sub n} (n = 1,2) peptides and their dimers were measured with 266 nm photons. The protonated monomers were found to have a fast dissociation channel with an exponential lifetime of {approx}200 ns while the protonated dimers show an additional slow dissociation component with a lifetime of {approx}2 {mu}s. Laser power dependence measurements enabled us to ascribe the fast channel in the monomer and the slow channel in the dimer to a one-photon process, whereas the fast dimer channel is from a two-photon process. The slow (1 photon) dissociation channel in themore » dimer was found to result in cleavage of the H-bonds after energy transfer through these H-bonds. In general, the dissociation of these protonated peptides is non-prompt and the decay time was found to increase with the size of the peptides. Quantum RRKM calculations of the microcanonical rate constants also confirmed a statistical nature of the photodissociation processes in the dipeptide monomers and dimers. The classical RRKM expression gives a rate constant as an analytical function of the number of active vibrational modes in the system, estimated separately on the basis of the equipartition theorem. It demonstrates encouraging results in predicting fragmentation lifetimes of protonated peptides. Finally, we present the first experimental evidence for a photo-induced conversion of tyrosine-containing peptides into monocyclic aromatic hydrocarbon along with a formamide molecule both found in space.« less

Exploiting hydrogen bonding interactions to probe smaller linear and cyclic diamines binding to G-quadruplexes: a DFT and molecular dynamics study.

PubMed

Kanti Si, Mrinal; Sen, Anik; Ganguly, Bishwajit

2017-05-10

G-quadruplexes are formed by the association of four guanine bases through Hoogsteen hydrogen bonding in guanine-rich sequences of DNA and exist in the telomere as well as in promoter regions of certain oncogenes. The sequences of G-quadruplex-DNA are targets for the design of molecules that can bind and can be developed as anti-cancer drugs. The linear and cyclic protonated diamines have been explored to bind to G-quadruplex-DNA through hydrogen bonding interactions. The quadruplex-DNA binders exploit π-stacking and hydrogen bonding interactions with the phosphate backbone of loops and grooves. In this study, linear and cyclic protonated diamines showed remarkable binding affinity for G-tetrads using hydrogen bonding interactions. The DFT M06-2X/6-31G(d)//B3LYP/6-31+G(d) level of theory showed that the cyclic ee-1,2-CHDA (equatorial-equatorial form of 1,2-disubstituted cyclohexadiamine di-cation) binds to the G-tetrads very strongly (∼70.0 kcal mol -1 ), with a much higher binding energy than the linear protonated diamines. The binding affinity of ligands for G-tetrads with counterions has also been examined. The binding preference of these small ligands for G-tetrads is higher than for DNA-duplex. The binding affinity of an intercalated acridine-based ligand (BRACO-19) for G-quadruplexes has been examined and the binding energy is relatively lower than that for the 1,2 disubstituted cyclohexadiamine di-cation with G-tetrads. The atoms-in-molecules (AIM) analysis reveals that the hydrogen bonding interactions between the organic systems with G-tetrads are primarily electrostatic in nature. The molecular dynamics simulations performed using a classical force field (GROMACS) also supported the phosphate backbone sites of G-quadruplex-DNA to bind to these diamines. To mimic the structural pattern of BRACO-19, the designed inhibitor N,2-bis-2(3,4-aminocyclohexyl) acetamide (9) examined possesses two 1,2-CHDA moieties linked through an acetamide group. The molecular dynamics results showed that the designed molecule 9 can efficiently bind to the base-pairs and the phosphate backbone of G quadruplex-DNA using H-bonding interactions. The binding affinity calculated for the intercalated acridine-based drug (BRACO-19) with G-quadruplexes is weaker compared to ee-1,2-CHDA. These ligands deliver a different binding motif (hydrogen bonding) compared to the reported G-quadruplex binders of π-delocalized systems and will kindle interest in examining such scaffolds to stabilize DNA.

Calorimetric and spectroscopic studies of the interaction between zidovudine and human serum albumin

NASA Astrophysics Data System (ADS)

Pîrnău, Adrian; Mic, Mihaela; Neamţu, Silvia; Floare, Călin G.; Bogdan, Mircea

2018-02-01

A quantitative analysis of the interaction between zidovudine (AZT) and human serum albumin (HSA) was achieved using Isothermal titration calorimetry (ITC) in combination with fluorescence and 1H NMR spectroscopy. ITC directly measure the heat during a biomolecular binding event and gave us thermodynamic parameters and the characteristic association constant. By fluorescence quenching, the binding parameters of AZT-HSA interaction was determined and location to binding site I of HSA was confirmed. Via T1 NMR selective relaxation time measurements the drug-protein binding extent was evaluated as dissociation constants Kd and the involvement of azido moiety of zidovudine in molecular complex formation was put in evidence. All three methods indicated a very weak binding interaction. The association constant determined by ITC (3.58 × 102 M- 1) is supported by fluorescence quenching data (2.74 × 102 M- 1). The thermodynamic signature indicates that at least hydrophobic and electrostatic type interactions played a main role in the binding process.

Revised Model of Calcium and Magnesium Binding to the Bacterial Cell Wall

PubMed Central

Thomas, Kieth J.; Rice, Charles V.

2014-01-01

Metals bind to the bacterial cell wall yet the binding mechanisms and affinity constants are not fully understood. The cell wall of gram positive bacteria is characterized by a thick layer of peptidoglycan and anionic teichoic acids anchored in the cytoplasmic membrane (lipoteichoic acid) or covalently bound to the cell wall (wall teichoic acid). The polyphosphate groups of teichoic acid provide one-half of the metal binding sites for calcium and magnesium, contradicting previous reports that calcium binding is 100% dependent on teichoic acid. The remaining binding sites are formed with the carboxyl units of peptidoglycan. In this work we report equilibrium association constants and total metal binding capacities for the interaction of calcium and magnesium ions with the bacterial cell wall. Metal binding is much stronger and previously reported. Curvature of Scatchard plots from the binding data and the resulting two regions of binding affinity suggest the presence of negative cooperative binding, meaning that the binding affinity decreases as more ions become bound to the sample. For Ca2+, Region I has a KA = (1.0 ± 0.2) × 106 M−1 and Region II has a KA = (0.075 ± 0.058) × 106 M−1. For Mg2+, KA1 = (1.5 ± 0.1) × 106 and KA2 = (0.17 ± 0.10) × 106. A binding capacity (η) is reported for both regions. However, since binding is still occurring in Region II, the total binding capacity is denoted by η2, which are 0.70 ± 0.04 µmol/mg and 0.67 ± 0.03 µmol/mg for Ca2+ and Mg2+ respectively. These data contradict the current paradigm of there being a single metal affinity value that is constant over a range of concentrations. We also find that measurement of equilibrium binding constants is highly sample dependent, suggesting a role for diffusion of metals through heterogeneous cell wall fragments. As a result, we are able to reconcile many contradictory theories that describe binding affinity and the binding mode of divalent metal cations. PMID:25315444

Binding of resveratrol with sodium caseinate in aqueous solutions.

PubMed

Acharya, Durga P; Sanguansri, Luz; Augustin, Mary Ann

2013-11-15

The interaction between resveratrol (Res) and sodium caseinate (Na-Cas) has been studied by measuring fluorescence quenching of the protein by resveratrol. Quenching constants were determined using Stern-Volmer equation, which suggests that both dynamic and static quenching occur between Na-Cas and Res. Binding constants for the complexation between Na-Cas and Res were determined at different temperatures. The large binding constants (3.7-5.1×10(5)M(-1)) suggest that Res has strong affinity for Na-Cas. This affinity decreases as the temperature is raised from 25 to 37°C. The binding involves both hydrogen bonding and hydrophobic interaction, as suggested by negative enthalpy change and positive entropy change for the binding reaction. The present study indicates that Na-Cas, a common food protein, may be used as a carrier of Res, a bioactive polyphenol which is insoluble in both water and oils. Copyright © 2013 Elsevier Ltd. All rights reserved.

Determination of the pairing-strength constants in the isovector plus isoscalar pairing case

NASA Astrophysics Data System (ADS)

Mokhtari, D.; Fellah, M.; Allal, N. H.

2016-05-01

A method for the determination of the pairing-strength constants, in the neutron-proton (n-p) isovector plus isoscalar pairing case, is proposed in the framework of the BCS theory. It is based on the fitting of these constants to reproduce the experimentally known pairing gap parameters as well as the root-mean-squared (r.m.s) charge radii values. The method is applied to some proton-rich even-even nuclei. The single-particle energies used are those of a deformed Woods-Saxon mean field. It is shown that the obtained value of the ratio GnpT=0/G npT=1 is of the same order as the ones, arbitrary chosen, of some previous works. The effect of the inclusion of the isoscalar n-p pairing in the r.m.s matter radii is then numerically studied for the same nuclei.

On proton excitation of forbidden lines in positive ions

NASA Astrophysics Data System (ADS)

Burgess, Alan; Tully, John A.

2005-08-01

The semi-classical impact parameter approximations used by Bahcall and Wolf and by Bely and Faucher, for proton excitation of electric quadrupole transitions in positive ions, both fail at high energies, giving cross sections which do not fall off correctly as constant/E. This is in contrast with the pioneering example of Seaton for Fe+13 and of Reid and Schwarz for S+3, both of whom achieve the correct functional form, but do not ensure the correct constant of proportionality. By combining the Born and semi-classical approximations one can obtain cross sections which have the full correct behaviour as E → ∞, and hence, rate coefficients which have the correct high temperature behaviour (~C/T1/2 with the correct value of C). We provide a computer program for calculating these. An error in Faucher's derivation of the Born formula is also discussed.

Electrochemical and spectroscopic studies of the interaction of proflavine with DNA.

PubMed

Aslanoglu, Mehmet

2006-03-01

The interaction of proflavine with herring sperm DNA has been investigated by cyclic voltammetry and UV-Vis spectroscopy as well as viscosity measurements. Shifts in the peak potentials in cyclic voltammetry, spectral changes in UV absorption titration, an increase in viscosity of DNA and the results of the effect of ionic strength on the binding constant strongly support the intercalation of proflavine into the DNA double helix. The binding constant for the interaction between proflavine and DNA was K = 2.32 (+/- 0.41) x 10(4) M(-1) and the binding site size was 2.07 (+/- 0.1) base pairs, estimated in voltammetric measurements. The value of the binding site size was determined to be closer to that expected for a planar intercalating agent. The standard Gibbs free-energy change is ca. -24.90 kJ/mol at 25 degrees C, indicating the spontaneity of the binding interaction. The binding constant determined by UV absorption measurements was K = 2.20 (+/- 0.48) x 10(4) M(-1), which is very close to the value determined by cyclic voltammetry assuming that the binding equilibrium is static.

Effect of temperature on the acid-base properties of the alumina surface: microcalorimetry and acid-base titration experiments.

PubMed

Morel, Jean-Pierre; Marmier, Nicolas; Hurel, Charlotte; Morel-Desrosiers, Nicole

2006-06-15

Sorption reactions on natural or synthetic materials that can attenuate the migration of pollutants in the geosphere could be affected by temperature variations. Nevertheless, most of the theoretical models describing sorption reactions are at 25 degrees C. To check these models at different temperatures, experimental data such as the enthalpies of sorption are thus required. Highly sensitive microcalorimeters can now be used to determine the heat effects accompanying the sorption of radionuclides on oxide-water interfaces, but enthalpies of sorption cannot be extracted from microcalorimetric data without a clear knowledge of the thermodynamics of protonation and deprotonation of the oxide surface. However, the values reported in the literature show large discrepancies and one must conclude that, amazingly, this fundamental problem of proton binding is not yet resolved. We have thus undertaken to measure by titration microcalorimetry the heat effects accompanying proton exchange at the alumina-water interface at 25 degrees C. Based on (i) the surface sites speciation provided by a surface complexation model (built from acid-base titrations at 25 degrees C) and (ii) results of the microcalorimetric experiments, calculations have been made to extract the enthalpic variations associated respectively to first and second deprotonation of the alumina surface. Values obtained are deltaH1 = 80+/-10 kJ mol(-1) and deltaH2 = 5+/-3 kJ mol(-1). In a second step, these enthalpy values were used to calculate the alumina surface acidity constants at 50 degrees C via the van't Hoff equation. Then a theoretical titration curve at 50 degrees C was calculated and compared to the experimental alumina surface titration curve. Good agreement between the predicted acid-base titration curve and the experimental one was observed.

Copper(II) binding by dissolved organic matter: Importance of the copper-to-dissolved organic matter ratio and implications for the Biotic Ligand Model

USGS Publications Warehouse

Craven, Alison M.; Aiken, George R.; Ryan, Joseph N.

2012-01-01

The ratio of copper to dissolved organic matter (DOM) is known to affect the strength of copper binding by DOM, but previous methods to determine the Cu2+–DOM binding strength have generally not measured binding constants over the same Cu:DOM ratios. In this study, we used a competitive ligand exchange–solid-phase extraction (CLE-SPE) method to determine conditional stability constants for Cu2+–DOM binding at pH 6.6 and 0.01 M ionic strength over a range of Cu:DOM ratios that bridge the detection windows of copper-ion-selective electrode and voltammetry measurements. As the Cu:DOM ratio increased from 0.0005 to 0.1 mg of Cu/mg of DOM, the measured conditional binding constant (cKCuDOM) decreased from 1011.5 to 105.6 M–1. A comparison of the binding constants measured by CLE-SPE with those measured by copper-ion-selective electrode and voltammetry demonstrates that the Cu:DOM ratio is an important factor controlling Cu2+–DOM binding strength even for DOM isolates of different types and different sources and for whole water samples. The results were modeled with Visual MINTEQ and compared to results from the biotic ligand model (BLM). The BLM was found to over-estimate Cu2+ at low total copper concentrations and under-estimate Cu2+ at high total copper concentrations.

Mechanism of electrocatalytic hydrogen production by a di-iron model of iron-iron hydrogenase: a density functional theory study of proton dissociation constants and electrode reduction potentials.

PubMed

Surawatanawong, Panida; Tye, Jesse W; Darensbourg, Marcetta Y; Hall, Michael B

2010-03-28

Simple dinuclear iron dithiolates such as (mu-SCH2CH2CH2S)[Fe(CO)3]2, (1) and (mu-SCH2CH2S)[Fe(CO)3]2 (2) are functional models for diiron-hydrogenases, [FeFe]-H2ases, that catalyze the reduction of protons to H2. The mechanism of H2 production with 2 as the catalyst and with both toluenesulfonic (HOTs) and acetic (HOAc) acids as the H+ source in CH3CN solvent has been examined by density functional theory (DFT). Proton dissociation constants (pKa) and electrode reduction potentials (E(o)) are directly computed and compared to the measured pKa of HOTs and HOAc acids and the experimental reduction potentials. Computations show that when the strong acid, HOTs, is used as a proton source the one-electron reduced species 2- can be protonated to form a bridging hydride complex as the most stable structure. Then, this species can be reduced and protonated to form dihydrogen and regenerate 2. This cycle produces H2 via an ECEC process at an applied potential of -1.8 V vs. Fc/Fc+. A second faster process opens for this system when the species produced at the ECEC step above is further reduced and H2 release returns the system to 2- rather than 2, an E[CECE] process. On the other hand, when the weak acid, HOAc, is the proton source a more negative applied reduction potential (-2.2 V vs. Fc/Fc+) is necessary. At this potential two one-electron reductions yield the dianion 2(2-) before the first protonation, which in this case occurs on the thiolate. Subsequent reduction and protonation form dihydrogen and regenerate 2- through an E[ECEC] process.

Titration ELISA as a Method to Determine the Dissociation Constant of Receptor Ligand Interaction.

PubMed

Eble, Johannes A

2018-02-15

The dissociation constant describes the interaction between two partners in the binding equilibrium and is a measure of their affinity. It is a crucial parameter to compare different ligands, e.g., competitive inhibitors, protein isoforms and mutants, for their binding strength to a binding partner. Dissociation constants are determined by plotting concentrations of bound versus free ligand as binding curves. In contrast, titration curves, in which a signal that is proportional to the concentration of bound ligand is plotted against the total concentration of added ligand, are much easier to record. The signal can be detected spectroscopically and by enzyme-linked immunosorbent assay (ELISA). This is exemplified in a protocol for a titration ELISA that measures the binding of the snake venom-derived rhodocetin to its immobilized target domain of α2β1 integrin. Titration ELISAs are versatile and widely used. Any pair of interacting proteins can be used as immobilized receptor and soluble ligand, provided that both proteins are pure, and their concentrations are known. The difficulty so far has been to determine the dissociation constant from a titration curve. In this study, a mathematical function underlying titration curves is introduced. Without any error-prone graphical estimation of a saturation yield, this algorithm allows processing of the raw data (signal intensities at different concentrations of added ligand) directly by mathematical evaluation via non-linear regression. Thus, several titration curves can be recorded simultaneously and transformed into a set of characteristic parameters, among them the dissociation constant and the concentration of binding-active receptor, and they can be evaluated statistically. When combined with this algorithm, titration ELISAs gain the advantage of directly presenting the dissociation constant. Therefore, they may be used more efficiently in the future.

[19F]fluorine nuclear-magnetic-resonance study of the interaction of difluoro-oxaloacetate with aspartate transaminase.

PubMed Central

Briley, P A; Eisenthal, R; Harrison, R; Smith, G D

1977-01-01

Difluoro-oxaloacetate interacts with the aldimine form of aspartate transaminase to give a complex, the dissociation constant of which has been determined spectrophotometrically and by 19F n.m.r. (nuclear magnetic resonance). The 19F n.m.r. line-width-pH and chemical-shift-pH profiles of difluoro-oxaloacetate in the presence of the aldimine form of the enzyme both show inflexion points in the pH5 and pH8 regions, which may arise from variations in the binding of difluoro-oxaloacetate as specific groups on the enzyme are successively protonated. Difluoro-oxaloacetate also interacts with apoenzyme to form a complex, the dissociation constant of which was determined by 19F n.m.r. The 19F n.m.r. line-width-pH and chemical-shift-pH profiles of difluoro-oxaloacetate in the presence of apoenzyme show a single inflexion point in the region of pH8. The absence, in this case, of an inflexion in the pH5 region indicates that the latter, present in the corresponding profiles for the aldimine form of the enzyme, results from ionization of an enzyme group associated with the pyridoxal phosphate cofactor. PMID:17399

Binding the diproton in stars: anthropic limits on the strength of gravity

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

Barnes, Luke A., E-mail: L.Barnes@physics.usyd.edu.au

2015-12-01

We calculate the properties and investigate the stability of stars that burn via strong (and electromagnetic) interactions, and compare their properties with those that, as in our Universe, include a rate-limiting weak interaction. It has been suggested that, if the diproton were bound, stars would burn ∼10{sup 18} times brighter and faster via strong interactions, resulting in a universe that would fail to support life. By considering the representative case of a star in our Universe with initially equal numbers of protons and deuterons, we find that stable, 'strong-burning' stars adjust their central densities and temperatures to have familiar surfacemore » temperatures, luminosities and lifetimes. There is no 'diproton disaster'. In addition, strong-burning stars are stable in a much larger region of the parameter space of fundamental constants, specifically the strength of electromagnetism and gravity. The strongest anthropic bound on stars in such universes is not their stability, as is the case for stars limited by the weak interaction, but rather their lifetime. Regardless of the strength of electromagnetism, all stars burn out in mere millions of years unless the gravitational coupling constant is extremely small, α{sub G}∼< 10{sup −30}.« less

Voltammetric behavior of Mammeisin (MA) at a glassy carbon electrode and its interaction with Bovine Serum Albumin (BSA).

PubMed

Leuna, Jules-Blaise Mabou; Sop, Sergeot Kungo; Makota, Suzanne; Njanja, Evangeline; Ebelle, Thiery Christophe; Azebaze, Anatole Guy; Ngameni, Emmanuel; Nassi, Achille

2018-02-01

The electrochemical oxidation of Mammeisin (MA) was studied in a solution containing acetone and 0.1M phosphate buffer +0.1M KCl (pH=5.3) at a glassy carbon electrode (GCE), using cyclic (CV) and square wave voltammetry (SWV). MA showed a quasi-reversible process, which is pH dependent and that involves the exchange of two electrons and two protons. The oxidation product was adsorbed by the electrode surface to form a film that blocks active sites over repetitive cyclic. Moreover, the interaction of MA and bovine serum albumin (BSA) was studied by CV and SWV at different pHs (5.4, 7.2, 9.5). As a result of the affinity binding with BSA, electrochemically inactive complex was formed. In addition, the oxidation potential of MA in the presence of BSA depends on the pH. The diffusion coefficients of both free and bound MA were estimated from the cyclic voltammetry data using the method developed by Randles-Sevich (D f =9.85×10 -5 cm 2 s -1 and D b =1.27×10 -9 cm 2 s -1 ) and the binding constant of MA-BSA complex, K=3.47×10 2 Lmol -1 , was obtained. Copyright © 2017. Published by Elsevier B.V.

Supramolecular polymeric chemosensor for biomedical applications: design and synthesis of a luminescent zinc metallopolymer as a chemosensor for adenine detection.

PubMed

Chow, Cheuk-Fai

2012-11-01

Adenine is an important bio-molecule that plays many crucial roles in food safety and biomedical diagnostics. Differentiating adenine from a mixture of adenosine and other nucleic bases (guanine, thymine, cytosine, and uracil) is particularly important for both biological and clinical applications. A neutral Zn(II) metallosupramolecular polymer based on acyl hydrazone derived coordination centres (P1) were generated through self-assembly polymerization. It is a linear coordination polymer that behaves like self-standing film. The synthesis, (1)H-NMR characterization, and spectroscopic properties of this supramolecular material are reported. P1 was found to be a chemosensor specific to adenine, with a luminescent enhancement. The binding properties of P1 with common nucleic bases and nucleosides reveal that this supramolecular polymer is very selective to adenine molecules (~20 to 420 times more selectivity than other nucleic bases). The formation constant (K) of P1 to adenine was found to be log K = 4.10 ± 0.02. This polymeric chemosensor produces a specific response to adenine down to 90 ppb. Spectrofluorimetric and (1)H-NMR titration studies showed that the P1 polymer allows each Zn(II) coordination centre to bind to two adenine molecules through hydrogen bonding with their imine and hydrazone protons.

Low capping group surface density on zinc oxide nanocrystals.

PubMed

Valdez, Carolyn N; Schimpf, Alina M; Gamelin, Daniel R; Mayer, James M

2014-09-23

The ligand shell of colloidal nanocrystals can dramatically affect their stability and reaction chemistry. We present a methodology to quantify the dodecylamine (DDA) capping shell of colloidal zinc oxide nanocrystals in a nonpolar solvent. Using NMR spectroscopy, three different binding regimes are observed: strongly bound, weakly associated, and free in solution. The surface density of bound DDA is constant over a range of nanocrystal sizes, and is low compared to both predictions of the number of surface cations and maximum coverages of self-assembled monolayers. The density of strongly bound DDA ligands on the as-prepared ZnO NCs is 25% of the most conservative estimate of the maximum surface DDA density. Thus, these NCs do not resemble the common picture of a densely capped surface ligand layer. Annealing the ZnO NCs in molten DDA for 12 h at 160 °C, which is thought to remove surface hydroxide groups, resulted in a decrease of the weakly associated DDA and an increase in the density of strongly bound DDA, to ca. 80% of the estimated density of a self-assembled monolayer on a flat ZnO surface. These findings suggest that as-prepared nanocrystal surfaces contain hydroxide groups (protons on the ZnO surfaces) that inhibit strong binding of DDA.

Rationalizing 5000-Fold Differences in Receptor-Binding Rate Constants of Four Cytokines

PubMed Central

Pang, Xiaodong; Qin, Sanbo; Zhou, Huan-Xiang

2011-01-01

The four cytokines erythropoietin (EPO), interleukin-4 (IL4), human growth hormone (hGH), and prolactin (PRL) all form four-helix bundles and bind to type I cytokine receptors. However, their receptor-binding rate constants span a 5000-fold range. Here, we quantitatively rationalize these vast differences in rate constants by our transient-complex theory for protein-protein association. In the transient complex, the two proteins have near-native separation and relative orientation, but have yet to form the short-range specific interactions of the native complex. The theory predicts the association rate constant as ka=ka0exp(−ΔGel∗/kBT) where ka0 is the basal rate constant for reaching the transient complex by random diffusion, and the Boltzmann factor captures the rate enhancement due to electrostatic attraction. We found that the vast differences in receptor-binding rate constants of the four cytokines arise mostly from the differences in charge complementarity among the four cytokine-receptor complexes. The basal rate constants (ka0) of EPO, IL4, hGH, and PRL were similar (5.2 × 105 M−1s−1, 2.4 × 105 M−1s−1, 1.7 × 105 M−1s−1, and 1.7 × 105 M−1s−1, respectively). However, the average electrostatic free energies (ΔGe1∗) were very different (−4.2 kcal/mol, −2.4 kcal/mol, −0.1 kcal/mol, and −0.5 kcal/mol, respectively, at ionic strength = 160 mM). The receptor-binding rate constants predicted without adjusting any parameters, 6.2 × 108 M−1s−1, 1.3 × 107 M−1s−1, 2.0 × 105 M−1s−1, and 7.6 × 104 M−1s−1, respectively, for EPO, IL4, hGH, and PRL agree well with experimental results. We uncover that these diverse rate constants are anticorrelated with the circulation concentrations of the cytokines, with the resulting cytokine-receptor binding rates very close to the limits set by the half-lives of the receptors, suggesting that these binding rates are functionally relevant and perhaps evolutionarily tuned. Our calculations also reproduced well-observed effects of mutations and ionic strength on the rate constants and produced a set of mutations on the complex of hGH with its receptor that putatively enhances the rate constant by nearly 100-fold through increasing charge complementarity. To quantify charge complementarity, we propose a simple index based on the charge distribution within the binding interface, which shows good correlation with ΔGe1∗. Together these results suggest that protein charges can be manipulated to tune ka and control biological function. PMID:21889455

Comparative study of substrate and product binding to the human ABO(H) blood group glycosyltransferases.

PubMed

Soya, Naoto; Shoemaker, Glen K; Palcic, Monica M; Klassen, John S

2009-11-01

The first comparative thermodynamic study of the human blood group glycosyltransferases, alpha-(1-->3)-N-acetylgalactosaminyltransferase (GTA) and alpha-(1-->3)-galactosyltransferase (GTB), interacting with donor substrates, donor and acceptor analogs, and trisaccharide products in vitro is reported. The binding constants, measured at 24 degrees C with the direct electrospray ionization mass spectrometry (ES-MS) assay, provide new insights into these model GTs and their interactions with substrate and product. Notably, the recombinant forms of GTA and GTB used in this study are shown to exist as homodimers, stabilized by noncovalent interactions at neutral pH. In the absence of divalent metal ion, neither GTA nor GTB exhibits any appreciable affinity for its native donors (UDP-GalNAc, UDP-Gal). Upon introduction of Mn(2+), both donors undergo enzyme-catalyzed hydrolysis in the presence of either GTA or GTB. Hydrolysis of UDP-GalNAc in the presence of GTA proceeds very rapidly under the solution conditions investigated and a binding constant could not be directly measured. In contrast, the rate of hydrolysis of UDP-Gal in the presence of GTB is significantly slower and, utilizing a modified approach to analyze the ES-MS data, a binding constant of 2 x 10(4) M(-1) was established. GTA and GTB bind the donor analogs UDP-GlcNAc, UDP-Glc with affinities similar to those measured for UDP-Gal and UDP-GalNAc (GTB only), suggesting that the native donors and donor analogs bind to the GTA and GTB through similar interactions. The binding constant determined for GTA and UDP-GlcNAc (approximately 1 x 10(4) M(-1)), therefore, provides an estimate for the binding constant for GTA and UDP-GalNAc. Binding of GTA and GTB with the A and B trisaccharide products was also investigated for the first time. In the absence of UDP and Mn(2+), both GTA and GTB recognize their respective trisaccharide products but with a low affinity approximately 10(3) M(-1); the presence of UDP and Mn(2+) has no effect on A trisaccharide binding but precludes B-trisaccharide binding.

Influence of sulfhydryl sites on metal binding by bacteria

NASA Astrophysics Data System (ADS)

Nell, Ryan M.; Fein, Jeremy B.

2017-02-01

The role of sulfhydryl sites within bacterial cell envelopes is still unknown, but the sites may control the fate and bioavailability of metals. Organic sulfhydryl compounds are important complexing ligands in aqueous systems and they can influence metal speciation in natural waters. Though representing only approximately 5-10% of the total available binding sites on bacterial surfaces, sulfhydryl sites exhibit high binding affinities for some metals. Due to the potential importance of bacterial sulfhydryl sites in natural systems, metal-bacterial sulfhydryl site binding constants must be determined in order to construct accurate models of the fate and distribution of metals in these systems. To date, only Cd-sulfhydryl binding has been quantified. In this study, the thermodynamic stabilities of Mn-, Co-, Ni-, Zn-, Sr- and Pb-sulfhydryl bacterial cell envelope complexes were determined for the bacterial species Shewanella oneidensis MR-1. Metal adsorption experiments were conducted as a function of both pH, ranging from 5.0 to 7.0, and metal loading, from 0.5 to 40.0 μmol/g (wet weight) bacteria, in batch experiments in order to determine if metal-sulfhydryl binding occurs. Initially, the data were used to calculate the value of the stability constants for the important metal-sulfhydryl bacterial complexes for each metal-loading condition studied, assuming a single binding reaction for the dominant metal-binding site type under the pH conditions of the experiments. For most of the metals that we studied, these calculated stability constant values increased significantly with decreasing metal loading, strongly suggesting that our initial assumption was not valid and that more than one type of binding occurs at the assumed binding site. We then modeled each dataset with two distinct site types with identical acidity constants: one site with a high metal-site stability constant value, which we take to represent metal-sulfhydryl binding and which dominates under low metal loading conditions, and another more abundant site that we term non-sulfhydryl sites that becomes important at high metal loadings. The resulting calculated stability constants do not vary significantly as a function of metal loading and yield reasonable fits to the observed adsorption behaviors as a function of both pH and metal loading. We use the results to calculate the speciation of metals bound by the bacterial envelope in realistic bacteria-bearing, heavy metal contaminated systems in order to demonstrate the potential importance of metal-sulfhydryl binding in the budget of bacterially-adsorbed metals under low metal-loading conditions.

On the interpretation of quantitative structure–function activity relationship data for lactate oxidase

PubMed Central

Yorita, Kazuko; Misaki, Hideo; Palfey, Bruce A.; Massey, Vincent

2000-01-01

The native flavin, FMN, has been removed from the l-lactate oxidase of Aerococcus viridans, and the apoprotein reconstituted with 12 FMN derivatives with various substituents at the flavin 6- and 8-positions. Impressive linear relationships are exhibited between the sum of the Hammett σpara and σortho parameters and the redox potentials of the free flavins, and between the redox potentials of the free and enzyme-bound flavins. Rapid reaction kinetics studies of the reconstituted enzymes with the substrates l-lactate and l-mandelate show an increase in the reduction rate constant with increasing redox potential, except that, with lactate, a limiting rate constant of ≈700 s−1 is obtained with flavins of high potential. Similar breakpoints are found in plots of the rate constants for flavin N5-sulfite adduct formation and for the reaction of the reduced enzymes with molecular oxygen. These results are interpreted in terms of a two-step equilibrium preceding the chemical reaction step, in which the second equilibrium step provides an upper limit to the rate with which the particular substrate or ligand is positioned with the flavin in the correct fashion for the observed chemical reaction to occur. The relationship of rate constants for flavin reduction and N5-sulfite adduct formation with flavin redox potential below the observed breakpoint indicate development of significant negative charge in the transition states of the reactions. In the case of reduction by substrate, the results are consistent either with a hydride transfer mechanism or with the so called “carbanion” mechanism, in which the substrate α-proton is abstracted by an enzyme base protected from exchange with solvent. These conclusions are supported by substrate α-deuterium isotope effects and by solvent viscosity effects on sulfite binding. PMID:10706608

Nuclear binding energy using semi empirical mass formula

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

Ankita,, E-mail: ankitagoyal@gmail.com; Suthar, B.

2016-05-06

In the present communication, semi empirical mass formula using the liquid drop model has been presented. Nuclear binding energies are calculated using semi empirical mass formula with various constants given by different researchers. We also compare these calculated values with experimental data and comparative study for finding suitable constants is added using the error plot. The study is extended to find the more suitable constant to reduce the error.

An integrated catch-and-hold mechanism activates nicotinic acetylcholine receptors.

PubMed

Jadey, Snehal; Auerbach, Anthony

2012-07-01

In neuromuscular acetylcholine (ACh) receptor channels (AChRs), agonist molecules bind with a low affinity (LA) to two sites that can switch to high affinity (HA) and increase the probability of channel opening. We measured (by using single-channel kinetic analysis) the rate and equilibrium constants for LA binding and channel gating for several different agonists of adult-type mouse AChRs. Almost all of the variation in the equilibrium constants for LA binding was from differences in the association rate constants. These were consistently below the limit set by diffusion and were substantially different even though the agonists had similar sizes and the same charge. This suggests that binding to resting receptors is not by diffusion alone and, hence, that each binding site can undergo two conformational changes ("catch" and "hold") that connect three different structures (apo-, LA-bound, and HA-bound). Analyses of ACh-binding protein structures suggest that this binding site, too, may adopt three discrete structures having different degrees of loop C displacement ("capping"). For the agonists we tested, the logarithms of the equilibrium constants for LA binding and LA↔HA gating were correlated. Although agonist binding and channel gating have long been considered to be separate processes in the activation of ligand-gated ion channels, this correlation implies that the catch-and-hold conformational changes are energetically linked and together comprise an integrated process having a common structural basis. We propose that loop C capping mainly reflects agonist binding, with its two stages corresponding to the formation of the LA and HA complexes. The catch-and-hold reaction coordinate is discussed in terms of preopening states and thermodynamic cycles of activation.

An integrated catch-and-hold mechanism activates nicotinic acetylcholine receptors

PubMed Central

Jadey, Snehal

2012-01-01

In neuromuscular acetylcholine (ACh) receptor channels (AChRs), agonist molecules bind with a low affinity (LA) to two sites that can switch to high affinity (HA) and increase the probability of channel opening. We measured (by using single-channel kinetic analysis) the rate and equilibrium constants for LA binding and channel gating for several different agonists of adult-type mouse AChRs. Almost all of the variation in the equilibrium constants for LA binding was from differences in the association rate constants. These were consistently below the limit set by diffusion and were substantially different even though the agonists had similar sizes and the same charge. This suggests that binding to resting receptors is not by diffusion alone and, hence, that each binding site can undergo two conformational changes (“catch” and “hold”) that connect three different structures (apo-, LA-bound, and HA-bound). Analyses of ACh-binding protein structures suggest that this binding site, too, may adopt three discrete structures having different degrees of loop C displacement (“capping”). For the agonists we tested, the logarithms of the equilibrium constants for LA binding and LA↔HA gating were correlated. Although agonist binding and channel gating have long been considered to be separate processes in the activation of ligand-gated ion channels, this correlation implies that the catch-and-hold conformational changes are energetically linked and together comprise an integrated process having a common structural basis. We propose that loop C capping mainly reflects agonist binding, with its two stages corresponding to the formation of the LA and HA complexes. The catch-and-hold reaction coordinate is discussed in terms of preopening states and thermodynamic cycles of activation. PMID:22732309

Experimental Determination of pK[subscript a] Values by Use of NMR Chemical Shifts, Revisited

ERIC Educational Resources Information Center

Gift, Alan D.; Stewart, Sarah M.; Bokashanga, Patrick Kwete

2012-01-01

This laboratory experiment, using proton NMR spectroscopy to determine the dissociation constant for heterocyclic bases, has been modified from a previously described experiment. A solution of a substituted pyridine is prepared using deuterium oxide (D[subscript 2]O) as the solvent. The pH of the solution is adjusted and proton NMR spectra are…

Nuclear magnetic resonance studies of phosphorus(v) pesticides. Part I. Chemical shifts of protons as a means of identification of pesticides

USGS Publications Warehouse

Babad, H.; Herbert, W.; Goldberg, M.C.

1968-01-01

Correlations of structural and proton chemical-hift data for 40 commercial phosphorus(V) pesticides are reported. Correlations of structure with the phosphorus coupling constants are discussed, and general trends are noted which aid in the use of NMR as a tool for identification and analysis of phosphorus(V) compounds. ?? 1968.

A dye-binding assay for measurement of the binding of Cu(II) to proteins.

PubMed

Wilkinson-White, Lorna E; Easterbrook-Smith, Simon B

2008-10-01

We analysed the theory of the coupled equilibria between a metal ion, a metal ion-binding dye and a metal ion-binding protein in order to develop a procedure for estimating the apparent affinity constant of a metal ion:protein complex. This can be done by analysing from measurements of the change in the concentration of the metal ion:dye complex with variation in the concentration of either the metal ion or the protein. Using experimentally determined values for the affinity constant of Cu(II) for the dye, 2-(5-bromo-2-pyridylaxo)-5-(N-propyl-N-sulfopropylamino) aniline (5-Br-PSAA), this procedure was used to estimate the apparent affinity constants for formation of Cu(II):transthyretin, yielding values which were in agreement with literature values. An apparent affinity constant for Cu(II) binding to alpha-synuclein of approximately 1 x 10(9)M(-1) was obtained from measurements of tyrosine fluorescence quenching by Cu(II). This value was in good agreement with that obtained using 5-Br-PSAA. Our analysis and data therefore show that measurement of changes in the equilibria between Cu(II) and 5-Br-PSAA by Cu(II)-binding proteins provides a general procedure for estimating the affinities of proteins for Cu(II).

Evaluation of the rate constants of sugar transport through maltoporin (LamB) of Escherichia coli from the sugar-induced current noise

PubMed Central

1995-01-01

LamB (maltoporin) of Escherichia coli outer membrane was reconstituted into artificial lipid bilayer membranes. The channel contains a binding site for sugars and is blocked for ions when the site is occupied by a sugar. The on and off reactions of sugar binding cause an increase of the noise of the current through the channel. The sugar-induced current noise of maltoporin was used for the evaluation of the sugar-binding kinetics for different sugars of the maltooligosaccharide series and for sucrose. The on rate constant for sugar binding was between 10(6) and 10(7) M-1.s-1 for the maltooligosaccharides and corresponds to the movement of the sugars from the aqueous phase to the central binding site. The off rate (corresponding to the release of the sugars from the channel) decreased with increasing number of glucose residues in the maltooligosaccharides from approximately 2,000 s-1 for maltotriose to 180 s-1 for maltoheptaose. The kinetics for sucrose movement was considerably slower. The activation energies of the stability constant and of the rate constants for sugar binding were evaluated from noise experiments at different temperatures. The role of LamB in the transport of maltooligosaccharides across the outer membrane is discussed. PMID:7539481

Copper complexes containing thiosemicarbazones derived from 6-nitropiperonal: Antimicrobial and biophysical properties

NASA Astrophysics Data System (ADS)

Beckford, Floyd A.; Webb, Kelsey R.

2017-08-01

A series of four thiosemicarbazones from 6-nitropiperonal along with the corresponding copper complexes were synthesized. The biophysical characteristics of the complexes were investigated by the binding to DNA and human serum albumin. The binding to DNA is moderate; the binding constants run from (0.49-7.50) × 104 M- 1. In relation to HSA, the complexes interact strongly with binding constants on the order of 105 M- 1. The complexes also display antioxidant behavior as determined by the ability to scavenge diphenylpicrylhydrazyl (dpph) and nitric oxide radicals. The antimicrobial profiles of the compounds, tested against a panel of microbes including five of the ESKAPE pathogens (Staphylococcus aureus, MRSA, Escherichia coli, Klebsiella pneumoniae, MDR, Acinetobacter baumannii, Pseudomonas aeruginosa) and two yeasts (Candida albicans and Cryptococcus neoformans var. grubii), are also described. The compounds contain a core moiety that is similar to oxolinic acid, a quinolone antibiotic that targets DNA gyrase and topoisomerase (IV). The binding interaction between the complexes and these important antibacterial targets were studied by computational methods, chiefly docking studies. The calculated dissociation constants for the interaction with DNA gyrase B (from Staphylococcus aureus) range from 4.32 to 24.65 μM; the binding was much stronger to topoisomerase IV, with dissociation constants ranging from 0.37 to 1.27 μM.

Effects of pH on the association between the inhibitor cystatin and the proteinase chymopapain.

PubMed

Reyes-Espinosa, Francisco; Arroyo-Reyna, Alfonso; Garcia-Gutierrez, Ponciano; Serratos, Iris N; Zubillaga, Rafael A

2014-01-01

Cysteine proteinases are involved in many aspects of physiological regulation. In humans, some cathepsins have shown another function in addition to their role as lysosomal proteases in intracellular protein degradation; they have been implicated in the pathogenesis of several heart and blood vessel diseases and in cancer development. In this work, we present a fluorometric and computational study of the binding of one representative plant cysteine proteinase, chymopapain, to one of the most studied inhibitors of these proteinases: chicken cystatin. The binding equilibrium constant, Kb, was determined in the pH range between 3.5 and 10.0, revealing a maximum in the affinity at pH 9.0. We constructed an atomic model for the chymopapain-cystatin dimer by docking the individual 3D protein structures; subsequently, the model was refined using a 100 ns NPT molecular dynamics simulation in explicit water. Upon scrutiny of this model, we identified 14 ionizing residues at the interface of the complex using a cutoff distance of 5.0 Å. Using the pKa values predicted with PROPKA and a modified proton-linkage model, we performed a regression analysis on our data to obtain the composite pKavalues for three isoacidic residues. We also calculated the electrostatic component of the binding energy (ΔGb,elec) at different pH values using an implicit solvent model and APBS software. The pH profile of this calculated energy compares well with the experimentally obtained binding energy, ΔGb. We propose that the residues that form an interchain ionic pair, Lys139A from chymopapain and Glu19B from cystatin, as well as Tyr61A and Tyr67A from chymopapain are the main residues responsible for the observed pH dependence in the chymopapain- cystatin affinity.

Detecting Low-Contrast Features in the Cosmic Ray Albedo Proton Map of the Moon

NASA Technical Reports Server (NTRS)

Wilson, J. K.; Schwadron, N.; Spence, H. E.; Golightly, M. J.; Case, A. W.; Smith, S.; Blake, J. B.; Kasper, J.; Looper, M. D.; Mazur, J. E.;

Water and tissue equivalence properties of biological materials for photons, electrons, protons and alpha particles in the energy region 10 keV-1 GeV: a comparative study.

PubMed

Kurudirek, Murat

2016-09-01

To compare some biological materials in respect to the water and tissue equivalence properties for photon, electron, proton and alpha particle interactions as means of the effective atomic number (Zeff) and electron density (Ne). A Z-wise interpolation procedure has been adopted for calculation of Zeff using the mass attenuation coefficients for photons and the mass stopping powers for charged particles. At relatively low energies (100 keV-3 MeV), Zeff and Ne for photons and electrons were found to be constant while they vary much more for protons and alpha particles. In contrast, Zeff and Ne for protons and alpha particles were found to be constant after 3 MeV whereas for photons and electrons they were found to increase with the increasing energy. Also, muscle eq. liquid (with sucrose) have Zeff and Ne values close to the Muscle Skeletal (ICRP) and Muscle Striated (ICRU) within low relative differences below 9%. Muscle eq. liquid (without sucrose) have Zeff and Ne values close to the Muscle Skeletal (ICRP) and Muscle Striated (ICRU) with difference below 10%. The reported data should be useful in determining best water as well as tissue equivalent materials for photon, electron, proton and alpha particle interactions.

Kinetic analysis of cooperative interactions induced by Mn2+ binding to the chloroplast H(+)-ATPase.

PubMed

Hiller, R; Carmeli, C

1990-07-03

The kinetics of Mn2+ binding to three cooperatively interacting sites in chloroplast H(+)-ATPase (CF1) were measured by EPR following rapid mixing of the enzyme with MnCl2 with a time resolution of 8 ms. Mixing of the enzyme-bound Mn2+ with MgCl2 gave a measure of the rate of exchange. The data could be best fitted to a kinetic model assuming three sequential, positively cooperative binding sites. (1) In the latent CF1, the binding to all three sites had a similar on-rate constants of (1.1 +/- 0.04) X 10(4) M-1s-1. (2) Site segregation was found in the release of ions with off-rate constants of 0.69 +/- 0.04 s-1 for the first two and 0.055 +/- 0.003 s-1 for the third. (3) Addition of one ADP per CF1 caused a decrease in the off-rate constants to 0.31 +/- 0.02 and 0.033 +/- 0.008 s-1 for the first two and the third sites, respectively. (4) Heat activation of CF1 increased the on-rate constant to (4.2 +/- 0.92) X 10(4) M-1s-1 and the off-rate constants of the first two and the third site to 1.34 +/- 0.08 and 0.16 +/- 0.07 s-1, respectively. (5) The calculated thermodynamic dissociation constants were similar to those previously obtained from equilibrium binding studies. These findings were correlated to the rate constants obtained from studies of the catalysis and regulation of the H(+)-ATPase. The data support the suggestion that regulation induces sequential progress of catalysis through the three active sites of the enzyme.

Analysis of cholera toxin-ganglioside interactions by flow cytometry.

PubMed

Lauer, Sabine; Goldstein, Byron; Nolan, Rhiannon L; Nolan, John P

2002-02-12

Cholera toxin entry into mammalian cells is mediated by binding of the pentameric B subunit (CTB) to ganglioside GM(1) in the cell membrane. We used flow cytometry to quantitatively measure in real time the interactions of fluorescently labeled pentameric cholera toxin B-subunit (FITC-CTB) with its ganglioside receptor on microsphere-supported phospholipid membranes. A model that describes the multiple steps of this mode of recognition was developed to guide our flow cytometric experiments and extract relevant equilibrium and kinetic rate constants. In contrast to previous studies, our approach takes into account receptor cross-linking, an important feature for multivalent interactions. From equilibrium measurements, we determined an equilibrium binding constant for a single subunit of FITC-CTB binding monovalently to GM(1) presented in bilayers of approximately 8 x 10(7) M(-1) while that for binding to soluble GM(1)-pentasaccharide was found to be approximately 4 x 10(6) M(-1). From kinetic measurements, we determined the rate constant for dissociation of a single site of FITC-CTB from microsphere-supported bilayers to be (3.21 +/- 0.03) x 10(-3) s(-1), and the rate of association of a site on FITC-CTB in solution to a GM(1) in the bilayer to be (2.8 +/- 0.4) x 10(4) M(-1) s(-1). These values yield a lower estimate for the equilibrium binding constant of approximately 1 x 10(7) M(-1). We determined the equilibrium surface cross-linking constant [(1.1 +/- 0.1) x 10(-12) cm(2)] and from this value and the value for the rate constant for dissociation derived a value of approximately 3.5 x 10(-15) cm(2) s(-1) for the forward rate constant for cross-linking. We also compared the interaction of the receptor binding B-subunit with that of the whole toxin (A- and B-subunits). Our results show that the whole toxin binds with approximately 100-fold higher avidity than the pentameric B-subunit alone which is most likely due to the additional interaction of the A(2)-subunit with the membrane surface. Interaction of cholera toxin B-subunit and whole cholera toxin with gangliosides other than GM(1) revealed specific binding only to GD1(b) and asialo-GM(1). These interactions, however, are marked by low avidity and require high receptor concentrations to be observed.

Analysis by metadynamics simulation of binding pathway of influenza virus M2 channel blockers.

PubMed

Sakai, Yuri; Kawaguchi, Atsushi; Nagata, Kyosuke; Hirokawa, Takatsugu

2018-01-01

M2 protein of influenza A virus is a proton channel spanning the viral envelope. Activity of this proton channel is required for uncoating of viral particles and equilibrating the pH across the trans Golgi apparatus, which prevents conformational change in hemagglutinin. Amantadine, an anti-influenza A virus drug, inhibits M2 proton channel activity by binding to the channel pore; however, most currently circulating influenza A viruses are amantadine-resistant. The most prevalent resistant mutation is a substitution from Ser31 to Asn31 in M2. Further atomistic analysis of ligand-M2 complexes is needed to provide new approaches for the design of novel M2 channel blockers. Here, the free energy profiles of the binding kinetics of M2 channel blockers were examined by well-tempered metadynamics simulations and it was found that amantadine first binds to Asp24 of S31 M2 and forms a metastable conformation. In contrast, the free energy profiles of adamantyl bromothiophene dual inhibitor with either S31 M2 or N31 M2 are broad funnel-shaped curves, suggesting that adamantyl bromothiophene does not form metastable complexes with M2. The trajectory of well-tempered metadynamics simulations revealed that steric hindrance between adamantyl bromothiophene and S31 M2 interrupts formation of a metastable conformation at Asp24 and that a halogen bond between the bromine atom and N31 is responsible for pulling down the ligand to the channel pore of N31 M2 in the absence of a metastable state. Binding pathways of M2 channel blockers to M2 are here proposed on the basis of these findings; they may provide new approaches to designing further M2 channel blockers. © 2017 The Societies and John Wiley & Sons Australia, Ltd.

Ab initio study of the binding of Trichostatin A (TSA) in the active site of histone deacetylase like protein (HDLP).

PubMed

Vanommeslaeghe, Kenno; Van Alsenoy, Christian; De Proft, Frank; Martins, José C; Tourwé, Dirk; Geerlings, Paul

2003-08-21

Histone deacetylase (HDAC) inhibitors have recently attracted considerable interest because of their therapeutic potential for the treatment of cell proliferative diseases. An X-ray structure of a very potent inhibitor, Trichostatin A (TSA), bound to HDLP (an HDAC analogue isolated from Aquifex aeolicus), is available. From this structure, an active site model (322 atoms), relevant for the binding of TSA and structural analogues, has been derived, and TSA has been minimized in this active site at HF 3-21G* level. The resulting conformation is in excellent accordance with the X-ray structure, and indicates a deprotonation of the hydroxamic acid in TSA by His 131. Also, a water molecule was minimized in the active site. In addition to a similar deprotonation, in accordance with a possible catalytic mechanism of HDAC as proposed by Finnin et al. (M. S. Finnin, J. R. Donigian, A. Cohen, V. M. Richon, R. A. Rifkind and P. A. Marks, Nature, 1999, 401, 188-193), a displacement of the resulting OH- ion in the active site was observed. Based on these results, the difference in energy of binding between TSA and water was calculated. The resulting value is realistic in respect to experimental binding affinities. Furthermore, the mechanism of action of the His 131-Asp 166 charge relay system was investigated. Although the Asp residue in this motif is known to substantially increase the basicity of the His residue, no proton transfer from His 131 to Asp 166 was observed on binding of TSA or water. However, in the empty protonated active site, this proton transfer does occur.

Properties of the anion-binding site of pharaonis Halorhodopsin studied by ultrafast pump-probe spectroscopy and low-temperature FTIR spectroscopy.

PubMed

Nakashima, Keisuke; Nakamura, Takumi; Takeuchi, Satoshi; Shibata, Mikihiro; Demura, Makoto; Tahara, Tahei; Kandori, Hideki

2009-06-18

Halorhodopsin (HR) is a light-driven chloride pump. Cl(-) is bound in the Schiff base region of the retinal chromophore, and unidirectional Cl(-) transport is probably enforced by the specific hydrogen-bonding interaction with the protonated Schiff base and internal water molecules. It is known that HR from Natronobacterium pharaonis (pHR) also pumps NO(3)(-) with similar efficiency, suggesting that NO(3)(-) binds to the Cl(-)-binding site. In the present study, we investigated the properties of the anion-binding site by means of ultrafast pump-probe spectroscopy and low-temperature FTIR spectroscopy. The obtained data were surprisingly similar between pHR-NO(3)(-) and pHR-Cl(-), even though the shapes and sizes of the two anions are quite different. Femtosecond pump-probe spectroscopy showed very similar excited-state dynamics between pHR-NO(3)(-) and pHR-Cl(-). Low-temperature FTIR spectroscopy of unlabeled and [zeta-(15)N]Lys-labeled pHR revealed almost identical hydrogen-bonding strengths of the protonated retinal Schiff base between pHR-NO(3)(-) and pHR-Cl(-), which is similarly strengthened after retinal isomerization. There were spectral variations for water stretching vibrations between pHR-NO(3)(-) and pHR-Cl(-), suggesting that the water molecules hydrate each anion. Nevertheless, the overall spectral features were similar for the two species. These observations strongly suggest that the anion-binding site has a flexible structure and that the interaction between retinal and the anions is weak, despite the presence of an electrostatic interaction. Such a flexible hydrogen-bonding network in the Schiff base region in HR appears to be in remarkable contrast to that in light-driven proton-pumping proteins.

Proton irradiation study of GFR candidate ceramics

NASA Astrophysics Data System (ADS)

Gan, Jian; Yang, Yong; Dickson, Clayton; Allen, Todd

2009-06-01

This work investigated the microstructural response of SiC, ZrC and ZrN irradiated with 2.6 MeV protons at 800 °C to a fluence of 2.75 × 10 19 protons/cm 2, corresponding to 0.71-1.8 displacement per atom (dpa), depending on the material. The change of lattice constant evaluated using HOLZ patterns is not observed. In comparison to Kr ion irradiation at 800 °C to 10 dpa from the previous studies, the proton irradiated ZrC and ZrN at 1.8 dpa show less irradiation damage to the lattice structure. The proton irradiated ZrC exhibits faulted loops which are not observed in the Kr ion irradiated sample. ZrN shows the least microstructural change from proton irradiation. The microstructure of 6H-SiC irradiated to 0.71 dpa consists of black dot defects at high density.

Energetic basis for drug resistance of HIV-1 protease mutants against amprenavir

NASA Astrophysics Data System (ADS)

Kar, Parimal; Knecht, Volker

2012-02-01

Amprenavir (APV) is a high affinity (0.15 nM) HIV-1 protease (PR) inhibitor. However, the affinities of the drug resistant protease variants V32I, I50V, I54V, I54M, I84V and L90M to amprenavir are decreased 3 to 30-fold compared to the wild-type. In this work, the popular molecular mechanics Poisson-Boltzmann surface area method has been used to investigate the effectiveness of amprenavir against the wild-type and these mutated protease variants. Our results reveal that the protonation state of Asp25/Asp25' strongly affects the dynamics, the overall affinity and the interactions of the inhibitor with individual residues. We emphasize that, in contrast to what is often assumed, the protonation state may not be inferred from the affinities but requires pKa calculations. At neutral pH, Asp25 and Asp25' are ionized or protonated, respectively, as suggested from pKa calculations. This protonation state was thus mainly considered in our study. Mutation induced changes in binding affinities are in agreement with the experimental findings. The decomposition of the binding free energy reveals the mechanisms underlying binding and drug resistance. Drug resistance arises from an increase in the energetic contribution from the van der Waals interactions between APV and PR (V32I, I50V, and I84V mutant) or a rise in the energetic contribution from the electrostatic interactions between the inhibitor and its target (I54M and I54V mutant). For the V32I mutant, also an increased free energy for the polar solvation contributes to the drug resistance. For the L90M mutant, a rise in the van der Waals energy for APV-PR interactions is compensated by a decrease in the polar solvation free energy such that the net binding affinity remains unchanged. Detailed understanding of the molecular forces governing binding and drug resistance might assist in the design of new inhibitors against HIV-1 PR variants that are resistant against current drugs.

Simulation of weak polyelectrolytes: a comparison between the constant pH and the reaction ensemble method

NASA Astrophysics Data System (ADS)

Landsgesell, Jonas; Holm, Christian; Smiatek, Jens

2017-03-01

The reaction ensemble and the constant pH method are well-known chemical equilibrium approaches to simulate protonation and deprotonation reactions in classical molecular dynamics and Monte Carlo simulations. In this article, we demonstrate the similarity between both methods under certain conditions. We perform molecular dynamics simulations of a weak polyelectrolyte in order to compare the titration curves obtained by both approaches. Our findings reveal a good agreement between the methods when the reaction ensemble is used to sweep the reaction constant. Pronounced differences between the reaction ensemble and the constant pH method can be observed for stronger acids and bases in terms of adaptive pH values. These deviations are due to the presence of explicit protons in the reaction ensemble method which induce a screening of electrostatic interactions between the charged titrable groups of the polyelectrolyte. The outcomes of our simulation hint to a better applicability of the reaction ensemble method for systems in confined geometries and titrable groups in polyelectrolytes with different pKa values.

Measurement of the ratio of the inclusive 3-jet cross section to the inclusive 2-jet cross section in pp collisions at and first determination of the strong coupling constant in the TeV range

NASA Astrophysics Data System (ADS)

Chatrchyan, S.; Khachatryan, V.; Sirunyan, A. M.; Tumasyan, A.; Adam, W.; Bergauer, T.; Dragicevic, M.; Erö, J.; Fabjan, C.; Friedl, M.; Frühwirth, R.; Ghete, V. M.; Hörmann, N.; Hrubec, J.; Jeitler, M.; Kiesenhofer, W.; Knünz, V.; Krammer, M.; Krätschmer, I.; Liko, D.; Mikulec, I.; Rabady, D.; Rahbaran, B.; Rohringer, C.; Rohringer, H.; Schöfbeck, R.; Strauss, J.; Taurok, A.; Treberer-Treberspurg, W.; Waltenberger, W.; Wulz, C.-E.; Mossolov, V.; Shumeiko, N.; Suarez Gonzalez, J.; Alderweireldt, S.; Bansal, M.; Bansal, S.; Cornelis, T.; De Wolf, E. A.; Janssen, X.; Knutsson, A.; Luyckx, S.; Mucibello, L.; Ochesanu, S.; Roland, B.; Rougny, R.; Van Haevermaet, H.; Van Mechelen, P.; Van Remortel, N.; Van Spilbeeck, A.; Blekman, F.; Blyweert, S.; D'Hondt, J.; Kalogeropoulos, A.; Keaveney, J.; Maes, M.; Olbrechts, A.; Tavernier, S.; Van Doninck, W.; Van Mulders, P.; Van Onsem, G. P.; Villella, I.; Clerbaux, B.; De Lentdecker, G.; Favart, L.; Gay, A. P. R.; Hreus, T.; Léonard, A.; Marage, P. E.; Mohammadi, A.; Reis, T.; Thomas, L.; Vander Velde, C.; Vanlaer, P.; Wang, J.; Adler, V.; Beernaert, K.; Benucci, L.; Cimmino, A.; Costantini, S.; Dildick, S.; Garcia, G.; Klein, B.; Lellouch, J.; Marinov, A.; Mccartin, J.; Ocampo Rios, A. A.; Ryckbosch, D.; Sigamani, M.; Strobbe, N.; Thyssen, F.; Tytgat, M.; Walsh, S.; Yazgan, E.; Zaganidis, N.; Basegmez, S.; Beluffi, C.; Bruno, G.; Castello, R.; Caudron, A.; Ceard, L.; Delaere, C.; du Pree, T.; Favart, D.; Forthomme, L.; Giammanco, A.; Hollar, J.; Lemaitre, V.; Liao, J.; Militaru, O.; Nuttens, C.; Pagano, D.; Pin, A.; Piotrzkowski, K.; Popov, A.; Selvaggi, M.; Vizan Garcia, J. M.; Beliy, N.; Caebergs, T.; Daubie, E.; Hammad, G. H.; Alves, G. A.; Correa Martins Junior, M.; Martins, T.; Pol, M. E.; Souza, M. H. G.; Aldá Júnior, W. L.; Carvalho, W.; Chinellato, J.; Custódio, A.; Da Costa, E. M.; De Jesus Damiao, D.; De Oliveira Martins, C.; Fonseca De Souza, S.; Malbouisson, H.; Malek, M.; Matos Figueiredo, D.; Mundim, L.; Nogima, H.; Prado Da Silva, W. L.; Santoro, A.; Soares Jorge, L.; Sznajder, A.; Tonelli Manganote, E. J.; Vilela Pereira, A.; Anjos, T. S.; Bernardes, C. A.; Dias, F. A.; Fernandez Perez Tomei, T. R.; Gregores, E. M.; Lagana, C.; Marinho, F.; Mercadante, P. G.; Novaes, S. F.; Padula, Sandra S.; Genchev, V.; Iaydjiev, P.; Piperov, S.; Rodozov, M.; Stoykova, S.; Sultanov, G.; Tcholakov, V.; Trayanov, R.; Vutova, M.; Dimitrov, A.; Hadjiiska, R.; Kozhuharov, V.; Litov, L.; Pavlov, B.; Petkov, P.; Bian, J. G.; Chen, G. M.; Chen, H. S.; Jiang, C. H.; Liang, D.; Liang, S.; Meng, X.; Tao, J.; Wang, J.; Wang, X.; Wang, Z.; Xiao, H.; Xu, M.; Asawatangtrakuldee, C.; Ban, Y.; Guo, Y.; Li, Q.; Li, W.; Liu, S.; Mao, Y.; Qian, S. J.; Wang, D.; Zhang, L.; Zou, W.; Avila, C.; Carrillo Montoya, C. A.; Gomez, J. P.; Gomez Moreno, B.; Sanabria, J. C.; Godinovic, N.; Lelas, D.; Plestina, R.; Polic, D.; Puljak, I.; Antunovic, Z.; Kovac, M.; Brigljevic, V.; Duric, S.; Kadija, K.; Luetic, J.; Mekterovic, D.; Morovic, S.; Tikvica, L.; Attikis, A.; Mavromanolakis, G.; Mousa, J.; Nicolaou, C.; Ptochos, F.; Razis, P. A.; Finger, M.; Finger, M.; Assran, Y.; Ellithi Kamel, A.; Mahmoud, M. A.; Mahrous, A.; Radi, A.; Kadastik, M.; Müntel, M.; Murumaa, M.; Raidal, M.; Rebane, L.; Tiko, A.; Eerola, P.; Fedi, G.; Voutilainen, M.; Härkönen, J.; Karimäki, V.; Kinnunen, R.; Kortelainen, M. J.; Lampén, T.; Lassila-Perini, K.; Lehti, S.; Lindén, T.; Luukka, P.; Mäenpää, T.; Peltola, T.; Tuominen, E.; Tuominiemi, J.; Tuovinen, E.; Wendland, L.; Korpela, A.; Tuuva, T.; Besancon, M.; Choudhury, S.; Couderc, F.; Dejardin, M.; Denegri, D.; Fabbro, B.; Faure, J. L.; Ferri, F.; Ganjour, S.; Givernaud, A.; Gras, P.; Hamel de Monchenault, G.; Jarry, P.; Locci, E.; Malcles, J.; Millischer, L.; Nayak, A.; Rander, J.; Rosowsky, A.; Titov, M.; Baffioni, S.; Beaudette, F.; Benhabib, L.; Bianchini, L.; Bluj, M.; Busson, P.; Charlot, C.; Daci, N.; Dahms, T.; Dalchenko, M.; Dobrzynski, L.; Florent, A.; Granier de Cassagnac, R.; Haguenauer, M.; Miné, P.; Mironov, C.; Naranjo, I. N.; Nguyen, M.; Ochando, C.; Paganini, P.; Sabes, D.; Salerno, R.; Sirois, Y.; Veelken, C.; Zabi, A.; Agram, J.-L.; Andrea, J.; Bloch, D.; Bodin, D.; Brom, J.-M.; Chabert, E. C.; Collard, C.; Conte, E.; Drouhin, F.; Fontaine, J.-C.; Gelé, D.; Goerlach, U.; Goetzmann, C.; Juillot, P.; Le Bihan, A.-C.; Van Hove, P.; Beauceron, S.; Beaupere, N.; Boudoul, G.; Brochet, S.; Chasserat, J.; Chierici, R.; Contardo, D.; Depasse, P.; El Mamouni, H.; Fay, J.; Gascon, S.; Gouzevitch, M.; Ille, B.; Kurca, T.; Lethuillier, M.; Mirabito, L.; Perries, S.; Sgandurra, L.; Sordini, V.; Tschudi, Y.; Vander Donckt, M.; Verdier, P.; Viret, S.; Tsamalaidze, Z.; Autermann, C.; Beranek, S.; Calpas, B.; Edelhoff, M.; Feld, L.; Heracleous, N.; Hindrichs, O.; Klein, K.; Merz, J.; Ostapchuk, A.; Perieanu, A.; Raupach, F.; Sammet, J.; Schael, S.; Sprenger, D.; Weber, H.; Wittmer, B.; Zhukov, V.; Ata, M.; Caudron, J.; Dietz-Laursonn, E.; Duchardt, D.; Erdmann, M.; Fischer, R.; Güth, A.; Hebbeker, T.; Heidemann, C.; Hoepfner, K.; Klingebiel, D.; Kreuzer, P.; Merschmeyer, M.; Meyer, A.; Olschewski, M.; Padeken, K.; Papacz, P.; Pieta, H.; Reithler, H.; Schmitz, S. A.; Sonnenschein, L.; Steggemann, J.; Teyssier, D.; Thüer, S.; Weber, M.; Cherepanov, V.; Erdogan, Y.; Flügge, G.; Geenen, H.; Geisler, M.; Haj Ahmad, W.; Hoehle, F.; Kargoll, B.; Kress, T.; Kuessel, Y.; Lingemann, J.; Nowack, A.; Nugent, I. M.; Perchalla, L.; Pooth, O.; Stahl, A.; Aldaya Martin, M.; Asin, I.; Bartosik, N.; Behr, J.; Behrenhoff, W.; Behrens, U.; Bergholz, M.; Bethani, A.; Borras, K.; Burgmeier, A.; Cakir, A.; Calligaris, L.; Campbell, A.; Costanza, F.; Diez Pardos, C.; Dorland, T.; Eckerlin, G.; Eckstein, D.; Flucke, G.; Geiser, A.; Glushkov, I.; Gunnellini, P.; Habib, S.; Hauk, J.; Hellwig, G.; Jung, H.; Kasemann, M.; Katsas, P.; Kleinwort, C.; Kluge, H.; Krämer, M.; Krücker, D.; Kuznetsova, E.; Lange, W.; Leonard, J.; Lipka, K.; Lohmann, W.; Lutz, B.; Mankel, R.; Marfin, I.; Melzer-Pellmann, I.-A.; Meyer, A. B.; Mnich, J.; Mussgiller, A.; Naumann-Emme, S.; Novgorodova, O.; Nowak, F.; Olzem, J.; Perrey, H.; Petrukhin, A.; Pitzl, D.; Raspereza, A.; Ribeiro Cipriano, P. M.; Riedl, C.; Ron, E.; Salfeld-Nebgen, J.; Schmidt, R.; Schoerner-Sadenius, T.; Sen, N.; Stein, M.; Walsh, R.; Wissing, C.; Blobel, V.; Enderle, H.; Erfle, J.; Gebbert, U.; Görner, M.; Gosselink, M.; Haller, J.; Heine, K.; Höing, R. S.; Kaussen, G.; Kirschenmann, H.; Klanner, R.; Lange, J.; Peiffer, T.; Pietsch, N.; Rathjens, D.; Sander, C.; Schettler, H.; Schleper, P.; Schlieckau, E.; Schmidt, A.; Schröder, M.; Schum, T.; Seidel, M.; Sibille, J.; Sola, V.; Stadie, H.; Steinbrück, G.; Thomsen, J.; Vanelderen, L.; Barth, C.; Baus, C.; Berger, J.; Böser, C.; Chwalek, T.; De Boer, W.; Descroix, A.; Dierlamm, A.; Feindt, M.; Guthoff, M.; Hackstein, C.; Hartmann, F.; Hauth, T.; Heinrich, M.; Held, H.; Hoffmann, K. H.; Husemann, U.; Katkov, I.; Komaragiri, J. R.; Kornmayer, A.; Lobelle Pardo, P.; Martschei, D.; Mueller, S.; Müller, Th.; Niegel, M.; Nürnberg, A.; Oberst, O.; Ott, J.; Quast, G.; Rabbertz, K.; Ratnikov, F.; Ratnikova, N.; Röcker, S.; Schilling, F.-P.; Schott, G.; Simonis, H. J.; Stober, F. M.; Troendle, D.; Ulrich, R.; Wagner-Kuhr, J.; Wayand, S.; Weiler, T.; Zeise, M.; Anagnostou, G.; Daskalakis, G.; Geralis, T.; Kesisoglou, S.; Kyriakis, A.; Loukas, D.; Markou, A.; Markou, C.; Ntomari, E.; Gouskos, L.; Mertzimekis, T. J.; Panagiotou, A.; Saoulidou, N.; Stiliaris, E.; Aslanoglou, X.; Evangelou, I.; Flouris, G.; Foudas, C.; Kokkas, P.; Manthos, N.; Papadopoulos, I.; Paradas, E.; Bencze, G.; Hajdu, C.; Hidas, P.; Horvath, D.; Radics, B.; Sikler, F.; Veszpremi, V.; Vesztergombi, G.; Zsigmond, A. J.; Beni, N.; Czellar, S.; Molnar, J.; Palinkas, J.; Szillasi, Z.; Karancsi, J.; Raics, P.; Trocsanyi, Z. L.; Ujvari, B.; Beri, S. B.; Bhatnagar, V.; Dhingra, N.; Gupta, R.; Kaur, M.; Mehta, M. Z.; Mittal, M.; Nishu, N.; Saini, L. K.; Sharma, A.; Singh, J. B.; Kumar, Ashok; Kumar, Arun; Ahuja, S.; Bhardwaj, A.; Choudhary, B. C.; Malhotra, S.; Naimuddin, M.; Ranjan, K.; Saxena, P.; Sharma, V.; Shivpuri, R. K.; Banerjee, S.; Bhattacharya, S.; Chatterjee, K.; Dutta, S.; Gomber, B.; Jain, Sa.; Jain, Sh.; Khurana, R.; Modak, A.; Mukherjee, S.; Roy, D.; Sarkar, S.; Sharan, M.; Abdulsalam, A.; Dutta, D.; Kailas, S.; Kumar, V.; Mohanty, A. K.; Pant, L. M.; Shukla, P.; Topkar, A.; Aziz, T.; Chatterjee, R. M.; Ganguly, S.; Guchait, M.; Gurtu, A.; Maity, M.; Majumder, G.; Mazumdar, K.; Mohanty, G. B.; Parida, B.; Sudhakar, K.; Wickramage, N.; Banerjee, S.; Dugad, S.; Arfaei, H.; Bakhshiansohi, H.; Etesami, S. M.; Fahim, A.; Hesari, H.; Jafari, A.; Khakzad, M.; Mohammadi Najafabadi, M.; Paktinat Mehdiabadi, S.; Safarzadeh, B.; Zeinali, M.; Grunewald, M.; Abbrescia, M.; Barbone, L.; Calabria, C.; Chhibra, S. S.; Colaleo, A.; Creanza, D.; De Filippis, N.; De Palma, M.; Fiore, L.; Iaselli, G.; Maggi, G.; Maggi, M.; Marangelli, B.; My, S.; Nuzzo, S.; Pacifico, N.; Pompili, A.; Pugliese, G.; Selvaggi, G.; Silvestris, L.; Singh, G.; Venditti, R.; Verwilligen, P.; Zito, G.; Abbiendi, G.; Benvenuti, A. C.; Bonacorsi, D.; Braibant-Giacomelli, S.; Brigliadori, L.; Campanini, R.; Capiluppi, P.; Castro, A.; Cavallo, F. R.; Cuffiani, M.; Dallavalle, G. M.; Fabbri, F.; Fanfani, A.; Fasanella, D.; Giacomelli, P.; Grandi, C.; Guiducci, L.; Marcellini, S.; Masetti, G.; Meneghelli, M.; Montanari, A.; Navarria, F. L.; Odorici, F.; Perrotta, A.; Primavera, F.; Rossi, A. M.; Rovelli, T.; Siroli, G. P.; Tosi, N.; Travaglini, R.; Albergo, S.; Chiorboli, M.; Costa, S.; Potenza, R.; Tricomi, A.; Tuve, C.; Barbagli, G.; Ciulli, V.; Civinini, C.; D'Alessandro, R.; Focardi, E.; Frosali, S.; Gallo, E.; Gonzi, S.; Lenzi, P.; Meschini, M.; Paoletti, S.; Sguazzoni, G.; Tropiano, A.; Benussi, L.; Bianco, S.; Fabbri, F.; Piccolo, D.; Fabbricatore, P.; Musenich, R.; Tosi, S.; Benaglia, A.; De Guio, F.; Di Matteo, L.; Fiorendi, S.; Gennai, S.; Ghezzi, A.; Govoni, P.; Lucchini, M. T.; Malvezzi, S.; Manzoni, R. A.; Martelli, A.; Massironi, A.; Menasce, D.; Moroni, L.; Paganoni, M.; Pedrini, D.; Ragazzi, S.; Redaelli, N.; Tabarelli de Fatis, T.; Buontempo, S.; Cavallo, N.; De Cosa, A.; Fabozzi, F.; Iorio, A. O. M.; Lista, L.; Meola, S.; Merola, M.; Paolucci, P.; Azzi, P.; Bacchetta, N.; Bellan, P.; Biasotto, M.; Bisello, D.; Branca, A.; Carlin, R.; Checchia, P.; Dorigo, T.; Fanzago, F.; Galanti, M.; Gasparini, F.; Gasparini, U.; Giubilato, P.; Gonella, F.; Gozzelino, A.; Kanishchev, K.; Lacaprara, S.; Lazzizzera, I.; Margoni, M.; Meneguzzo, A. T.; Montecassiano, F.; Pazzini, J.; Pozzobon, N.; Ronchese, P.; Sgaravatto, M.; Simonetto, F.; Torassa, E.; Tosi, M.; Zotto, P.; Gabusi, M.; Ratti, S. P.; Riccardi, C.; Vitulo, P.; Biasini, M.; Bilei, G. M.; Fanò, L.; Lariccia, P.; Mantovani, G.; Menichelli, M.; Nappi, A.; Romeo, F.; Saha, A.; Santocchia, A.; Spiezia, A.; Androsov, K.; Azzurri, P.; Bagliesi, G.; Boccali, T.; Broccolo, G.; Castaldi, R.; D'Agnolo, R. T.; Dell'Orso, R.; Fiori, F.; Foà, L.; Giassi, A.; Kraan, A.; Ligabue, F.; Lomtadze, T.; Martini, L.; Messineo, A.; Palla, F.; Rizzi, A.; Serban, A. T.; Spagnolo, P.; Squillacioti, P.; Tenchini, R.; Tonelli, G.; Venturi, A.; Verdini, P. G.; Vernieri, C.; Barone, L.; Cavallari, F.; Del Re, D.; Diemoz, M.; Fanelli, C.; Grassi, M.; Longo, E.; Margaroli, F.; Meridiani, P.; Micheli, F.; Nourbakhsh, S.; Organtini, G.; Paramatti, R.; Rahatlou, S.; Soffi, L.; Amapane, N.; Arcidiacono, R.; Argiro, S.; Arneodo, M.; Biino, C.; Cartiglia, N.; Casasso, S.; Costa, M.; Dellacasa, G.; Demaria, N.; Mariotti, C.; Maselli, S.; Migliore, E.; Monaco, V.; Musich, M.; Obertino, M. M.; Pastrone, N.; Pelliccioni, M.; Potenza, A.; Romero, A.; Ruspa, M.; Sacchi, R.; Solano, A.; Staiano, A.; Tamponi, U.; Belforte, S.; Candelise, V.; Casarsa, M.; Cossutti, F.; Della Ricca, G.; Gobbo, B.; La Licata, C.; Marone, M.; Montanino, D.; Penzo, A.; Schizzi, A.; Zanetti, A.; Kim, T. Y.; Nam, S. K.; Chang, S.; Kim, D. H.; Kim, G. N.; Kim, J. E.; Kong, D. J.; Oh, Y. D.; Park, H.; Son, D. C.; Kim, J. Y.; Kim, Zero J.; Song, S.; Choi, S.; Gyun, D.; Hong, B.; Jo, M.; Kim, H.; Kim, T. J.; Lee, K. S.; Park, S. K.; Roh, Y.; Choi, M.; Kim, J. H.; Park, C.; Park, I. C.; Park, S.; Ryu, G.; Choi, Y.; Choi, Y. K.; Goh, J.; Kim, M. S.; Kwon, E.; Lee, B.; Lee, J.; Lee, S.; Seo, H.; Yu, I.; Grigelionis, I.; Juodagalvis, A.; Castilla-Valdez, H.; De La Cruz-Burelo, E.; Heredia-de La Cruz, I.; Lopez-Fernandez, R.; Martínez-Ortega, J.; Sanchez-Hernandez, A.; Villasenor-Cendejas, L. M.; Carrillo Moreno, S.; Vazquez Valencia, F.; Salazar Ibarguen, H. A.; Casimiro Linares, E.; Morelos Pineda, A.; Reyes-Santos, M. A.; Krofcheck, D.; Bell, A. J.; Butler, P. H.; Doesburg, R.; Reucroft, S.; Silverwood, H.; Ahmad, M.; Asghar, M. I.; Butt, J.; Hoorani, H. R.; Khalid, S.; Khan, W. A.; Khurshid, T.; Qazi, S.; Shah, M. A.; Shoaib, M.; Bialkowska, H.; Boimska, B.; Frueboes, T.; Górski, M.; Kazana, M.; Nawrocki, K.; Romanowska-Rybinska, K.; Szleper, M.; Wrochna, G.; Zalewski, P.; Brona, G.; Bunkowski, K.; Cwiok, M.; Dominik, W.; Doroba, K.; Kalinowski, A.; Konecki, M.; Krolikowski, J.; Misiura, M.; Wolszczak, W.; Almeida, N.; Bargassa, P.; David, A.; Faccioli, P.; Ferreira Parracho, P. G.; Gallinaro, M.; Rodrigues Antunes, J.; Seixas, J.; Varela, J.; Vischia, P.; Bunin, P.; Gavrilenko, M.; Golutvin, I.; Gorbunov, I.; Kamenev, A.; Karjavin, V.; Konoplyanikov, V.; Kozlov, G.; Lanev, A.; Malakhov, A.; Matveev, V.; Moisenz, P.; Palichik, V.; Perelygin, V.; Shmatov, S.; Skatchkov, N.; Smirnov, V.; Zarubin, A.; Evstyukhin, S.; Golovtsov, V.; Ivanov, Y.; Kim, V.; Levchenko, P.; Murzin, V.; Oreshkin, V.; Smirnov, I.; Sulimov, V.; Uvarov, L.; Vavilov, S.; Vorobyev, A.; Vorobyev, An.; Andreev, Yu.; Dermenev, A.; Gninenko, S.; Golubev, N.; Kirsanov, M.; Krasnikov, N.; Pashenkov, A.; Tlisov, D.; Toropin, A.; Epshteyn, V.; Erofeeva, M.; Gavrilov, V.; Lychkovskaya, N.; Popov, V.; Safronov, G.; Semenov, S.; Spiridonov, A.; Stolin, V.; Vlasov, E.; Zhokin, A.; Andreev, V.; Azarkin, M.; Dremin, I.; Kirakosyan, M.; Leonidov, A.; Mesyats, G.; Rusakov, S. V.; Vinogradov, A.; Belyaev, A.; Boos, E.; Dubinin, M.; Dudko, L.; Ershov, A.; Gribushin, A.; Klyukhin, V.; Kodolova, O.; Lokhtin, I.; Markina, A.; Obraztsov, S.; Petrushanko, S.; Savrin, V.; Snigirev, A.; Azhgirey, I.; Bayshev, I.; Bitioukov, S.; Kachanov, V.; Kalinin, A.; Konstantinov, D.; Krychkine, V.; Petrov, V.; Ryutin, R.; Sobol, A.; Tourtchanovitch, L.; Troshin, S.; Tyurin, N.; Uzunian, A.; Volkov, A.; Adzic, P.; Ekmedzic, M.; Krpic, D.; Milosevic, J.; Aguilar-Benitez, M.; Alcaraz Maestre, J.; Battilana, C.; Calvo, E.; Cerrada, M.; Chamizo Llatas, M.; Colino, N.; De La Cruz, B.; Delgado Peris, A.; Domínguez Vázquez, D.; Fernandez Bedoya, C.; Fernández Ramos, J. P.; Ferrando, A.; Flix, J.; Fouz, M. C.; Garcia-Abia, P.; Gonzalez Lopez, O.; Goy Lopez, S.; Hernandez, J. M.; Josa, M. I.; Merino, G.; Navarro De Martino, E.; Puerta Pelayo, J.; Quintario Olmeda, A.; Redondo, I.; Romero, L.; Santaolalla, J.; Soares, M. S.; Willmott, C.; Albajar, C.; de Trocóniz, J. F.; Brun, H.; Cuevas, J.; Fernandez Menendez, J.; Folgueras, S.; Gonzalez Caballero, I.; Lloret Iglesias, L.; Piedra Gomez, J.; Brochero Cifuentes, J. A.; Cabrillo, I. J.; Calderon, A.; Chuang, S. H.; Duarte Campderros, J.; Fernandez, M.; Gomez, G.; Gonzalez Sanchez, J.; Graziano, A.; Jorda, C.; Lopez Virto, A.; Marco, J.; Marco, R.; Martinez Rivero, C.; Matorras, F.; Munoz Sanchez, F. J.; Rodrigo, T.; Rodríguez-Marrero, A. Y.; Ruiz-Jimeno, A.; Scodellaro, L.; Vila, I.; Vilar Cortabitarte, R.; Abbaneo, D.; Auffray, E.; Auzinger, G.; Bachtis, M.; Baillon, P.; Ball, A. H.; Barney, D.; Bendavid, J.; Benitez, J. F.; Bernet, C.; Bianchi, G.; Bloch, P.; Bocci, A.; Bonato, A.; Bondu, O.; Botta, C.; Breuker, H.; Camporesi, T.; Cerminara, G.; Christiansen, T.; Coarasa Perez, J. A.; Colafranceschi, S.; d'Enterria, D.; Dabrowski, A.; De Roeck, A.; De Visscher, S.; Di Guida, S.; Dobson, M.; Dupont-Sagorin, N.; Elliott-Peisert, A.; Eugster, J.; Funk, W.; Georgiou, G.; Giffels, M.; Gigi, D.; Gill, K.; Giordano, D.; Girone, M.; Giunta, M.; Glege, F.; Gomez-Reino Garrido, R.; Gowdy, S.; Guida, R.; Hammer, J.; Hansen, M.; Harris, P.; Hartl, C.; Hegner, B.; Hinzmann, A.; Innocente, V.; Janot, P.; Kaadze, K.; Karavakis, E.; Kousouris, K.; Krajczar, K.; Lecoq, P.; Lee, Y.-J.; Lourenço, C.; Magini, N.; Malberti, M.; Malgeri, L.; Mannelli, M.; Masetti, L.; Meijers, F.; Mersi, S.; Meschi, E.; Moser, R.; Mulders, M.; Musella, P.; Nesvold, E.; Orsini, L.; Palencia Cortezon, E.; Perez, E.; Perrozzi, L.; Petrilli, A.; Pfeiffer, A.; Pierini, M.; Pimiä, M.; Piparo, D.; Polese, G.; Quertenmont, L.; Racz, A.; Reece, W.; Rojo, J.; Rolandi, G.; Rovelli, C.; Rovere, M.; Sakulin, H.; Santanastasio, F.; Schäfer, C.; Schwick, C.; Segoni, I.; Sekmen, S.; Sharma, A.; Siegrist, P.; Silva, P.; Simon, M.; Sphicas, P.; Spiga, D.; Stoye, M.; Tsirou, A.; Veres, G. I.; Vlimant, J. R.; Wöhri, H. K.; Worm, S. D.; Zeuner, W. D.; Bertl, W.; Deiters, K.; Erdmann, W.; Gabathuler, K.; Horisberger, R.; Ingram, Q.; Kaestli, H. C.; König, S.; Kotlinski, D.; Langenegger, U.; Meier, F.; Renker, D.; Rohe, T.; Bachmair, F.; Bäni, L.; Bortignon, P.; Buchmann, M. A.; Casal, B.; Chanon, N.; Deisher, A.; Dissertori, G.; Dittmar, M.; Donegà, M.; Dünser, M.; Eller, P.; Grab, C.; Hits, D.; Lecomte, P.; Lustermann, W.; Marini, A. C.; Martinez Ruiz del Arbol, P.; Mohr, N.; Moortgat, F.; Nägeli, C.; Nef, P.; Nessi-Tedaldi, F.; Pandolfi, F.; Pape, L.; Pauss, F.; Peruzzi, M.; Ronga, F. J.; Rossini, M.; Sala, L.; Sanchez, A. K.; Starodumov, A.; Stieger, B.; Takahashi, M.; Tauscher, L.; Thea, A.; Theofilatos, K.; Treille, D.; Urscheler, C.; Wallny, R.; Weber, H. A.; Amsler, C.; Chiochia, V.; Favaro, C.; Ivova Rikova, M.; Kilminster, B.; Millan Mejias, B.; Otiougova, P.; Robmann, P.; Snoek, H.; Taroni, S.; Tupputi, S.; Verzetti, M.; Cardaci, M.; Chen, K. H.; Ferro, C.; Kuo, C. M.; Li, S. W.; Lin, W.; Lu, Y. J.; Volpe, R.; Yu, S. S.; Bartalini, P.; Chang, P.; Chang, Y. H.; Chang, Y. W.; Chao, Y.; Chen, K. F.; Dietz, C.; Grundler, U.; Hou, W.-S.; Hsiung, Y.; Kao, K. Y.; Lei, Y. J.; Lu, R.-S.; Majumder, D.; Petrakou, E.; Shi, X.; Shiu, J. G.; Tzeng, Y. M.; Wang, M.; Asavapibhop, B.; Suwonjandee, N.; Adiguzel, A.; Bakirci, M. N.; Cerci, S.; Dozen, C.; Dumanoglu, I.; Eskut, E.; Girgis, S.; Gokbulut, G.; Gurpinar, E.; Hos, I.; Kangal, E. E.; Kayis Topaksu, A.; Onengut, G.; Ozdemir, K.; Ozturk, S.; Polatoz, A.; Sogut, K.; Sunar Cerci, D.; Tali, B.; Topakli, H.; Vergili, M.; Akin, I. V.; Aliev, T.; Bilin, B.; Bilmis, S.; Deniz, M.; Gamsizkan, H.; Guler, A. M.; Karapinar, G.; Ocalan, K.; Ozpineci, A.; Serin, M.; Sever, R.; Surat, U. E.; Yalvac, M.; Zeyrek, M.; Gülmez, E.; Isildak, B.; Kaya, M.; Kaya, O.; Ozkorucuklu, S.; Sonmez, N.; Bahtiyar, H.; Barlas, E.; Cankocak, K.; Günaydin, Y. O.; Vardarlı, F. I.; Yücel, M.; Levchuk, L.; Sorokin, P.; Brooke, J. J.; Clement, E.; Cussans, D.; Flacher, H.; Frazier, R.; Goldstein, J.; Grimes, M.; Heath, G. P.; Heath, H. F.; Kreczko, L.; Metson, S.; Newbold, D. M.; Nirunpong, K.; Poll, A.; Senkin, S.; Smith, V. J.; Williams, T.; Basso, L.; Bell, K. W.; Belyaev, A.; Brew, C.; Brown, R. M.; Cockerill, D. J. A.; Coughlan, J. A.; Harder, K.; Harper, S.; Jackson, J.; Olaiya, E.; Petyt, D.; Radburn-Smith, B. C.; Shepherd-Themistocleous, C. H.; Tomalin, I. R.; Womersley, W. J.; Bainbridge, R.; Buchmuller, O.; Burton, D.; Colling, D.; Cripps, N.; Cutajar, M.; Dauncey, P.; Davies, G.; Della Negra, M.; Ferguson, W.; Fulcher, J.; Futyan, D.; Gilbert, A.; Guneratne Bryer, A.; Hall, G.; Hatherell, Z.; Hays, J.; Iles, G.; Jarvis, M.; Karapostoli, G.; Kenzie, M.; Lane, R.; Lucas, R.; Lyons, L.; Magnan, A.-M.; Marrouche, J.; Mathias, B.; Nandi, R.; Nash, J.; Nikitenko, A.; Pela, J.; Pesaresi, M.; Petridis, K.; Pioppi, M.; Raymond, D. M.; Rogerson, S.; Rose, A.; Seez, C.; Sharp, P.; Sparrow, A.; Tapper, A.; Vazquez Acosta, M.; Virdee, T.; Wakefield, S.; Wardle, N.; Whyntie, T.; Chadwick, M.; Cole, J. E.; Hobson, P. R.; Khan, A.; Kyberd, P.; Leggat, D.; Leslie, D.; Martin, W.; Reid, I. D.; Symonds, P.; Teodorescu, L.; Turner, M.; Dittmann, J.; Hatakeyama, K.; Kasmi, A.; Liu, H.; Scarborough, T.; Charaf, O.; Cooper, S. I.; Henderson, C.; Rumerio, P.; Avetisyan, A.; Bose, T.; Fantasia, C.; Heister, A.; Lawson, P.; Lazic, D.; Rohlf, J.; Sperka, D.; John, J. St.; Sulak, L.; Alimena, J.; Bhattacharya, S.; Christopher, G.; Cutts, D.; Demiragli, Z.; Ferapontov, A.; Garabedian, A.; Heintz, U.; Kukartsev, G.; Laird, E.; Landsberg, G.; Luk, M.; Narain, M.; Segala, M.; Sinthuprasith, T.; Speer, T.; Breedon, R.; Breto, G.; Calderon De La Barca Sanchez, M.; Chauhan, S.; Chertok, M.; Conway, J.; Conway, R.; Cox, P. T.; Erbacher, R.; Gardner, M.; Houtz, R.; Ko, W.; Kopecky, A.; Lander, R.; Mall, O.; Miceli, T.; Nelson, R.; Pellett, D.; Ricci-Tam, F.; Rutherford, B.; Searle, M.; Smith, J.; Squires, M.; Tripathi, M.; Wilbur, S.; Yohay, R.; Andreev, V.; Cline, D.; Cousins, R.; Erhan, S.; Everaerts, P.; Farrell, C.; Felcini, M.; Hauser, J.; Ignatenko, M.; Jarvis, C.; Rakness, G.; Schlein, P.; Takasugi, E.; Traczyk, P.; Valuev, V.; Weber, M.; Babb, J.; Clare, R.; Dinardo, M. E.; Ellison, J.; Gary, J. W.; Giordano, F.; Hanson, G.; Liu, H.; Long, O. R.; Luthra, A.; Nguyen, H.; Paramesvaran, S.; Sturdy, J.; Sumowidagdo, S.; Wilken, R.; Wimpenny, S.; Andrews, W.; Branson, J. G.; Cerati, G. B.; Cittolin, S.; Evans, D.; Holzner, A.; Kelley, R.; Lebourgeois, M.; Letts, J.; Macneill, I.; Mangano, B.; Padhi, S.; Palmer, C.; Petrucciani, G.; Pieri, M.; Sani, M.; Sharma, V.; Simon, S.; Sudano, E.; Tadel, M.; Tu, Y.; Vartak, A.; Wasserbaech, S.; Würthwein, F.; Yagil, A.; Yoo, J.; Barge, D.; Bellan, R.; Campagnari, C.; D'Alfonso, M.; Danielson, T.; Flowers, K.; Geffert, P.; George, C.; Golf, F.; Incandela, J.; Justus, C.; Kalavase, P.; Kovalskyi, D.; Krutelyov, V.; Lowette, S.; Magaña Villalba, R.; Mccoll, N.; Pavlunin, V.; Ribnik, J.; Richman, J.; Rossin, R.; Stuart, D.; To, W.; West, C.; Apresyan, A.; Bornheim, A.; Bunn, J.; Chen, Y.; Di Marco, E.; Duarte, J.; Kcira, D.; Ma, Y.; Mott, A.; Newman, H. B.; Rogan, C.; Spiropulu, M.; Timciuc, V.; Veverka, J.; Wilkinson, R.; Xie, S.; Yang, Y.; Zhu, R. Y.; Azzolini, V.; Calamba, A.; Carroll, R.; Ferguson, T.; Iiyama, Y.; Jang, D. W.; Liu, Y. F.; Paulini, M.; Russ, J.; Vogel, H.; Vorobiev, I.; Cumalat, J. P.; Drell, B. R.; Ford, W. T.; Gaz, A.; Luiggi Lopez, E.; Nauenberg, U.; Smith, J. G.; Stenson, K.; Ulmer, K. A.; Wagner, S. R.; Alexander, J.; Chatterjee, A.; Eggert, N.; Gibbons, L. K.; Hopkins, W.; Khukhunaishvili, A.; Kreis, B.; Mirman, N.; Nicolas Kaufman, G.; Patterson, J. R.; Ryd, A.; Salvati, E.; Sun, W.; Teo, W. D.; Thom, J.; Thompson, J.; Tucker, J.; Weng, Y.; Winstrom, L.; Wittich, P.; Winn, D.; Abdullin, S.; Albrow, M.; Anderson, J.; Apollinari, G.; Bauerdick, L. A. T.; Beretvas, A.; Berryhill, J.; Bhat, P. C.; Burkett, K.; Butler, J. N.; Chetluru, V.; Cheung, H. W. K.; Chlebana, F.; Cihangir, S.; Elvira, V. D.; Fisk, I.; Freeman, J.; Gao, Y.; Gottschalk, E.; Gray, L.; Green, D.; Gutsche, O.; Harris, R. M.; Hirschauer, J.; Hooberman, B.; Jindariani, S.; Johnson, M.; Joshi, U.; Klima, B.; Kunori, S.; Kwan, S.; Linacre, J.; Lincoln, D.; Lipton, R.; Lykken, J.; Maeshima, K.; Marraffino, J. M.; Martinez Outschoorn, V. I.; Maruyama, S.; Mason, D.; McBride, P.; Mishra, K.; Mrenna, S.; Musienko, Y.; Newman-Holmes, C.; O'Dell, V.; Prokofyev, O.; Sexton-Kennedy, E.; Sharma, S.; Spalding, W. J.; Spiegel, L.; Taylor, L.; Tkaczyk, S.; Tran, N. V.; Uplegger, L.; Vaandering, E. W.; Vidal, R.; Whitmore, J.; Wu, W.; Yang, F.; Yun, J. C.; Acosta, D.; Avery, P.; Bourilkov, D.; Chen, M.; Cheng, T.; Das, S.; De Gruttola, M.; Di Giovanni, G. P.; Dobur, D.; Drozdetskiy, A.; Field, R. D.; Fisher, M.; Fu, Y.; Furic, I. K.; Hugon, J.; Kim, B.; Konigsberg, J.; Korytov, A.; Kropivnitskaya, A.; Kypreos, T.; Low, J. F.; Matchev, K.; Milenovic, P.; Mitselmakher, G.; Muniz, L.; Remington, R.; Rinkevicius, A.; Skhirtladze, N.; Snowball, M.; Yelton, J.; Zakaria, M.; Gaultney, V.; Hewamanage, S.; Lebolo, L. M.; Linn, S.; Markowitz, P.; Martinez, G.; Rodriguez, J. L.; Adams, T.; Askew, A.; Bochenek, J.; Chen, J.; Diamond, B.; Gleyzer, S. V.; Haas, J.; Hagopian, S.; Hagopian, V.; Johnson, K. F.; Prosper, H.; Veeraraghavan, V.; Weinberg, M.; Baarmand, M. M.; Dorney, B.; Hohlmann, M.; Kalakhety, H.; Yumiceva, F.; Adams, M. R.; Apanasevich, L.; Bazterra, V. E.; Betts, R. R.; Bucinskaite, I.; Callner, J.; Cavanaugh, R.; Evdokimov, O.; Gauthier, L.; Gerber, C. E.; Hofman, D. J.; Khalatyan, S.; Kurt, P.; Lacroix, F.; Moon, D. H.; O'Brien, C.; Silkworth, C.; Strom, D.; Turner, P.; Varelas, N.; Akgun, U.; Albayrak, E. A.; Bilki, B.; Clarida, W.; Dilsiz, K.; Duru, F.; Griffiths, S.; Merlo, J.-P.; Mermerkaya, H.; Mestvirishvili, A.; Moeller, A.; Nachtman, J.; Newsom, C. R.; Ogul, H.; Onel, Y.; Ozok, F.; Sen, S.; Tan, P.; Tiras, E.; Wetzel, J.; Yetkin, T.; Yi, K.; Barnett, B. A.; Blumenfeld, B.; Bolognesi, S.; Fehling, D.; Giurgiu, G.; Gritsan, A. V.; Hu, G.; Maksimovic, P.; Swartz, M.; Whitbeck, A.; Baringer, P.; Bean, A.; Benelli, G.; Kenny, R. P., III; Murray, M.; Noonan, D.; Sanders, S.; Stringer, R.; Wood, J. S.; Barfuss, A. F.; Chakaberia, I.; Ivanov, A.; Khalil, S.; Makouski, M.; Maravin, Y.; Shrestha, S.; Svintradze, I.; Gronberg, J.; Lange, D.; Rebassoo, F.; Wright, D.; Baden, A.; Calvert, B.; Eno, S. C.; Gomez, J. A.; Hadley, N. J.; Kellogg, R. G.; Kolberg, T.; Lu, Y.; Marionneau, M.; Mignerey, A. C.; Pedro, K.; Peterman, A.; Skuja, A.; Temple, J.; Tonjes, M. B.; Tonwar, S. C.; Apyan, A.; Bauer, G.; Busza, W.; Butz, E.; Cali, I. A.; Chan, M.; Dutta, V.; Gomez Ceballos, G.; Goncharov, M.; Kim, Y.; Klute, M.; Lai, Y. S.; Levin, A.; Luckey, P. D.; Ma, T.; Nahn, S.; Paus, C.; Ralph, D.; Roland, C.; Roland, G.; Stephans, G. S. F.; Stöckli, F.; Sumorok, K.; Sung, K.; Velicanu, D.; Wolf, R.; Wyslouch, B.; Yang, M.; Yilmaz, Y.; Yoon, A. S.; Zanetti, M.; Zhukova, V.; Dahmes, B.; De Benedetti, A.; Franzoni, G.; Gude, A.; Haupt, J.; Kao, S. C.; Klapoetke, K.; Kubota, Y.; Mans, J.; Pastika, N.; Rusack, R.; Sasseville, M.; Singovsky, A.; Tambe, N.; Turkewitz, J.; Cremaldi, L. M.; Kroeger, R.; Perera, L.; Rahmat, R.; Sanders, D. A.; Summers, D.; Avdeeva, E.; Bloom, K.; Bose, S.; Claes, D. R.; Dominguez, A.; Eads, M.; Gonzalez Suarez, R.; Keller, J.; Kravchenko, I.; Lazo-Flores, J.; Malik, S.; Snow, G. R.; Dolen, J.; Godshalk, A.; Iashvili, I.; Jain, S.; Kharchilava, A.; Kumar, A.; Rappoccio, S.; Wan, Z.; Alverson, G.; Barberis, E.; Baumgartel, D.; Chasco, M.; Haley, J.; Nash, D.; Orimoto, T.; Trocino, D.; Wood, D.; Zhang, J.; Anastassov, A.; Hahn, K. A.; Kubik, A.; Lusito, L.; Mucia, N.; Odell, N.; Pollack, B.; Pozdnyakov, A.; Schmitt, M.; Stoynev, S.; Velasco, M.; Won, S.; Berry, D.; Brinkerhoff, A.; Chan, K. M.; Hildreth, M.; Jessop, C.; Karmgard, D. J.; Kolb, J.; Lannon, K.; Luo, W.; Lynch, S.; Marinelli, N.; Morse, D. M.; Pearson, T.; Planer, M.; Ruchti, R.; Slaunwhite, J.; Valls, N.; Wayne, M.; Wolf, M.; Antonelli, L.; Bylsma, B.; Durkin, L. S.; Hill, C.; Hughes, R.; Kotov, K.; Ling, T. Y.; Puigh, D.; Rodenburg, M.; Smith, G.; Vuosalo, C.; Williams, G.; Winer, B. L.; Wolfe, H.; Berry, E.; Elmer, P.; Halyo, V.; Hebda, P.; Hegeman, J.; Hunt, A.; Jindal, P.; Koay, S. A.; Lopes Pegna, D.; Lujan, P.; Marlow, D.; Medvedeva, T.; Mooney, M.; Olsen, J.; Piroué, P.; Quan, X.; Raval, A.; Saka, H.; Stickland, D.; Tully, C.; Werner, J. S.; Zenz, S. C.; Zuranski, A.; Brownson, E.; Lopez, A.; Mendez, H.; Ramirez Vargas, J. E.; Alagoz, E.; Benedetti, D.; Bolla, G.; Bortoletto, D.; De Mattia, M.; Everett, A.; Hu, Z.; Jones, M.; Jung, K.; Koybasi, O.; Kress, M.; Leonardo, N.; Maroussov, V.; Merkel, P.; Miller, D. H.; Neumeister, N.; Shipsey, I.; Silvers, D.; Svyatkovskiy, A.; Vidal Marono, M.; Wang, F.; Xu, L.; Yoo, H. D.; Zablocki, J.; Zheng, Y.; Guragain, S.; Parashar, N.; Adair, A.; Akgun, B.; Ecklund, K. M.; Geurts, F. J. M.; Li, W.; Padley, B. P.; Redjimi, R.; Roberts, J.; Zabel, J.; Betchart, B.; Bodek, A.; Covarelli, R.; de Barbaro, P.; Demina, R.; Eshaq, Y.; Ferbel, T.; Garcia-Bellido, A.; Goldenzweig, P.; Han, J.; Harel, A.; Miner, D. C.; Petrillo, G.; Vishnevskiy, D.; Zielinski, M.; Bhatti, A.; Ciesielski, R.; Demortier, L.; Goulianos, K.; Lungu, G.; Malik, S.; Mesropian, C.; Arora, S.; Barker, A.; Chou, J. P.; Contreras-Campana, C.; Contreras-Campana, E.; Duggan, D.; Ferencek, D.; Gershtein, Y.; Gray, R.; Halkiadakis, E.; Hidas, D.; Lath, A.; Panwalkar, S.; Park, M.; Patel, R.; Rekovic, V.; Robles, J.; Rose, K.; Salur, S.; Schnetzer, S.; Seitz, C.; Somalwar, S.; Stone, R.; Thomas, S.; Walker, M.; Cerizza, G.; Hollingsworth, M.; Spanier, S.; Yang, Z. C.; York, A.; Eusebi, R.; Flanagan, W.; Gilmore, J.; Kamon, T.; Khotilovich, V.; Montalvo, R.; Osipenkov, I.; Pakhotin, Y.; Perloff, A.; Roe, J.; Safonov, A.; Sakuma, T.; Suarez, I.; Tatarinov, A.; Toback, D.; Akchurin, N.; Damgov, J.; Dragoiu, C.; Dudero, P. R.; Jeong, C.; Kovitanggoon, K.; Lee, S. W.; Libeiro, T.; Volobouev, I.; Appelt, E.; Delannoy, A. G.; Greene, S.; Gurrola, A.; Johns, W.; Maguire, C.; Mao, Y.; Melo, A.; Sharma, M.; Sheldon, P.; Snook, B.; Tuo, S.; Velkovska, J.; Arenton, M. W.; Boutle, S.; Cox, B.; Francis, B.; Goodell, J.; Hirosky, R.; Ledovskoy, A.; Lin, C.; Neu, C.; Wood, J.; Gollapinni, S.; Harr, R.; Karchin, P. E.; Kottachchi Kankanamge Don, C.; Lamichhane, P.; Sakharov, A.; Anderson, M.; Belknap, D. A.; Borrello, L.; Carlsmith, D.; Cepeda, M.; Dasu, S.; Friis, E.; Grogg, K. S.; Grothe, M.; Hall-Wilton, R.; Herndon, M.; Hervé, A.; Klabbers, P.; Klukas, J.; Lanaro, A.; Lazaridis, C.; Loveless, R.; Mohapatra, A.; Mozer, M. U.; Ojalvo, I.; Pierro, G. A.; Ross, I.; Savin, A.; Smith, W. H.; Swanson, J.

2013-10-01

A measurement is presented of the ratio of the inclusive 3-jet cross section to the inclusive 2-jet cross section as a function of the average transverse momentum, , of the two leading jets in the event. The data sample was collected during 2011 at a proton-proton centre-of-mass energy of 7 TeV with the CMS detector at the LHC, corresponding to an integrated luminosity of 5.0 fb-1. The strong coupling constant at the scale of the Z boson mass is determined to be α S ( M Z)=0.1148±0.0014 (exp.)±0.0018 (PDF)±0.0050(theory), by comparing the ratio in the range to the predictions of perturbative QCD at next-to-leading order. This is the first determination of α S ( M Z) from measurements at momentum scales beyond 0.6 TeV. The predicted ratio depends only indirectly on the evolution of the parton distribution functions of the proton such that this measurement also serves as a test of the evolution of the strong coupling constant. No deviation from the expected behaviour is observed.

Surface Protonation at the Rutile (110) Interface: Explicit Incorporation of Solvation Structure within the Refined MUSIC Model Framework

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

Machesky, Michael L.; Predota, M.; Wesolowski, David J

The detailed solvation structure at the (110) surface of rutile ({alpha}-TiO{sub 2}) in contact with bulk liquid water has been obtained primarily from experimentally verified classical molecular dynamics (CMD) simulations of the ab initio-optimized surface in contact with SPC/E water. The results are used to explicitly quantify H-bonding interactions, which are then used within the refined MUSIC model framework to predict surface oxygen protonation constants. Quantum mechanical molecular dynamics (QMD) simulations in the presence of freely dissociable water molecules produced H-bond distributions around deprotonated surface oxygens very similar to those obtained by CMD with nondissociable SPC/E water, thereby confirming thatmore » the less computationally intensive CMD simulations provide accurate H-bond information. Utilizing this H-bond information within the refined MUSIC model, along with manually adjusted Ti-O surface bond lengths that are nonetheless within 0.05 {angstrom} of those obtained from static density functional theory (DFT) calculations and measured in X-ray reflectivity experiments (as well as bulk crystal values), give surface protonation constants that result in a calculated zero net proton charge pH value (pHznpc) at 25 C that agrees quantitatively with the experimentally determined value (5.4 {+-} 0.2) for a specific rutile powder dominated by the (110) crystal face. Moreover, the predicted pH{sub znpc} values agree to within 0.1 pH unit with those measured at all temperatures between 10 and 250 C. A slightly smaller manual adjustment of the DFT-derived Ti-O surface bond lengths was sufficient to bring the predicted pH{sub znpc} value of the rutile (110) surface at 25 C into quantitative agreement with the experimental value (4.8 {+-} 0.3) obtained from a polished and annealed rutile (110) single crystal surface in contact with dilute sodium nitrate solutions using second harmonic generation (SHG) intensity measurements as a function of ionic strength. Additionally, the H-bond interactions between protolyzable surface oxygen groups and water were found to be stronger than those between bulk water molecules at all temperatures investigated in our CMD simulations (25, 150 and 250 C). Comparison with the protonation scheme previously determined for the (110) surface of isostructural cassiterite ({alpha}-SnO{sub 2}) reveals that the greater extent of H-bonding on the latter surface, and in particular between water and the terminal hydroxyl group (Sn-OH) results in the predicted protonation constant for that group being lower than for the bridged oxygen (Sn-O-Sn), while the reverse is true for the rutile (110) surface. These results demonstrate the importance of H-bond structure in dictating surface protonation behavior, and that explicit use of this solvation structure within the refined MUSIC model framework results in predicted surface protonation constants that are also consistent with a variety of other experimental and computational data.« less

Surface protonation at the rutile (110) interface: explicit incorporation of solvation structure within the refined MUSIC model framework.

PubMed

Machesky, Michael L; Predota, Milan; Wesolowski, David J; Vlcek, Lukas; Cummings, Peter T; Rosenqvist, Jörgen; Ridley, Moira K; Kubicki, James D; Bandura, Andrei V; Kumar, Nitin; Sofo, Jorge O

2008-11-04

The detailed solvation structure at the (110) surface of rutile (alpha-TiO2) in contact with bulk liquid water has been obtained primarily from experimentally verified classical molecular dynamics (CMD) simulations of the ab initio-optimized surface in contact with SPC/E water. The results are used to explicitly quantify H-bonding interactions, which are then used within the refined MUSIC model framework to predict surface oxygen protonation constants. Quantum mechanical molecular dynamics (QMD) simulations in the presence of freely dissociable water molecules produced H-bond distributions around deprotonated surface oxygens very similar to those obtained by CMD with nondissociable SPC/E water, thereby confirming that the less computationally intensive CMD simulations provide accurate H-bond information. Utilizing this H-bond information within the refined MUSIC model, along with manually adjusted Ti-O surface bond lengths that are nonetheless within 0.05 A of those obtained from static density functional theory (DFT) calculations and measured in X-ray reflectivity experiments (as well as bulk crystal values), give surface protonation constants that result in a calculated zero net proton charge pH value (pHznpc) at 25 degrees C that agrees quantitatively with the experimentally determined value (5.4+/-0.2) for a specific rutile powder dominated by the (110) crystal face. Moreover, the predicted pHznpc values agree to within 0.1 pH unit with those measured at all temperatures between 10 and 250 degrees C. A slightly smaller manual adjustment of the DFT-derived Ti-O surface bond lengths was sufficient to bring the predicted pHznpcvalue of the rutile (110) surface at 25 degrees C into quantitative agreement with the experimental value (4.8+/-0.3) obtained from a polished and annealed rutile (110) single crystal surface in contact with dilute sodium nitrate solutions using second harmonic generation (SHG) intensity measurements as a function of ionic strength. Additionally, the H-bond interactions between protolyzable surface oxygen groups and water were found to be stronger than those between bulk water molecules at all temperatures investigated in our CMD simulations (25, 150 and 250 degrees C). Comparison with the protonation scheme previously determined for the (110) surface of isostructural cassiterite (alpha-SnO2) reveals that the greater extent of H-bonding on the latter surface, and in particular between water and the terminal hydroxyl group (Sn-OH) results in the predicted protonation constant for that group being lower than for the bridged oxygen (Sn-O-Sn), while the reverse is true for the rutile (110) surface. These results demonstrate the importance of H-bond structure in dictating surface protonation behavior, and that explicit use of this solvation structure within the refined MUSIC model framework results in predicted surface protonation constants that are also consistent with a variety of other experimental and computational data.

Determination of the molecular complexation constant between alprostadil and alpha-cyclodextrin by conductometry: implications for a freeze-dried formulation.

PubMed

Sheehy, Philip M; Ramstad, Tore

2005-10-04

The binding constant between alprostadil (PGE1) and alpha-cyclodextrin (alpha-CD) was determined at four temperatures using conductance measurements. Alpha-cyclodextrin is an excipient material in Caverject dual chamber syringe (DCS) that was added to enhance stability. The binding constant was used to calculate the amount of PGE1 free upon reconstitution and injection, since only the free drug is clinically active. The conductivity measurement is based on a decrease in specific conductance as alprostadil is titrated with alpha-CD. The change in conductivity was plotted versus free ligand concentration (alpha-CD) to generate a binding curve. As the value of the binding constant proved to be dependent on substrate concentration, it is really a pseudo binding constant. A value of 742+/-60 M(-1) was obtained for a 0.5 mM solution of alprostadil at 27 degrees C and a value of 550+/-52 M(-1) at 37 degrees C. These results compare favorably to values previously obtained by NMR and capillary electrophoresis. Calculation of the fraction PGE1 free upon reconstitution and injection show it to approach the desired outcome of one. Hence, the amount of drug delivered by Caverject DCS is nominally equivalent to that delivered by Caverject S. Po., a predecessor product that contains no alpha-cyclodextrin.

Spectroscopic analyses on interaction of o-Vanillin- D-Phenylalanine, o-Vanillin- L-Tyrosine and o-Vanillin- L-Levodopa Schiff Bases with bovine serum albumin (BSA)

NASA Astrophysics Data System (ADS)

Gao, Jingqun; Guo, Yuwei; Wang, Jun; Wang, Zhiqiu; Jin, Xudong; Cheng, Chunping; Li, Ying; Li, Kai

2011-04-01

In this work, three o-Vanillin Schiff Bases (o-VSB: o-Vanillin- D-Phenylalanine (o-VDP), o-Vanillin- L-Tyrosine (o-VLT) and o-Vanillin- L-Levodopa (o-VLL)) with alanine constituent were synthesized by direct reflux method in ethanol solution, and then were used to study the interaction to bovine serum albumin (BSA) molecules by fluorescence spectroscopy. Based on the fluorescence quenching calculation, the bimolecular quenching constant ( Kq), apparent quenching constant ( Ksv), effective binding constant ( KA) and corresponding dissociation constant ( KD) as well as binding site number ( n) were obtained. In addition, the binding distance ( r) was also calculated according to Foster's non-radioactive energy transfer theory. The results show that these three o-VSB can efficiently bind to BSA molecules, but the binding array order is o-VDP-BSA > o-VLT-BSA > o-VLL-BSA. Synchronous fluorescence spectroscopy indicates that the o-VDP is more accessibility to tryptophan (Trp) residues of BSA molecules than to tyrosine (Tyr) residues. Nevertheless, the o-VLT and o-VLL are more accessibility to Tyr residues than to Trp residues.

Spectroscopic analyses on interaction of o-Vanillin-D-Phenylalanine, o-Vanillin-L-Tyrosine and o-Vanillin-L-Levodopa Schiff Bases with bovine serum albumin (BSA).

PubMed

Gao, Jingqun; Guo, Yuwei; Wang, Jun; Wang, Zhiqiu; Jin, Xudong; Cheng, Chunping; Li, Ying; Li, Kai

2011-04-01

In this work, three o-Vanillin Schiff Bases (o-VSB: o-Vanillin-D-Phenylalanine (o-VDP), o-Vanillin-L-Tyrosine (o-VLT) and o-Vanillin-L-Levodopa (o-VLL)) with alanine constituent were synthesized by direct reflux method in ethanol solution, and then were used to study the interaction to bovine serum albumin (BSA) molecules by fluorescence spectroscopy. Based on the fluorescence quenching calculation, the bimolecular quenching constant (K(q)), apparent quenching constant (K(sv)), effective binding constant (K(A)) and corresponding dissociation constant (K(D)) as well as binding site number (n) were obtained. In addition, the binding distance (r) was also calculated according to Foster's non-radioactive energy transfer theory. The results show that these three o-VSB can efficiently bind to BSA molecules, but the binding array order is o-VDP-BSA>o-VLT-BSA>o-VLL-BSA. Synchronous fluorescence spectroscopy indicates that the o-VDP is more accessibility to tryptophan (Trp) residues of BSA molecules than to tyrosine (Tyr) residues. Nevertheless, the o-VLT and o-VLL are more accessibility to Tyr residues than to Trp residues. Copyright © 2010 Elsevier B.V. All rights reserved.

Intrinsic thermodynamics of inhibitor binding to human carbonic anhydrase IX.

PubMed

Linkuvienė, Vaida; Matulienė, Jurgita; Juozapaitienė, Vaida; Michailovienė, Vilma; Jachno, Jelena; Matulis, Daumantas

2016-04-01

Human carbonic anhydrase 9th isoform (CA IX) is an important marker of numerous cancers and is increasingly interesting as a potential anticancer drug target. Various synthetic aromatic sulfonamide-bearing compounds are being designed as potent inhibitors of CA IX. However, sulfonamide compound binding to CA IX is linked to several reactions, the deprotonation of the sulfonamide amino group and the protonation of the CA active site Zn(II)-bound hydroxide. These linked reactions significantly affect the affinities and other thermodynamic parameters such as enthalpies and entropies of binding. The observed and intrinsic affinities of compound binding to CA IX were determined by the fluorescent thermal shift assay. The enthalpies and entropies of binding were determined by the isothermal titration calorimetry. The pKa of CA IX was determined to be 6.8 and the enthalpy of CA IX-Zn(II)-bound hydroxide protonation was -24 kJ/mol. These values enabled the analysis of intrinsic thermodynamics of a library of compounds binding to CA IX. The most strongly binding compounds exhibited the intrinsic affinity of 0.01 nM and the observed affinity of 2 nM. The intrinsic thermodynamic parameters of compound binding to CA IX helped to draw the compound structure to thermodynamics relationship. It is important to distinguish the intrinsic from observed parameters of any disease target protein interaction with its inhibitors as drug candidates when drawing detailed compound structure to thermodynamics correlations. Copyright © 2016 Elsevier B.V. All rights reserved.

Constant-pH Hybrid Nonequilibrium Molecular Dynamics–Monte Carlo Simulation Method

PubMed Central

2016-01-01

A computational method is developed to carry out explicit solvent simulations of complex molecular systems under conditions of constant pH. In constant-pH simulations, preidentified ionizable sites are allowed to spontaneously protonate and deprotonate as a function of time in response to the environment and the imposed pH. The method, based on a hybrid scheme originally proposed by H. A. Stern (J. Chem. Phys.2007, 126, 164112), consists of carrying out short nonequilibrium molecular dynamics (neMD) switching trajectories to generate physically plausible configurations with changed protonation states that are subsequently accepted or rejected according to a Metropolis Monte Carlo (MC) criterion. To ensure microscopic detailed balance arising from such nonequilibrium switches, the atomic momenta are altered according to the symmetric two-ends momentum reversal prescription. To achieve higher efficiency, the original neMD–MC scheme is separated into two steps, reducing the need for generating a large number of unproductive and costly nonequilibrium trajectories. In the first step, the protonation state of a site is randomly attributed via a Metropolis MC process on the basis of an intrinsic pKa; an attempted nonequilibrium switch is generated only if this change in protonation state is accepted. This hybrid two-step inherent pKa neMD–MC simulation method is tested with single amino acids in solution (Asp, Glu, and His) and then applied to turkey ovomucoid third domain and hen egg-white lysozyme. Because of the simple linear increase in the computational cost relative to the number of titratable sites, the present method is naturally able to treat extremely large systems. PMID:26300709

Protons migrate along interfacial water without significant contributions from jumps between ionizable groups on the membrane surface

PubMed Central

Springer, Andreas; Hagen, Volker; Cherepanov, Dmitry A.; Antonenko, Yuri N.; Pohl, Peter

2011-01-01

Proton diffusion along membrane surfaces is thought to be essential for many cellular processes such as energy transduction. Commonly, it is treated as a succession of jumps between membrane-anchored proton-binding sites. Our experiments provide evidence for an alternative model. We released membrane-bound caged protons by UV flashes and monitored their arrival at distant sites by fluorescence measurements. The kinetics of the arrival is probed as a function of distance for different membranes and for different water isotopes. We found that proton diffusion along the membrane is fast even in the absence of ionizable groups in the membrane, and it decreases strongly in D2O as compared to H2O. We conclude that the fast proton transport along the membrane is dominated by diffusion via interfacial water, and not via ionizable lipid moieties. PMID:21859952

The structure of binding curves and practical identifiability of equilibrium ligand-binding parameters

PubMed Central

Middendorf, Thomas R.

2017-01-01

A critical but often overlooked question in the study of ligands binding to proteins is whether the parameters obtained from analyzing binding data are practically identifiable (PI), i.e., whether the estimates obtained from fitting models to noisy data are accurate and unique. Here we report a general approach to assess and understand binding parameter identifiability, which provides a toolkit to assist experimentalists in the design of binding studies and in the analysis of binding data. The partial fraction (PF) expansion technique is used to decompose binding curves for proteins with n ligand-binding sites exactly and uniquely into n components, each of which has the form of a one-site binding curve. The association constants of the PF component curves, being the roots of an n-th order polynomial, may be real or complex. We demonstrate a fundamental connection between binding parameter identifiability and the nature of these one-site association constants: all binding parameters are identifiable if the constants are all real and distinct; otherwise, at least some of the parameters are not identifiable. The theory is used to construct identifiability maps from which the practical identifiability of binding parameters for any two-, three-, or four-site binding curve can be assessed. Instructions for extending the method to generate identifiability maps for proteins with more than four binding sites are also given. Further analysis of the identifiability maps leads to the simple rule that the maximum number of structurally identifiable binding parameters (shown in the previous paper to be equal to n) will also be PI only if the binding curve line shape contains n resolved components. PMID:27993951

Energetic particles in the pre-dawn magnetotail of Jupiter

NASA Technical Reports Server (NTRS)

Schardt, A. W.; Mcdonald, F. B.; Trainor, J. H.

1980-01-01

A detailed account is given of the energetic electron and proton populations as observed with Voyagers 1 and 2 during their passes through the dawn magnetotail of Jupiter. The region between 20 and 150 R sub J is dominated by a thin plasma sheet, where trapped energetic electron and proton fluxes reach their maximum. Proton spectra can be represented by an exponential in rigidity with a characteristic energy of approximately 50 keV. Proton anisotropies were consistent with corotation even at 100 R sub J. A major proton acceleration event as well as several cases of field aligned proton streaming were observed. The flux of 0.4 MeV protons decreases by three orders of magnitude between 30 and 90 R sub J and then remains relatively constant to the magnetopause. Fine structure in the data indicate longitudinal asymmetries with respect to the dipole orientation. Electron spectra in the magnetosheath and interplanetary space are modulated by the Jovian longitude relative to the subsolar point.

CONSTANTS FOR MERCURY BINDING BY DISSOLVED ORGANIC MATTER ISOLATES FROM THE FLORIDA EVERGLADES. (R827653)

EPA Science Inventory

Dissolved organic matter (DOM) has been implicated as an important complexing agent for Hg that can affect its mobility and bioavailability in aquatic ecosystems. However, binding constants for natural Hg-DOM complexes are not well known. We employed a competitive ligand appro...

Characterization of the binding of metoprolol tartrate and guaifenesin drugs to human serum albumin and human hemoglobin proteins by fluorescence and circular dichroism spectroscopy.

PubMed

Duman, Osman; Tunç, Sibel; Kancı Bozoğlan, Bahar

2013-07-01

The interactions of metoprolol tartrate (MPT) and guaifenesin (GF) drugs with human serum albumin (HSA) and human hemoglobin (HMG) proteins at pH 7.4 were studied by fluorescence and circular dichroism (CD) spectroscopy. Drugs quenched the fluorescence spectra of HSA and HMG proteins through a static quenching mechanism. For each protein-drug system, the values of Stern-Volmer quenching constant, bimolecular quenching constant, binding constant and number of binding site on the protein molecules were determined at 288.15, 298.15, 310.15 and 318.15 K. It was found that the binding constants of HSA-MPT and HSA-GF systems were smaller than those of HMG-MPT and HMG-GF systems. For both drugs, the affinity of HMG was much higher than that of HSA. An increase in temperature caused a negative effect on the binding reactions. The number of binding site on blood proteins for MPT and GF drugs was approximately one. Thermodynamic parameters showed that MPT interacted with HSA through electrostatic attraction forces. However, hydrogen bonds and van der Waals forces were the main interaction forces in the formation of HSA-GF, HMG-MPT and HMG-GF complexes. The binding processes between protein and drug molecules were exothermic and spontaneous owing to negative ∆H and ∆G values, respectively. The values of binding distance between protein and drug molecules were calculated from Förster resonance energy transfer theory. It was found from CD analysis that the bindings of MPT and GF drugs to HSA and HMG proteins altered the secondary structure of HSA and HMG proteins.

Unconventional hydrogen bonding to organic ions in the gas phase: Stepwise association of hydrogen cyanide with the pyridine and pyrimidine radical cations and protonated pyridine

NASA Astrophysics Data System (ADS)

Hamid, Ahmed M.; El-Shall, M. Samy; Hilal, Rifaat; Elroby, Shaaban; Aziz, Saadullah G.

2014-08-01

Equilibrium thermochemical measurements using the ion mobility drift cell technique have been utilized to investigate the binding energies and entropy changes for the stepwise association of HCN molecules with the pyridine and pyrimidine radical cations forming the C5H5N+.(HCN)n and C4H4N2+.(HCN)n clusters, respectively, with n = 1-4. For comparison, the binding of 1-4 HCN molecules to the protonated pyridine C5H5NH+(HCN)n has also been investigated. The binding energies of HCN to the pyridine and pyrimidine radical cations are nearly equal (11.4 and 12.0 kcal/mol, respectively) but weaker than the HCN binding to the protonated pyridine (14.0 kcal/mol). The pyridine and pyrimidine radical cations form unconventional carbon-based ionic hydrogen bonds with HCN (CHδ+⋯NCH). Protonated pyridine forms a stronger ionic hydrogen bond with HCN (NH+⋯NCH) which can be extended to a linear chain with the clustering of additional HCN molecules (NH+⋯NCH..NCH⋯NCH) leading to a rapid decrease in the bond strength as the length of the chain increases. The lowest energy structures of the pyridine and pyrimidine radical cation clusters containing 3-4 HCN molecules show a strong tendency for the internal solvation of the radical cation by the HCN molecules where bifurcated structures involving multiple hydrogen bonding sites with the ring hydrogen atoms are formed. The unconventional H-bonds (CHδ+⋯NCH) formed between the pyridine or the pyrimidine radical cations and HCN molecules (11-12 kcal/mol) are stronger than the similar (CHδ+⋯NCH) bonds formed between the benzene radical cation and HCN molecules (9 kcal/mol) indicating that the CHδ+ centers in the pyridine and pyrimidine radical cations have more effective charges than in the benzene radical cation.

On the Rutherford-Santilli neutron model

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

Burande, Chandrakant S.

2015-03-10

In 1920 H. Rutherford conjectured that the first particle synthesized in stars is neutron from a proton and an electron after which all known matter is progressively synthesized. However, Pauli objected Rutherford’s version of neutron synthesis because inability to represent spin 1/2 of the neutron. Using this objection E. Fermi proposed emission of massless particle, called “neutrino”. However, Santilli has dismissed the neutrino hypothesis following certain ambiguities such as positive binding energy required in synthesis of neutron. He found that celebrated Schrödinger’s equation of quantum physics is not suitable for obtaining positive binding energy for bound state at the dimensionmore » of 10{sup −13}cm. In order to remove these shortcomings, Santilli has developed isomathematics and then hadronic mechanics, which allowed the time invariant representation of Hamiltonian and non-Hamiltonian interactions as needed for the neutron synthesis (see for example: References cited at [1]).Thus the anomalies pertaining to the binding energy, the spin and the magnetic moment got resolved. He successfully calculated missing positive binding energy via isonormalization of the mass for electron when totally immersed within the hyper-dense medium inside the proton. Considering Rutherford’s compression of the isoelectron within the proton in the singlet coupling, he also identified the spin 1/2 for neutron and calculated the magnetic moment of the neutron. In order to verify his logical concept, he repeated the Don Carlo Borghi experiment of synthesis of the neutron from proton and electrons and verified that the said setup indeed produces neutron-type particles called “neutroids” which latter is absorbed by the activated detector substances that produces known nuclear reactions. He dismissed the neutrino hypothesis and replaced it with a longitudinal impulse originating from the ether as a universal substratum, named, “etherino”. He pointed out that all the physical quantities missing in the neutron synthesis, such as energy and spin, are delivered by said impulse.« less

Unconventional hydrogen bonding to organic ions in the gas phase: stepwise association of hydrogen cyanide with the pyridine and pyrimidine radical cations and protonated pyridine.

PubMed

Hamid, Ahmed M; El-Shall, M Samy; Hilal, Rifaat; Elroby, Shaaban; Aziz, Saadullah G

2014-08-07

Equilibrium thermochemical measurements using the ion mobility drift cell technique have been utilized to investigate the binding energies and entropy changes for the stepwise association of HCN molecules with the pyridine and pyrimidine radical cations forming the C5H5N(+·)(HCN)n and C4H4N2 (+·)(HCN)n clusters, respectively, with n = 1-4. For comparison, the binding of 1-4 HCN molecules to the protonated pyridine C5H5NH(+)(HCN)n has also been investigated. The binding energies of HCN to the pyridine and pyrimidine radical cations are nearly equal (11.4 and 12.0 kcal/mol, respectively) but weaker than the HCN binding to the protonated pyridine (14.0 kcal/mol). The pyridine and pyrimidine radical cations form unconventional carbon-based ionic hydrogen bonds with HCN (CH(δ+)⋯NCH). Protonated pyridine forms a stronger ionic hydrogen bond with HCN (NH(+)⋯NCH) which can be extended to a linear chain with the clustering of additional HCN molecules (NH(+)⋯NCH··NCH⋯NCH) leading to a rapid decrease in the bond strength as the length of the chain increases. The lowest energy structures of the pyridine and pyrimidine radical cation clusters containing 3-4 HCN molecules show a strong tendency for the internal solvation of the radical cation by the HCN molecules where bifurcated structures involving multiple hydrogen bonding sites with the ring hydrogen atoms are formed. The unconventional H-bonds (CH(δ+)⋯NCH) formed between the pyridine or the pyrimidine radical cations and HCN molecules (11-12 kcal/mol) are stronger than the similar (CH(δ+)⋯NCH) bonds formed between the benzene radical cation and HCN molecules (9 kcal/mol) indicating that the CH(δ+) centers in the pyridine and pyrimidine radical cations have more effective charges than in the benzene radical cation.

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

Johnston, Ryne C.; Zhou, Jing; Smith, Jeremy C.

In redox processes in complex transition metal-containing species are often intimately associated with changes in ligand protonation states and metal coordination number. Moreover, a major challenge is therefore to develop consistent computational approaches for computing pH-dependent redox and ligand dissociation properties of organometallic species. Reduction of the Co center in the vitamin B12 derivative aquacobalamin can be accompanied by ligand dissociation, protonation, or both, making these properties difficult to compute accurately. We examine this challenge here by using density functional theory and continuum solvation to compute Co ligand binding equilibrium constants (Kon/off), pKas and reduction potentials for models of aquacobalaminmore » in aqueous solution. We consider two models for cobalamin ligand coordination: the first follows the hexa, penta, tetra coordination scheme for Co III, Co II, and Co I species, respectively, and the second model features saturation of each vacant axial coordination site on Co II and Co I species with a single, explicit water molecule to maintain six directly interacting ligands or water molecules in each oxidation state. Comparing these two coordination schemes in combination with five dispersion-corrected density functionals, we find that the accuracy of the computed properties is largely independent of the scheme used, but including only a continuum representation of the solvent yields marginally better results than saturating the first solvation shell around Co throughout. PBE performs best, displaying balanced accuracy and superior performance overall, with RMS errors of 80 mV for seven reduction potentials, 2.0 log units for five pK as and 2.3 log units for two log K on/off values for the aquacobalamin system. Furthermore, we find that the BP86 functional commonly used in corrinoid studies suffers from erratic behavior and inaccurate descriptions of Co axial ligand binding, leading to substantial errors in predicted pK as and K on/off values. Finally, these findings demonstrate the effectiveness of the present approach for computing electrochemical and thermodynamic properties of a complex transition metal-containing cofactor.« less

Basic Residues R260 and K357 Affect the Conformational Dynamics of the Major Facilitator Superfamily Multidrug Transporter LmrP

PubMed Central

Wang, Wei; van Veen, Hendrik W.

2012-01-01

Secondary-active multidrug transporters can confer resistance on cells to pharmaceuticals by mediating their extrusion away from intracellular targets via substrate/H+(Na+) antiport. While the interactions of catalytic carboxylates in these transporters with coupling ions and substrates (drugs) have been studied in some detail, the functional importance of basic residues has received much less attention. The only two basic residues R260 and K357 in transmembrane helices in the Major Facilitator Superfamily transporter LmrP from Lactococcus lactis are present on the outer surface of the protein, where they are exposed to the phospholipid head group region of the outer leaflet (R260) and inner leaflet (K357) of the cytoplasmic membrane. Although our observations on the proton-motive force dependence and kinetics of substrate transport, and substrate-dependent proton transport demonstrate that K357A and R260A mutants are affected in ethidium-proton and benzalkonium-proton antiport compared to wildtype LmrP, our findings suggest that R260 and K357 are not directly involved in the binding of substrates or the translocation of protons. Secondary-active multidrug transporters are thought to operate by a mechanism in which binding sites for substrates are alternately exposed to each face of the membrane. Disulfide crosslinking experiments were performed with a double cysteine mutant of LmrP that reports the substrate-stimulated transition from the outward-facing state to the inward-facing state with high substrate-binding affinity. In the experiments, the R260A and K357A mutations were found to influence the dynamics of these major protein conformations in the transport cycle, potentially by removing the interactions of R260 and K357 with phospholipids and/or other residues in LmrP. The R260A and K357A mutations therefore modify the maximum rate at which the transport cycle can operate and, as the transitions between conformational states are differently affected by components of the proton-motive force, the mutations also influence the energetics of transport. PMID:22761697

A two-metal ion mechanism operates in the hammerhead ribozyme-mediated cleavage of an RNA substrate

PubMed Central

Lott, William B.; Pontius, Brian W.; von Hippel, Peter H.

1998-01-01

Evidence for a two-metal ion mechanism for cleavage of the HH16 hammerhead ribozyme is provided by monitoring the rate of cleavage of the RNA substrate as a function of La3+ concentration in the presence of a constant concentration of Mg2+. We show that a bell-shaped curve of cleavage activation is obtained as La3+ is added in micromolar concentrations in the presence of 8 mM Mg2+, with a maximal rate of cleavage being attained in the presence of 3 μM La3+. These results show that two-metal ion binding sites on the ribozyme regulate the rate of the cleavage reaction and, on the basis of earlier estimates of the Kd values for Mg2+ of 3.5 mM and >50 mM, that these sites bind La3+ with estimated Kd values of 0.9 and >37.5 μM, respectively. Furthermore, given the very different effects of these metal ions at the two binding sites, with displacement of Mg2+ by La3+ at the stronger (relative to Mg2+) binding site activating catalysis and displacement of Mg2+ by La3+ at the weaker (relative to Mg2+) (relative to Mg2+) binding site inhibiting catalysis, we show that the metal ions at these two sites play very different roles. We argue that the metal ion at binding site 1 coordinates the attacking 2′-oxygen species in the reaction and lowers the pKa of the attached proton, thereby increasing the concentration of the attacking alkoxide nucleophile in an equilibrium process. In contrast, the role of the metal ion at binding site 2 is to catalyze the reaction by absorbing the negative charge that accumulates at the leaving 5′-oxygen in the transition state. We suggest structural reasons why the Mg2+–La3+ ion combination is particularly suited to demonstrating these different roles of the two-metal ions in the ribozyme cleavage reaction. PMID:9435228

A global reaction route mapping-based kinetic Monte Carlo algorithm

NASA Astrophysics Data System (ADS)

Mitchell, Izaac; Irle, Stephan; Page, Alister J.

2016-07-01

We propose a new on-the-fly kinetic Monte Carlo (KMC) method that is based on exhaustive potential energy surface searching carried out with the global reaction route mapping (GRRM) algorithm. Starting from any given equilibrium state, this GRRM-KMC algorithm performs a one-step GRRM search to identify all surrounding transition states. Intrinsic reaction coordinate pathways are then calculated to identify potential subsequent equilibrium states. Harmonic transition state theory is used to calculate rate constants for all potential pathways, before a standard KMC accept/reject selection is performed. The selected pathway is then used to propagate the system forward in time, which is calculated on the basis of 1st order kinetics. The GRRM-KMC algorithm is validated here in two challenging contexts: intramolecular proton transfer in malonaldehyde and surface carbon diffusion on an iron nanoparticle. We demonstrate that in both cases the GRRM-KMC method is capable of reproducing the 1st order kinetics observed during independent quantum chemical molecular dynamics simulations using the density-functional tight-binding potential.

Biolayer interferometry of lipid nanodisc‐reconstituted yeast vacuolar H+‐ATPase

PubMed Central

Sharma, Stuti

2017-01-01

Abstract Vacuolar H+‐ATPase (V‐ATPase) is a large, multisubunit membrane protein complex responsible for the acidification of subcellular compartments and the extracellular space. V‐ATPase activity is regulated by reversible disassembly, resulting in cytosolic V 1‐ATPase and membrane‐integral V 0 proton channel sectors. Reversible disassembly is accompanied by transient interaction with cellular factors and assembly chaperones. Quantifying protein‐protein interactions involving membrane proteins, however, is challenging. Here we present a novel method to determine kinetic constants of membrane protein–protein interactions using biolayer interferometry (BLI). Yeast vacuoles are solubilized, vacuolar proteins are reconstituted into lipid nanodiscs with native vacuolar lipids and biotinylated membrane scaffold protein (MSP) followed by affinity purification of nanodisc‐reconstituted V‐ATPase (V 1 V 0ND). We show that V 1 V 0ND can be immobilized on streptavidin‐coated BLI sensors to quantitate binding of a pathogen derived inhibitor and to measure the kinetics of nucleotide dependent enzyme dissociation. PMID:28241399

Biolayer interferometry of lipid nanodisc-reconstituted yeast vacuolar H+ -ATPase.

PubMed

Sharma, Stuti; Wilkens, Stephan

2017-05-01

Vacuolar H + -ATPase (V-ATPase) is a large, multisubunit membrane protein complex responsible for the acidification of subcellular compartments and the extracellular space. V-ATPase activity is regulated by reversible disassembly, resulting in cytosolic V 1 -ATPase and membrane-integral V 0 proton channel sectors. Reversible disassembly is accompanied by transient interaction with cellular factors and assembly chaperones. Quantifying protein-protein interactions involving membrane proteins, however, is challenging. Here we present a novel method to determine kinetic constants of membrane protein-protein interactions using biolayer interferometry (BLI). Yeast vacuoles are solubilized, vacuolar proteins are reconstituted into lipid nanodiscs with native vacuolar lipids and biotinylated membrane scaffold protein (MSP) followed by affinity purification of nanodisc-reconstituted V-ATPase (V 1 V 0 ND). We show that V 1 V 0 ND can be immobilized on streptavidin-coated BLI sensors to quantitate binding of a pathogen derived inhibitor and to measure the kinetics of nucleotide dependent enzyme dissociation. © 2017 The Protein Society.

A global reaction route mapping-based kinetic Monte Carlo algorithm.

PubMed

Mitchell, Izaac; Irle, Stephan; Page, Alister J

2016-07-14

We propose a new on-the-fly kinetic Monte Carlo (KMC) method that is based on exhaustive potential energy surface searching carried out with the global reaction route mapping (GRRM) algorithm. Starting from any given equilibrium state, this GRRM-KMC algorithm performs a one-step GRRM search to identify all surrounding transition states. Intrinsic reaction coordinate pathways are then calculated to identify potential subsequent equilibrium states. Harmonic transition state theory is used to calculate rate constants for all potential pathways, before a standard KMC accept/reject selection is performed. The selected pathway is then used to propagate the system forward in time, which is calculated on the basis of 1st order kinetics. The GRRM-KMC algorithm is validated here in two challenging contexts: intramolecular proton transfer in malonaldehyde and surface carbon diffusion on an iron nanoparticle. We demonstrate that in both cases the GRRM-KMC method is capable of reproducing the 1st order kinetics observed during independent quantum chemical molecular dynamics simulations using the density-functional tight-binding potential.

Fragmentation analysis of α-induced reactions using clusterization approach

NASA Astrophysics Data System (ADS)

Kaur, Amandeep; Sharma, Manoj K.

2018-01-01

The dynamics of α-induced reactions are worked out over an incident beam energy Eα ∼ 10- 15 MeV using targets of different masses. The decay patterns of odd mass compound systems 117Sb*, 145Pm* and 191Ir* formed in α +113In, α +141Pr and α +187Re reactions are investigated in view of n-evaporation data. The methodology of collective clusterization is applied by optimizing the neck-length parameter ΔR and the DCM calculated cross-sections find nice agreement with the experimental data. The resulting compound systems with ACN = 117- 191 cover a wide range of compound nucleus mass, and hence give an opportunity to explore various aspects related to the dynamics involved. Moreover the neutron-proton asymmetry dependence is explored in terms of the Bulk constant (α) (in the liquid drop binding energy expression) and radius term Ri and its consequent influence on the fragmentation structure of these compound systems is investigated.

Binding equilibrium and kinetics of membrane-anchored receptors and ligands in cell adhesion: Insights from computational model systems and theory.

PubMed

Weikl, Thomas R; Hu, Jinglei; Xu, Guang-Kui; Lipowsky, Reinhard

2016-09-02

The adhesion of cell membranes is mediated by the binding of membrane-anchored receptor and ligand proteins. In this article, we review recent results from simulations and theory that lead to novel insights on how the binding equilibrium and kinetics of these proteins is affected by the membranes and by the membrane anchoring and molecular properties of the proteins. Simulations and theory both indicate that the binding equilibrium constant [Formula: see text] and the on- and off-rate constants of anchored receptors and ligands in their 2-dimensional (2D) membrane environment strongly depend on the membrane roughness from thermally excited shape fluctuations on nanoscales. Recent theory corroborated by simulations provides a general relation between [Formula: see text] and the binding constant [Formula: see text] of soluble variants of the receptors and ligands that lack the membrane anchors and are free to diffuse in 3 dimensions (3D).

Binding equilibrium and kinetics of membrane-anchored receptors and ligands in cell adhesion: Insights from computational model systems and theory

PubMed Central

Weikl, Thomas R.; Hu, Jinglei; Xu, Guang-Kui; Lipowsky, Reinhard

2016-01-01

ABSTRACT The adhesion of cell membranes is mediated by the binding of membrane-anchored receptor and ligand proteins. In this article, we review recent results from simulations and theory that lead to novel insights on how the binding equilibrium and kinetics of these proteins is affected by the membranes and by the membrane anchoring and molecular properties of the proteins. Simulations and theory both indicate that the binding equilibrium constant K2D and the on- and off-rate constants of anchored receptors and ligands in their 2-dimensional (2D) membrane environment strongly depend on the membrane roughness from thermally excited shape fluctuations on nanoscales. Recent theory corroborated by simulations provides a general relation between K2D and the binding constant K3D of soluble variants of the receptors and ligands that lack the membrane anchors and are free to diffuse in 3 dimensions (3D). PMID:27294442

Kinetics and equilibria of cyanide binding to prostaglandin H synthase.

PubMed

MacDonald, I D; Dunford, H B

1989-09-01

Cyanide binding to prostaglandin H (PGH) synthase results in a spectral shift in the Soret region. This shift was exploited to determine equilibrium and kinetic parameters of the cyanide binding process. At pH 8.0, ionic strength 0.22 M, 4 degrees C, the cyanide dissociation constant, determined from equilibrium experiments, is (65 +/- 10) microM. The binding rate constant is (2.8 +/- 0.2) x 10(3) M-1 s-1, and the dissociation rate constant is zero within experimental error. Through a kinetic study of the binding process as a function of pH, from pH 3.96 to 8.00, it was possible to determine the pKa of a heme-linked acid group on the enzyme of 4.15 +/- 0.10 with citrate buffer. An apparent pKa of 4.75 +/- 0.03 was determined with acetate buffer; this different value is attributed to complexation of the enzyme with one of the components of the acetate buffer.

Size and molecular flexibility affect the binding of ellagitannins to bovine serum albumin.

PubMed

Dobreva, Marina A; Green, Rebecca J; Mueller-Harvey, Irene; Salminen, Juha-Pekka; Howlin, Brendan J; Frazier, Richard A

2014-09-17

Binding to bovine serum albumin of monomeric (vescalagin and pedunculagin) and dimeric ellagitannins (roburin A, oenothein B, and gemin A) was investigated by isothermal titration calorimetry and fluorescence spectroscopy, which indicated two types of binding sites. Stronger and more specific sites exhibited affinity constants, K1, of 10(4)-10(6) M(-1) and stoichiometries, n1, of 2-13 and dominated at low tannin concentrations. Weaker and less-specific binding sites had K2 constants of 10(3)-10(5) M(-1) and stoichiometries, n2, of 16-30 and dominated at higher tannin concentrations. Binding to stronger sites appeared to be dependent on tannin flexibility and the presence of free galloyl groups. Positive entropies for all but gemin A indicated that hydrophobic interactions dominated during complexation. This was supported by an exponential relationship between the affinity, K1, and the modeled hydrophobic accessible surface area and by a linear relationship between K1 and the Stern-Volmer quenching constant, K(SV).

Measurement of monomolecular binding constants of neutral phenols into the beta-cyclodextrin by continuous frontal analysis in capillary and microchip electrophoresis via a competitive assay.

PubMed

Le Saux, Thomas; Hisamoto, Hideaki; Terabe, Shigeru

2006-02-03

Measurement of binding constant by chip electrophoresis is a very promising technique for the high throughput screening of non-covalent interactions. Among the different electrophoretic methods available that yield the binding parameters, continuous frontal analysis is the most appropriate for a transposition from capillary electrophoresis (CE) to microchip electrophoresis. Implementation of this methodology in microchip was exemplified by the measurement of inclusion constants of 2-naphtalenesulfonate and neutral phenols (phenol, 4-chlorophenol and 4-nitrophenol) into beta-cyclodextrin by competitive assays. The issue of competitor choice is discussed in relation to its appropriateness for proper monitoring of the interaction.

Circular dichroism spectroscopic investigation of double-decker phthalocyanine with G-Quadruplex as promising telomerase inhibitor

NASA Astrophysics Data System (ADS)

Baǧda, Efkan; Baǧda, Esra; Yabaş, Ebru

2017-01-01

In the present study, interaction of a double-decker phthalocyanine with two G-quadruplex DNA, Tel 21 and cMYC, was investigated. To the best of our knowledge, this is the first study about G-quadruplex-double decker phthalocyanine interaction. The spectrophotometric titration method was used for binding constant calculations. From the binding constants, it can be said that double-decker phthalocyanine more likely to bind Tel 21 rather than cMYC. The conformational changes upon binding were monitored via circular dichroism spectroscopy. The ethidium bromide replacement assay was investigated spectrofluorometrically.

Study of Proton Transfer in E. Coli Photolyase

NASA Astrophysics Data System (ADS)

Zhang, Meng; Liu, Zheyun; Li, Jiang; Wang, Lijuan; Zhong, Dongping

2013-06-01

Photolyase is a flavoprotein which utilizes blue-light energy to repair UV-light damaged DNA. The catalytic cofactor of photolyase, flavin adenine dinucleotide (FAD), has five redox states. Conversions between these redox states involve intraprotein electron transfer and proton transfer, which play important role in protein function. Here we systematically studied proton transfer in E. coli photolyase in vitro by site-directed mutagenesis and steady-state UV-vis spectroscopy, and proposed the proton channel in photolyase. We found that in the mutant N378C/E363L, proton channel was completely eliminated when DNA substrate was bound to the protein. Proton is suggested to be transported from protein surface to FAD by two pathways: the proton relay pathway through E363 and surface water to N378 and then to FAD; and the proton diffusion pathway through the substrate binding pocket. In addition, reaction kinetics of conversions between the redox states was then solved and redox potentials of the redox states were determined. These results described a complete picture of FAD redox changes, which are fundamental to the functions of all flavoenzymes.

The effects of structural variations of thiophene-containing Ru(II) complexes on the acid-base and DNA binding properties.

PubMed

Yuan, Cui-Li; Zhang, An-Guo; Zheng, Ze-Bo; Wang, Ke-Zhi

2013-03-01

A phenylthiophenyl-bearing Ru(II) complex of [Ru(bpy)₂(Hbptip)](PF₆)₂ {bpy = 2,2'-bipyridine, Hbptip = 2-(4-phenylthiophen-2-yl)-1H-imidazo[4,5-f][1,10]phenanthroline} was synthesized and characterized by elemental analysis, ¹H NMR spectroscopy, and electrospray ionization mass spectrometry. The ground- and excited-state acid-base properties of the complex were studied by UV-visible absorption and photoluminescence spectrophotometric pH titrations and the negative logarithm values of the ground-state acid ionization constants were derived to be pK(a1) = 1.31 ± 0.09 and pK(a2) = 5.71 ± 0.11 with the pK(a2) associated deprotonation/protonation process occurring over 3 pK(a) units more acidic than thiophenyl-free parent complex of [Ru(bpy)₂(Hpip)]²⁺ {Hpip = 2-phenyl-1H-imidazo[4,5-f][1,10]phenanthroline}. The calf thymus DNA-binding properties of [Ru(bpy)₂(Hbptip)]²⁺ in Tris-HCl buffer (pH 7.1 and 50 mM NaCl) were investigated by DNA viscosities and density functional theoretical calculations as well as UV-visible and emission spectroscopy techniques of UV-visible and luminescence titrations, steady-state emission quenching by [Fe(CN)₆]⁴⁻, DNA competitive binding with ethidium bromide, DNA melting experiments, and reverse salt effects. The complex was evidenced to bind to the DNA intercalatively with binding affinity being greater than those for previously reported analogs of [Ru(bpy)₂(Hip)]²⁺, [Ru(bpy)₂(Htip)]²⁺, and [Ru(bpy)₂(Haptip)]²⁺ {Hip = 1H-imidazo[4,5-f][1,10]phenanthroline, Htip = 2-thiophenimidazo[4,5-f][1,10]phenanthroline, Haptip = 2-(5-phenylthiophen-2-yl)-1H-imidazo[4,5-f][1,10]phenanthroline}.

Site-Directed Spin Labeling Reveals Pentameric Ligand-Gated Ion Channel Gating Motions

PubMed Central

Dellisanti, Cosma D.; Ghosh, Borna; Hanson, Susan M.; Raspanti, James M.; Grant, Valerie A.; Diarra, Gaoussou M.; Schuh, Abby M.; Satyshur, Kenneth; Klug, Candice S.; Czajkowski, Cynthia

2013-01-01

Pentameric ligand-gated ion channels (pLGICs) are neurotransmitter-activated receptors that mediate fast synaptic transmission. In pLGICs, binding of agonist to the extracellular domain triggers a structural rearrangement that leads to the opening of an ion-conducting pore in the transmembrane domain and, in the continued presence of neurotransmitter, the channels desensitize (close). The flexible loops in each subunit that connect the extracellular binding domain (loops 2, 7, and 9) to the transmembrane channel domain (M2–M3 loop) are essential for coupling ligand binding to channel gating. Comparing the crystal structures of two bacterial pLGIC homologues, ELIC and the proton-activated GLIC, suggests channel gating is associated with rearrangements in these loops, but whether these motions accurately predict the motions in functional lipid-embedded pLGICs is unknown. Here, using site-directed spin labeling (SDSL) electron paramagnetic resonance (EPR) spectroscopy and functional GLIC channels reconstituted into liposomes, we examined if, and how far, the loops at the ECD/TMD gating interface move during proton-dependent gating transitions from the resting to desensitized state. Loop 9 moves ∼9 Å inward toward the channel lumen in response to proton-induced desensitization. Loop 9 motions were not observed when GLIC was in detergent micelles, suggesting detergent solubilization traps the protein in a nonactivatable state and lipids are required for functional gating transitions. Proton-induced desensitization immobilizes loop 2 with little change in position. Proton-induced motion of the M2–M3 loop was not observed, suggesting its conformation is nearly identical in closed and desensitized states. Our experimentally derived distance measurements of spin-labeled GLIC suggest ELIC is not a good model for the functional resting state of GLIC, and that the crystal structure of GLIC does not correspond to a desensitized state. These findings advance our understanding of the molecular mechanisms underlying pLGIC gating. PMID:24260024

Potentiation of TRPM7 Inward Currents by Protons

PubMed Central

Jiang, Jianmin; Li, Mingjiang; Yue, Lixia

2005-01-01

TRPM7 is unique in being both an ion channel and a protein kinase. It conducts a large outward current at +100 mV but a small inward current at voltages ranging from −100 to −40 mV under physiological ionic conditions. Here we show that the small inward current of TRPM7 was dramatically enhanced by a decrease in extracellular pH, with an ∼10-fold increase at pH 4.0 and 1–2-fold increase at pH 6.0. Several lines of evidence suggest that protons enhance TRPM7 inward currents by competing with Ca2+ and Mg2+ for binding sites, thereby releasing blockade of divalent cations on inward monovalent currents. First, extracellular protons significantly increased monovalent cation permeability. Second, higher proton concentrations were required to induce 50% of maximal increase in TRPM7 currents when the external Ca2+ and Mg2+ concentrations were increased. Third, the apparent affinity for Ca2+ and Mg2+ was significantly diminished at elevated external H+ concentrations. Fourth, the anomalous-mole fraction behavior of H+ permeation further suggests that protons compete with divalent cations for binding sites in the TRPM7 pore. Taken together, it appears that at physiological pH (7.4), Ca2+ and Mg2+ bind to TRPM7 and inhibit the monovalent cationic currents; whereas at high H+ concentrations, the affinity of TRPM7 for Ca2+ and Mg2+ is decreased, thereby allowing monovalent cations to pass through TRPM7. Furthermore, we showed that the endogenous TRPM7-like current, which is known as Mg2+-inhibitable cation current (MIC) or Mg nucleotide–regulated metal ion current (MagNuM) in rat basophilic leukemia (RBL) cells was also significantly potentiated by acidic pH, suggesting that MIC/MagNuM is encoded by TRPM7. The pH sensitivity represents a novel feature of TRPM7 and implies that TRPM7 may play a role under acidic pathological conditions. PMID:16009728

Relativistic force field: parametric computations of proton-proton coupling constants in (1)H NMR spectra.

PubMed

Kutateladze, Andrei G; Mukhina, Olga A

2014-09-05

Spin-spin coupling constants in (1)H NMR carry a wealth of structural information and offer a powerful tool for deciphering molecular structures. However, accurate ab initio or DFT calculations of spin-spin coupling constants have been very challenging and expensive. Scaling of (easy) Fermi contacts, fc, especially in the context of recent findings by Bally and Rablen (Bally, T.; Rablen, P. R. J. Org. Chem. 2011, 76, 4818), offers a framework for achieving practical evaluation of spin-spin coupling constants. We report a faster and more precise parametrization approach utilizing a new basis set for hydrogen atoms optimized in conjunction with (i) inexpensive B3LYP/6-31G(d) molecular geometries, (ii) inexpensive 4-31G basis set for carbon atoms in fc calculations, and (iii) individual parametrization for different atom types/hybridizations, not unlike a force field in molecular mechanics, but designed for the fc's. With the training set of 608 experimental constants we achieved rmsd <0.19 Hz. The methodology performs very well as we illustrate with a set of complex organic natural products, including strychnine (rmsd 0.19 Hz), morphine (rmsd 0.24 Hz), etc. This precision is achieved with much shorter computational times: accurate spin-spin coupling constants for the two conformers of strychnine were computed in parallel on two 16-core nodes of a Linux cluster within 10 min.

IR spectroscopy of protonation in benzene-water nanoclusters: hydronium, zundel, and eigen at a hydrophobic interface.

PubMed

Cheng, Timothy C; Bandyopadhyay, Biswajit; Mosley, Jonathan D; Duncan, Michael A

2012-08-08

The structure of ions in water at a hydrophobic interface influences important processes throughout chemistry and biology. However, experiments to measure these structures are limited by the distribution of configurations present and the inability to selectively probe the interfacial region. Here, protonated nanoclusters containing benzene and water are produced in the gas phase, size-selected, and investigated with infrared laser spectroscopy. Proton stretch, free OH, and hydrogen-bonding vibrations uniquely define protonation sites and hydrogen-bonding networks. The structures consist of protonated water clusters binding to the hydrophobic interface of neutral benzene via one or more π-hydrogen bonds. Comparison to the spectra of isolated hydronium, zundel, or eigen ions reveals the inductive effects and local ordering induced by the interface. The structures and interactions revealed here represent key features expected for aqueous hydrophobic interfaces.

Kinetic studies of amino acid-based surfactant binding to DNA.

PubMed

Santhiya, Deenan; Dias, Rita S; Dutta, Sounak; Das, Prasanta Kumar; Miguel, Maria G; Lindman, Björn; Maiti, Souvik

2012-05-24

In this work, the binding kinetics of amino acid-based surfactants, presenting different linkers and head groups, with calf thymus (CT)-DNA was studied using stopped-flow fluorescence spectroscopy. The kinetic studies were carried out as a function of Na(+) concentration and surfactant-to-DNA charge ratio. The surfactant binding on DNA took place in two consecutive steps, for which the corresponding first and second relative rate constants (k(1) and k(2)) were determined. The fast step was attributed to the surfactant binding to DNA and micelle formation in its vicinity, the slower step to DNA condensation and possible rearrangement of the surfactant aggregates. In general, both relative rate constants increase with surfactant concentration and decrease with the ionic strength of the medium. The architecture of the surfactant was found to have a significant impact on the kinetics of the DNA-surfactant complexation. Surfactants with amide linkers showed larger relative rate constants than those with ester linkers. The variation of the relative rate constants with the head groups of the surfactants, alanine and proline, was found to be less obvious, being partially dependent on the surfactant concentration.

NMR and rotational angles in solution conformation of polypeptides

NASA Astrophysics Data System (ADS)

Bystrov, V. F.

1985-01-01

Professor San-Ichiro Mizushima and Professor Yonezo Morino's classical contributions provided unique means and firm basis for understanding of conformational states and internal rotation in polypeptide molecules. Now the NMR spectroscopy is the best choice to study molecular conformation, mechanism of action and structure-functional relationships of peptide and proteins in solution under conditions approaching those of their physiological environments. Crucial details of spatial structure and interactions of these molecules in solution are revealed by using proton-proton and carbon-proton vicinal coupling constants, proton nuclear Overhauser effect and spectral perturbation techniques. The results of NMR conformational analysis are presented for valinomycin "bracelet", gramicidin A double helices, honey-bee neurotoxin apamin, scorpion insectotoxins and snake neurotoxins of long and short types.

On the role of water density fluctuations in the inhibition of a proton channel

PubMed Central

Gianti, Eleonora; Delemotte, Lucie; Klein, Michael L.; Carnevale, Vincenzo

2016-01-01

Hv1 is a transmembrane four-helix bundle that transports protons in a voltage-controlled manner. Its crucial role in many pathological conditions, including cancer and ischemic brain damage, makes Hv1 a promising drug target. Starting from the recently solved crystal structure of Hv1, we used structural modeling and molecular dynamics simulations to characterize the channel’s most relevant conformations along the activation cycle. We then performed computational docking of known Hv1 inhibitors, 2-guanidinobenzimidazole (2GBI) and analogs. Although salt-bridge patterns and electrostatic potential profiles are well-defined and distinctive features of activated versus nonactivated states, the water distribution along the channel lumen is dynamic and reflects a conformational heterogeneity inherent to each state. In fact, pore waters assemble into intermittent hydrogen-bonded clusters that are replaced by the inhibitor moieties upon ligand binding. The entropic gain resulting from releasing these conformationally restrained waters to the bulk solvent is likely a major contributor to the binding free energy. Accordingly, we mapped the water density fluctuations inside the pore of the channel and identified the regions of maximum fluctuation within putative binding sites. Two sites appear as outstanding: One is the already known binding pocket of 2GBI, which is accessible to ligands from the intracellular side; the other is a site located at the exit of the proton permeation pathway. Our analysis of the waters confined in the hydrophobic cavities of Hv1 suggests a general strategy for drug discovery that can be applied to any ion channel. PMID:27956641

Chemical Rescue of Enzymes: Proton Transfer in Mutants of Human Carbonic Anhydrase II

PubMed Central

Maupin, C. Mark; Castillo, Norberto; Taraphder, Srabani; Tu, Chingkuang; McKenna, Robert; Silverman, David N.; Voth, Gregory A.

2011-01-01

In human carbonic anhydrase II (HCA II) the mutation of position 64 from histidine to alanine (H64A) disrupts the rate limiting proton transfer (PT) event, resulting in a reduction of the catalytic activity of the enzyme as compared to the wild-type. Potential of mean force (PMF) calculations utilizing the multistate empirical valence bond (MS-EVB) methodology for H64A HCA II give a PT free energy barrier significantly higher than that found in the wild-type enzyme. This high barrier, determined in the absence of exogenous buffer and assuming no additional ionizable residues in the PT pathway, indicates the likelihood of alternate enzyme pathways that utilize either ionizable enzyme residues (self-rescue) and/or exogenous buffers (chemical rescue). It has been shown experimentally that the catalytic activity of H64A HCA II can be chemically rescued to near wild type levels by the addition of the exogenous buffer 4-methylimidazole (4MI). Crystallographic studies have identified two 4MI binding sites, yet site specific mutations intended to disrupt 4MI binding have demonstrated these sites to be non-productive. In the present work MS-EVB simulations show that binding of 4MI near Thr199 in the H64A HCA II mutant, a binding site determined by NMR spectroscopy, results in a viable chemical rescue pathway. Additional viable rescue pathways are also identified where 4MI acts as a proton transport intermediary from the active site to ionizable residues on the rim of the active site, revealing a probable mode of action for the chemical rescue pathway PMID:21452838

High-affinity binding of laurate to naturally occurring mutants of human serum albumin and proalbumin.

PubMed

Kragh-Hansen, U; Pedersen, A O; Galliano, M; Minchiotti, L; Brennan, S O; Tárnoky, A L; Franco, M H; Salzano, F M

1996-12-15

Binding of laurate (n-dodecanoate) to genetic variants of albumin or its proprotein and to normal albumin isolated from the same heterozygous carriers was studied by a kinetic dialysis technique at physiological pH. The first stoichiometric association constant for binding to proalbumin Lille (Arg-2-->His) and albumin (Alb) Roma (Glu321-->Lys) was increased to 126% and 136% respectively compared with that for binding to normal albumin, whereas the constant for Alb Maku (Lys541-->Glu) was decreased to 80%. In contrast, normal laurate-binding properties were found for as many as nine other albumin variants with single amino acid substitutions. Because the net charges of all these mutants were different from that of normal albumin, the results suggest that the examples of modified laurate binding are not caused by long-range electrostatic effects. Rather, the three positions mentioned are located close to different binding sites for the fatty acid anion. The most pronounced effect was observed for the glycosylated Alb Casebrook, the binding constant of which was decreased to 20%. Binding to the glycosylated Alb Redhill was also decreased, but to a smaller extent (68%). These decreases in binding are caused by partial or total blocking of the high-affinity site by the oligosaccharides, by the negative charges of the oligosaccharides, and/or by conformational changes induced by these bulky moieties. Laurate binding to two chain-termination mutants (Alb Catania and Alb Venezia) was normal, indicating that the C-terminus of albumin is not important for binding. By using different preparations of normal albumin as controls in the binding experiments, it was also possible to compare the effect of various methods for isolation and defatting on laurate binding.

Alchemical free energy simulations for biological complexes: powerful but temperamental....

PubMed

Aleksandrov, Alexey; Thompson, Damien; Simonson, Thomas

2010-01-01

Free energy simulations compare multiple ligand:receptor complexes by "alchemically" transforming one into another, yielding binding free energy differences. Since their introduction in the 1980s, many technical and theoretical obstacles were surmounted, and the method ("MDFE," since molecular dynamics are often used) has matured into a powerful tool. We describe its current status, its effectiveness, and the challenges it faces. MDFE has provided chemical accuracy for many systems but remains expensive, with significant human overhead costs. The bottlenecks have shifted, partly due to increased computer power. To study diverse sets of ligands, force field availability and accuracy can be a major difficulty. Another difficulty is the frequent need to consider multiple states, related to sidechain protonation or buried waters, for example. Sophisticated, automated methods to sample these states are maturing, such as constant pH simulations. Meanwhile, combinations of MDFE and simpler approaches, like continuum dielectric models, can be very effective. As illustrations, we show how, with careful force field parameterization, MDFE accurately predicts binding specificities between complex tetracycline ligands and their targets. We describe substrate binding to the aspartyl-tRNA synthetase enzyme, where many distinct electrostatic states play a role, and a histidine and a Mg(2+) ion act as coupled switches that help enforce a strict preference for the aspartate substrate, relative to several analogs. Overall, MDFE has achieved a predictive status, where novel ligands can be studied and molecular recognition elucidated in depth. It should play an increasing role in the analysis of complex cellular processes and biomolecular engineering. 2009 John Wiley & Sons, Ltd.

Bioinformatic Analysis of the Contribution of Primer Sequences to Aptamer Structures

PubMed Central

Ellington, Andrew D.

2009-01-01

Aptamers are nucleic acid molecules selected in vitro to bind a particular ligand. While numerous experimental studies have examined the sequences, structures, and functions of individual aptamers, considerably fewer studies have applied bioinformatics approaches to try to infer more general principles from these individual studies. We have used a large Aptamer Database to parse the contributions of both random and constant regions to the secondary structures of more than 2000 aptamers. We find that the constant, primer-binding regions do not, in general, contribute significantly to aptamer structures. These results suggest that (a) binding function is not contributed to nor constrained by constant regions; (b) in consequence, the landscape of functional binding sequences is sparse but robust, favoring scenarios for short, functional nucleic acid sequences near origins; and (c) many pool designs for the selection of aptamers are likely to prove robust. PMID:18594898

Spectrophotometric analysis of flavonoid-DNA binding interactions at physiological conditions

NASA Astrophysics Data System (ADS)

Janjua, Naveed Kausar; Siddiqa, Asima; Yaqub, Azra; Sabahat, Sana; Qureshi, Rumana; Haque, Sayed ul

2009-12-01

Mode of interactions of three flavonoids [morin (M), quercetin (Q), and rutin (R)] with chicken blood ds.DNA (ck.DNA) has been investigated spectrophotometrically at different temperatures including body temperature (310 K) and at two physiological pH values, i.e. 7.4 (human blood pH) and 4.7 (stomach pH). The binding constants, Kf, evaluated using Benesi-Hildebrand equation showed that the flavonoids bind effectively through intercalation at both pH values and body temperature. Quercetin, somehow, showed greater binding capabilities with DNA. The free energies of flavonoid-DNA complexes indicated the spontaneity of their binding. The order of binding constants of three flavonoids at both pH values were found to be Kf(Q) > Kf(R) > Kf(M) and at 310 K.

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

Negative Dielectric Constant Material Based on Ion Conducting Materials

NASA Technical Reports Server (NTRS)

Gordon, Keith L. (Inventor); Kang, Jin Ho (Inventor); Park, Cheol (Inventor); Lillehei, Peter T. (Inventor); Harrison, Joycelyn S. (Inventor)

2017-01-01

Metamaterials or artificial negative index materials (NIMs) have generated great attention due to their unique and exotic electromagnetic properties. One exemplary negative dielectric constant material, which is an essential key for creating the NIMs, was developed by doping ions into a polymer, a protonated poly (benzimidazole) (PBI). The doped PBI showed a negative dielectric constant at megahertz (MHz) frequencies due to its reduced plasma frequency and an induction effect. The magnitude of the negative dielectric constant and the resonance frequency were tunable by doping concentration. The highly doped PBI showed larger absolute magnitude of negative dielectric constant at just above its resonance frequency than the less doped PBI.

Negative Dielectric Constant Material Based on Ion Conducting Materials

NASA Technical Reports Server (NTRS)

Gordon, Keith L. (Inventor); Kang, Jin Ho (Inventor); Harrison, Joycelyn S. (Inventor); Park, Cheol (Inventor); Lillehei, Peter T. (Inventor)

2014-01-01

Metamaterials or artificial negative index materials (NIMs) have generated great attention due to their unique and exotic electromagnetic properties. One exemplary negative dielectric constant material, which is an essential key for creating the NIMs, was developed by doping ions into a polymer, a protonated poly(benzimidazole) (PBI). The doped PBI showed a negative dielectric constant at megahertz (MHz) frequencies due to its reduced plasma frequency and an induction effect. The magnitude of the negative dielectric constant and the resonance frequency were tunable by doping concentration. The highly doped PBI showed larger absolute magnitude of negative dielectric constant at just above its resonance frequency than the less doped PBI.

Vicinal fluorine-fluorine coupling constants: Fourier analysis.

PubMed

San Fabián, J; Westra Hoekzema, A J A

2004-10-01

Stereochemical dependences of vicinal fluorine-fluorine nuclear magnetic resonance coupling constants (3JFF) have been studied with the multiconfigurational self-consistent field in the restricted active space approach, with the second-order polarization propagator approximation (SOPPA), and with density functional theory. The SOPPA results show the best overall agreement with experimental couplings. The relationship with the dihedral angle between the coupled fluorines has been studied by Fourier analysis, the result is very different from that of proton-proton couplings. The Fourier coefficients do not resemble those of a typical Karplus equation. The four nonrelativistic contributions to the coupling constants of 1,2-difluoroethane configurations have been studied separately showing that up to six Fourier coefficients are required to reproduce the calculated values satisfactorily. Comparison with Fourier coefficients for matching hydrogen fluoride dimer configurations suggests that the higher order Fourier coefficients (Cn> or =3) originate mainly from through-space Fermi contact interaction. The through-space interaction is the main reason 3JFF do not follow the Karplus equation. (c) 2004 American Institute of Physics

Vicinal fluorine-fluorine coupling constants: Fourier analysis

NASA Astrophysics Data System (ADS)

San Fabián, J.; Westra Hoekzema, A. J. A.

2004-10-01

Stereochemical dependences of vicinal fluorine-fluorine nuclear magnetic resonance coupling constants (3JFF) have been studied with the multiconfigurational self-consistent field in the restricted active space approach, with the second-order polarization propagator approximation (SOPPA), and with density functional theory. The SOPPA results show the best overall agreement with experimental couplings. The relationship with the dihedral angle between the coupled fluorines has been studied by Fourier analysis, the result is very different from that of proton-proton couplings. The Fourier coefficients do not resemble those of a typical Karplus equation. The four nonrelativistic contributions to the coupling constants of 1,2-difluoroethane configurations have been studied separately showing that up to six Fourier coefficients are required to reproduce the calculated values satisfactorily. Comparison with Fourier coefficients for matching hydrogen fluoride dimer configurations suggests that the higher order Fourier coefficients (Cn⩾3) originate mainly from through-space Fermi contact interaction. The through-space interaction is the main reason 3JFF do not follow the Karplus equation.

Measuring Positive Cooperativity Using the Direct ESI-MS Assay. Cholera Toxin B Subunit Homopentamer Binding to GM1 Pentasaccharide

NASA Astrophysics Data System (ADS)

Lin, Hong; Kitova, Elena N.; Klassen, John S.

2014-01-01

Direct electrospray ionization mass spectrometry (ESI-MS) assay was used to investigate the stepwise binding of the GM1 pentasaccharide β- D-Gal p-(1→3)-β-D-Gal pNAc-(1→4)[α-D-Neu5Ac-(2→3)]-β- D-Gal p-(1→4)-β-D-Glc p (GM1os) to the cholera toxin B subunit homopentamer (CTB5) and to establish conclusively whether GM1os binding is cooperative. Apparent association constants were measured for the stepwise addition of one to five GM1os to CTB5 at pH 6.9 and 22 °C. The intrinsic association constant, which was established from the apparent association constant for the addition of a single GM1os to CTB5, was found to be (3.2 ± 0.2) × 106 M-1. This is in reasonable agreement with the reported value of (6.4 ± 0.3) × 106 M-1, which was measured at pH 7.4 and 25 °C using isothermal titration calorimetry (ITC). Analysis of the apparent association constants provides direct and unambiguous evidence that GM1os binding exhibits small positive cooperativity. Binding was found to be sensitive to the number of ligand-bound nearest neighbor subunits, with the affinities enhanced by a factor of 1.7 and 2.9 when binding occurs next to one or two ligand-bound subunits, respectively. These findings, which provide quantitative support for the binding model proposed by Homans and coworkers [14], highlight the unique strengths of the direct ESI-MS assay for measuring cooperative ligand binding.

2D IR spectroscopy reveals the role of water in the binding of channel-blocking drugs to the influenza M2 channel.

PubMed

Ghosh, Ayanjeet; Wang, Jun; Moroz, Yurii S; Korendovych, Ivan V; Zanni, Martin; DeGrado, William F; Gai, Feng; Hochstrasser, Robin M

2014-06-21

Water is an integral part of the homotetrameric M2 proton channel of the influenza A virus, which not only assists proton conduction but could also play an important role in stabilizing channel-blocking drugs. Herein, we employ two dimensional infrared (2D IR) spectroscopy and site-specific IR probes, i.e., the amide I bands arising from isotopically labeled Ala30 and Gly34 residues, to probe how binding of either rimantadine or 7,7-spiran amine affects the water dynamics inside the M2 channel. Our results show, at neutral pH where the channel is non-conducting, that drug binding leads to a significant increase in the mobility of the channel water. A similar trend is also observed at pH 5.0 although the difference becomes smaller. Taken together, these results indicate that the channel water facilitates drug binding by increasing its entropy. Furthermore, the 2D IR spectral signatures obtained for both probes under different conditions collectively support a binding mechanism whereby amantadine-like drugs dock in the channel with their ammonium moiety pointing toward the histidine residues and interacting with a nearby water cluster, as predicted by molecular dynamics simulations. We believe these findings have important implications for designing new anti-influenza drugs.

Porphyrin-substrate binding to murine ferrochelatase: effect on the thermal stability of the enzyme

PubMed Central

2004-01-01

Ferrochelatase (EC 4.99.1.1), the terminal enzyme of the haem biosynthetic pathway, catalyses the chelation of Fe(II) into the protoporphyrin IX ring. The energetics of the binding between murine ferrochelatase and mesoporphyrin were determined using isothermal titration calorimetry, which revealed a stoichiometry of one molecule of mesoporphyrin bound per protein monomer. The binding is strongly exothermic, with a large intrinsic enthalpy (ΔH=−97.1 kJ · mol−1), and is associated with the uptake of two protons from the buffer. This proton transfer suggests that hydrogen bonding between ferrochelatase and mesoporphyrin is a key factor in the thermodynamics of the binding reaction. Differential scanning calorimetry thermograms indicated a co-operative two-state denaturation process with a single transition temperature of 56 °C for wild-type murine ferrochelatase. An increase in the thermal stability of ferrochelatase is dependent upon mesoporphyrin binding. Similarly, murine ferrochelatase variants, in which the active site Glu-289 was replaced by either glutamine or alanine and, when purified, contained specifically-bound protoporphyrin, exhibited enhanced protein stability when compared with wild-type ferrochelatase. However, in contrast with the wild-type enzyme, the thermal denaturation of ferrochelatase variants was best described as a non-co-operative denaturation process. PMID:15496139

The design features cells use to build their transmembrane proton gradient

NASA Astrophysics Data System (ADS)

Gunner, M. R.; Koder, Ronald

2017-02-01

Organisms store energy from food and sunlight as an electrochemical gradient across the membranes of mitochondria, chloroplasts and bacteria. The gradient arises from differences in the concentration of protons and ions on the negative (N) and positive (P) sides of these membranes. This perspective describes how the proton gradient is formed. One strategy is the movement of electrons but not protons across a membrane-embedded protein from a site of proton-releasing oxidative chemistry on the P-side of the protein to a site of proton-binding reductive chemistry on the N-side. Alternately, protons are directly pumped across membrane-embedded proteins, which have gated proton transfer pathways that are opened and closed, as well as internal sites where the proton affinity varies as the protein goes through the reaction cycle. The molecules that carry out these roles are complex, utilizing non-amino acid cofactors and earth-abundant metals. However, these are also potential sources of high-energy toxic byproducts. Understanding these reactions can open the door to their rational redesign, with possible beneficial effects as far-reaching as improving the global food supply, preventing neurodegenerative diseases, and better understanding the role of metabolism in aging.

The design features cells use to build their transmembrane proton gradient.

PubMed

Gunner, M R; Koder, Ronald

2017-02-07

Organisms store energy from food and sunlight as an electrochemical gradient across the membranes of mitochondria, chloroplasts and bacteria. The gradient arises from differences in the concentration of protons and ions on the negative (N) and positive (P) sides of these membranes. This perspective describes how the proton gradient is formed. One strategy is the movement of electrons but not protons across a membrane-embedded protein from a site of proton-releasing oxidative chemistry on the P-side of the protein to a site of proton-binding reductive chemistry on the N-side. Alternately, protons are directly pumped across membrane-embedded proteins, which have gated proton transfer pathways that are opened and closed, as well as internal sites where the proton affinity varies as the protein goes through the reaction cycle. The molecules that carry out these roles are complex, utilizing non-amino acid cofactors and earth-abundant metals. However, these are also potential sources of high-energy toxic byproducts. Understanding these reactions can open the door to their rational redesign, with possible beneficial effects as far-reaching as improving the global food supply, preventing neurodegenerative diseases, and better understanding the role of metabolism in aging.

The pH-sensitive structure of the C-terminal domain of voltage-gated proton channel and the thermodynamic characteristics of Zn{sup 2+} binding to this domain

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

Zhao, Qing; Li, Chuanyong; Li, Shu Jie, E-mail: shujieli@nankai.edu.cn

2015-01-02

Highlights: • The α-helical content of the C-terminus is decreased with a pH increase. • The thermostability of the C-terminus is decreased with a pH increase. • Zn{sup 2+} binds to His{sup 244} and His{sup 266} residues within the C-terminal domain. • The binding of Zn{sup 2+} to His{sup 244} residue is an endothermic heat reaction. • The binding of Zn{sup 2+} to His{sup 266} residue is an exothermic heat reaction. - Abstract: The voltage-gated proton channel Hv1 is strongly sensitive to Zn{sup 2+}. The H{sup +} conduction is decreased at a high concentration of Zn{sup 2+} and Hv1 channelmore » closing is slowed by the internal application of Zn{sup 2+}. Although the recent studies demonstrated that Zn{sup 2+} interacts with the intracellular C-terminal domain, the binding sites and details of the interaction remain unknown. Here, we studied the pH-dependent structural stability of the intracellular C-terminal domain of human Hv1 and showed that Zn{sup 2+} binds to His{sup 244} and His{sup 266} residues. The thermodynamics signature of Zn{sup 2+} binding to the two sites was investigated by isothermal titration calorimetry. The binding of Zn{sup 2+} to His{sup 244} (mutant H266A) and His{sup 266} (mutant H244A) were an endothermic heat reaction and an exothermic heat reaction, respectively.« less

NMR studies of multiple conformations in complexes of Lactobacillus casei dihydrofolate reductase with analogues of pyrimethamine

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

Birdsall, B.; Tendler, S.J.B.; Feeney, J.

1990-10-01

{sup 1}H and {sup 19}F NMR signals from bound ligands have been assigned in one- and two-dimensional NMR spectra of complexes of Lactobacillus casei dihydrofolate reductase with various pyrimethamine analogues. The signals were identified mainly by correlating signals from bound and free ligands by using 2D exchange experiments. Analogues with symmetrically substituted phenyl rings give rise to {sup 1}H signals from four nonequivalent aromatic protons, clearly indicating the presence of hindered rotation about the pyrimidine-phenyl bond. Analogues with symmetrically substituted phenyl rings give rise to {sup 1}H signals from four nonequivalent aromatic protons, clearly indicating the presence of hindered rotationmore » about the pyrimidine-phenyl bond. Analogues containing asymmetrically substituted aromatic rings exist as mixtures of two rotational isomers (an enantiomeric pair) because of this hindered rotation and the NMR spectra revealed that both isomers (forms A and B) bind to the enzyme with comparable, though unequal, binding energies. In this case two complete sets of bound proton signals were observed. The relative orientations of the two forms have been determined from NOE through-space connections between protons on the ligand and protein. Ternary complexes with NADP{sup {plus}} were also examined.« less

Neutron structures of the Helicobacter pylori 5'-methylthioadenosine nucleosidase highlight proton sharing and protonation states

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

Banco, Michael T.; Mishra, Vidhi; Ostermann, Andreas

2016-11-16

MTAN (5'-methylthioadenosine nucleosidase) catalyzes the hydrolysis of the N-ribosidic bond of a variety of adenosine-containing metabolites. The Helicobacter pylori MTAN (HpMTAN) hydrolyzes 6-amino-6-deoxyfutalosine in the second step of the alternative menaquinone biosynthetic pathway. Substrate binding of the adenine moiety is mediated almost exclusively by hydrogen bonds, and the proposed catalytic mechanism requires multiple proton-transfer events. Of particular interest is the protonation state of residue D198, which possesses a pKa above 8 and functions as a general acid to initiate the enzymatic reaction. In this study we present three corefined neutron/X-ray crystal structures of wild-type HpMTAN cocrystallized with S-adenosylhomocysteine (SAH), Formycinmore » A (FMA), and (3R,4S)-4-(4-Chlorophenylthiomethyl)-1-[(9-deaza-adenin-9-yl)methyl]-3-hydroxypyrrolidine (p-ClPh-Thio-DADMe-ImmA) as well as one neutron/X-ray crystal structure of an inactive variant (HpMTAN-D198N) cocrystallized with SAH. These results support a mechanism of D198 pKa elevation through the unexpected sharing of a proton with atom N7 of the adenine moiety possessing unconventional hydrogen-bond geometry. Additionally, the neutron structures also highlight active site features that promote the stabilization of the transition state and slight variations in these interactions that result in 100-fold difference in binding affinities between the DADMe-ImmA and ImmA analogs.« less

Detailed Analysis of Apoptosis and Delayed Luminescence of Human Leukemia Jurkat T Cells after Proton Irradiation and Treatments with Oxidant Agents and Flavonoids

PubMed Central

Baran, Irina; Ganea, Constanta; Privitera, Simona; Scordino, Agata; Barresi, Vincenza; Musumeci, Francesco; Mocanu, Maria Magdalena; Condorelli, Daniele F.; Ursu, Ioan; Grasso, Rosaria; Gulino, Marisa; Garaiman, Alexandru; Musso, Nicolò; Cirrone, Giuseppe A. Pablo; Cuttone, Giacomo

2012-01-01

Following previous work, we investigated in more detail the relationship between apoptosis and delayed luminescence (DL) in human leukemia Jurkat T cells under a wide variety of treatments. We used menadione and hydrogen peroxide to induce oxidative stress and two flavonoids, quercetin, and epigallocatechin gallate, applied alone or in combination with menadione or H2O2. 62 MeV proton beams were used to irradiate cells under a uniform dose of 2 or 10 Gy, respectively. We assessed apoptosis, cell cycle distributions, and DL. Menadione, H2O2 and quercetin were potent inducers of apoptosis and DL inhibitors. Quercetin decreased clonogenic survival and the NAD(P)H level in a dose-dependent manner. Proton irradiation with 2 Gy but not 10 Gy increased the apoptotic rate. However, both doses induced a substantial G2/M arrest. Quercetin reduced apoptosis and prolonged the G2/M arrest induced by radiation. DL spectroscopy indicated that proton irradiation disrupted the electron flow within Complex I of the mitochondrial respiratory chain, thus explaining the massive necrosis induced by 10 Gy of protons and also suggested an equivalent action of menadione and quercetin at the level of the Fe/S center N2, which may be mediated by their binding to a common site within Complex I, probably the rotenone-binding site. PMID:22829956

Neutron structures of the Helicobacter pylori 5'-methylthioadenosine nucleosidase highlight proton sharing and protonation states

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

Banco, Michael T.; Mishra, Vidhi; Ostermann, Andreas

MTAN (5'-methylthioadenosine nucleosidase) catalyzes the hydrolysis of the N-ribosidic bond of a variety of adenosine-containing metabolites. The Helicobacter pylori MTAN (HpMTAN) hydrolyzes 6-amino-6-deoxyfutalosine in the second step of the alternative menaquinone biosynthetic pathway. Substrate binding of the adenine moiety is mediated almost exclusively by hydrogen bonds, and the proposed catalytic mechanism requires multiple proton-transfer events. Of particular interest is the protonation state of residue D198, which possesses a pK a above 8 and functions as a general acid to initiate the enzymatic reaction. In this study we present three corefined neutron/X-ray crystal structures of wild-type HpMTAN cocrystallized with S-adenosylhomocysteine (SAH),more » Formycin A (FMA), and (3R,4S)-4-(4-Chlorophenylthiomethyl)-1-[(9-deaza-adenin-9-yl)methyl]-3-hydroxypyrrolidine (p-ClPh-Thio-DADMe-ImmA) as well as one neutron/X-ray crystal structure of an inactive variant (HpMTAN-D198N) cocrystallized with SAH. These results support a mechanism of D198 pKa elevation through the unexpected sharing of a proton with atom N7 of the adenine moiety possessing unconventional hydrogen-bond geometry. Additionally, the neutron structures also highlight active site features that promote the stabilization of the transition state and slight variations in these interactions that result in 100-fold difference in binding affinities between the DADMe-ImmA and ImmA analogs.« less

Neutron structures of the Helicobacter pylori 5'-methylthioadenosine nucleosidase highlight proton sharing and protonation states

DOE PAGES

Banco, Michael T.; Mishra, Vidhi; Ostermann, Andreas; ...

2016-10-01

MTAN (5'-methylthioadenosine nucleosidase) catalyzes the hydrolysis of the N-ribosidic bond of a variety of adenosine-containing metabolites. The Helicobacter pylori MTAN (HpMTAN) hydrolyzes 6-amino-6-deoxyfutalosine in the second step of the alternative menaquinone biosynthetic pathway. Substrate binding of the adenine moiety is mediated almost exclusively by hydrogen bonds, and the proposed catalytic mechanism requires multiple proton-transfer events. Of particular interest is the protonation state of residue D198, which possesses a pK a above 8 and functions as a general acid to initiate the enzymatic reaction. In this study we present three corefined neutron/X-ray crystal structures of wild-type HpMTAN cocrystallized with S-adenosylhomocysteine (SAH),more » Formycin A (FMA), and (3R,4S)-4-(4-Chlorophenylthiomethyl)-1-[(9-deaza-adenin-9-yl)methyl]-3-hydroxypyrrolidine (p-ClPh-Thio-DADMe-ImmA) as well as one neutron/X-ray crystal structure of an inactive variant (HpMTAN-D198N) cocrystallized with SAH. These results support a mechanism of D198 pKa elevation through the unexpected sharing of a proton with atom N7 of the adenine moiety possessing unconventional hydrogen-bond geometry. Additionally, the neutron structures also highlight active site features that promote the stabilization of the transition state and slight variations in these interactions that result in 100-fold difference in binding affinities between the DADMe-ImmA and ImmA analogs.« less

Protonation of key acidic residues is critical for the K+-selectivity of the Na/K pump

PubMed Central

Yu, Haibo; Ratheal, Ian; Artigas, Pablo; Roux, Benoît

2011-01-01

The sodium-potassium (Na/K) pump is a P-type ATPase that generates Na+ and K+ concentration gradients across the cell membrane. For each ATP molecule, the pump extrudes three Na+ and imports two K+ by alternating between outward- and inward-facing conformations that preferentially bind K+ or Na+, respectively. Remarkably, the selective K+ and Na+ binding sites share several residues, and how the pump is able to achieve the selectivity required for the functional cycle is unclear. Here, free energy perturbation molecular dynamics (FEP/MD) simulations based on the crystal structures of the Na/K pump in a K+-loaded state (E2·Pi) reveal that protonation of the high-field acidic side-chains involved in the binding sites is critical to achieve the proper K+ selectivity. This prediction is tested with electrophysiological experiments showing that the selectivity of the E2P state for K+ over Na+ is affected by extracellular pH. PMID:21909093

Mineral catalysis of the formation of the phosphodiester bond in aqueous solution - The possible role of montmorillonite clays

NASA Technical Reports Server (NTRS)

Ferris, James P.; Ertem, Gozen; KAMALUDDIN; Agarwal, Vipin; Hua, Lu Lin

1989-01-01

The possible role of montmorillonite clays in the spontaneous formation on the primitive earth of the phosphodiester bond in the presence of water was investigated in experiments measuring the binding of various nucleosides and nucleotides with Na(+)-montmorillonite 22A and the reactions of these compounds with a water-soluble carbodiimide. It was found that, at neutral pH, adenine derivatives bind stronger than the corresponding uracil derivatives, consistent with the protonation of the adenine by the acidic clay surface and a cationic binding of the protonated ring to the anionic clay surface. The reaction of the 5-prime-AMP with carbodiimide resulted in the formation of 2-prime,5-prime-pApA (18.9 percent), 3-prime,5-prime-pApA (11 percent), and AppA (4.8 percent). The yields of these oligomers obtained when poly(U) was used in place of the clay were 15.5 percent, 3.7 percent, and 14.9 percent AppA, respectively.

The kinetics of the decompositions of the proton bound dimers of 1,4-dimethylpyridine and dimethyl methylphosphonate from atmospheric pressure ion mobility spectra

NASA Astrophysics Data System (ADS)

Ewing, R. G.; Eiceman, G. A.; Harden, C. S.; Stone, J. A.

2006-09-01

The rate constants for the dissociations, A2H+ --> AH+ + A, of the symmetrical proton bound dimers of 2,4-dimethylpyridine and dimethyl methylphosphonate have been determined using an ion mobility spectrometer operating with air as drift gas at ambient pressure. Reaction time was varied by varying the drift electric field. The rate constants were derived from the mobility spectra by determining the rate at which ions decomposed in the drift region. Arrhenius plots with a drift gas containing water vapor at 5 ppmv gave the following activation energies and pre-exponential factors: 2,4-dimethylpyridine, 94 +/- 2 kJ mol-1, log A (s-1) = 15.9 +/- 0.4; dimethyl methylphosphonate, 127 +/- 3 kJ mol-1, log A (s-1) = 15.6 +/- 0.3. The enthalpy changes for the decompositions calculated from the activation energies are in accord with literature values for symmetrical proton bound dimers of oxygen and nitrogen bases. The results for dimethyl methylphosphonate were obtained over the temperature range 478-497 K and are practically independent of water concentration (5-2000 ppmv). The activation energy for 2,4-dimethylpyridine, obtained over the temperature range 340-359 K, decreased to 31 kJ mol-1 in the presence of 2.0 x 103 ppmv of water. At the low temperature, a displacement reaction involving water may account for the decrease. The reduced mobilities of the protonated molecules and the proton bound dimers have been determined over a wide temperature range. While the values for the dimers are essentially independent of the water concentration in the drift gas, those of the protonated molecules show a strong dependence.

Structural and photodynamic properties of the anti-cancer drug irinotecan in aqueous solutions of different pHs.

PubMed

di Nunzio, Maria Rosaria; Douhal, Yasmin; Organero, Juan Angel; Douhal, Abderrazzak

2018-05-23

This work reports on photophysical studies of the irinotecan (IRT) anti-cancer drug in water solutions of different acidities (pH = 1.11-9.46). We found that IRT co-exists as mono-cationic (C1), di-cationic (C2), or neutral (N) forms. The population of each prototropic species depends on the pH of the solution. At pH = 1.11-3.01, the C1 and C2 structures are stabilized. At pH = 7.00, the most populated species is C1, while at pH values larger than 9.46 the N form is the most stable species. In the 1.11-2.61 pH range, the C1* emission is efficiently quenched by protons to give rise to the emission from C2*. The dynamic quenching constant, KD, is ∼32 M-1. While the diffusion governs the rate of excited-state proton-transfer (ESPT) under these conditions, the reaction rate increases with the proton concentration. A two-step diffusive Debye-Smoluchowski model was applied at pH = 1.11-2.61 to describe the protonation of C1*. The ESPT time constants derived for C1* are 382 and 1720 ps at pH = 1.11 and 1.95, respectively. We found that one proton species is involved in the protonation of C1* to give C2*, in the analyzed acidic pH range. Under alkaline conditions (pH = 9.46), the N form is the most stable structure of IRT. These results indicate the influence of the pH of the medium on the structural and dynamical properties of IRT in water solution. They may help to provide a better understanding on the relationship between the structure and biological activity of IRT.

Quality assurance in proton beam therapy using a plastic scintillator and a commercially available digital camera.

PubMed

Almurayshid, Mansour; Helo, Yusuf; Kacperek, Andrzej; Griffiths, Jennifer; Hebden, Jem; Gibson, Adam

2017-09-01

In this article, we evaluate a plastic scintillation detector system for quality assurance in proton therapy using a BC-408 plastic scintillator, a commercial camera, and a computer. The basic characteristics of the system were assessed in a series of proton irradiations. The reproducibility and response to changes of dose, dose-rate, and proton energy were determined. Photographs of the scintillation light distributions were acquired, and compared with Geant4 Monte Carlo simulations and with depth-dose curves measured with an ionization chamber. A quenching effect was observed at the Bragg peak of the 60 MeV proton beam where less light was produced than expected. We developed an approach using Birks equation to correct for this quenching. We simulated the linear energy transfer (LET) as a function of depth in Geant4 and found Birks constant by comparing the calculated LET and measured scintillation light distribution. We then used the derived value of Birks constant to correct the measured scintillation light distribution for quenching using Geant4. The corrected light output from the scintillator increased linearly with dose. The system is stable and offers short-term reproducibility to within 0.80%. No dose rate dependency was observed in this work. This approach offers an effective way to correct for quenching, and could provide a method for rapid, convenient, routine quality assurance for clinical proton beams. Furthermore, the system has the advantage of providing 2D visualization of individual radiation fields, with potential application for quality assurance of complex, time-varying fields. © 2017 The Authors. Journal of Applied Clinical Medical Physics published by Wiley Periodicals, Inc. on behalf of American Association of Physicists in Medicine.

Nearly constant ratio between the proton inertial scale and the spectrum break length scale in the plasma beta range from 0.2 to 1.4 in the solar wind turbulence

NASA Astrophysics Data System (ADS)

Wang, X.; Tu, C. Y.; He, J.; Wang, L.

2017-12-01

The spectrum break at the ion scale of the solar wind magnetic fluctuations are considered to give important clue on the turbulence dissipation mechanism. Among several possible mechanisms, the most notable ones are the two mechanisms that related respectively with proton thermal gyro-radius and proton inertial length. However, no definite conclusion has been given for which one is more reasonable because the two parameters have similar values in the normal plasma beta range. Here we do a statistical study for the first time to see if the two mechanism predictions have different dependence on the solar wind velocity and on the plasma beta in the normal plasma beta range in the solar wind at 1 AU. From magnetic measurements by Wind, Ulysses and Messenger, we select 60 data sets with duration longer than 8 hours. We found that the ratio between the proton inertial scale and the spectrum break scale do not change considerably with both varying the solar wind speed from 300km/s to 800km/s and varying the plasma beta from 0.2 to 1.4. The average value of the ratio times 2pi is 0.46 ± 0.08. However, the ratio between the proton gyro-radius and the break scale changes clearly. This new result shows that the proton inertial scale could be a single factor that determines the break length scale and hence gives a strong evidence to support the dissipation mechanism related to it in the normal plasma beta range. The value of the constant ratio may relate with the dissipation mechanism, but it needs further theoretical study to give detailed explanation.

A GAS-PHASE FORMATION ROUTE TO INTERSTELLAR TRANS-METHYL FORMATE

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

Cole, Callie A.; Wehres, Nadine; Yang Zhibo

2012-07-20

The abundance of methyl formate in the interstellar medium has previously been underpredicted by chemical models. Additionally, grain surface chemistry cannot account for the relative abundance of the cis- and trans-conformers of methyl formate, and the trans-conformer is not even formed at detectable abundance on these surfaces. This highlights the importance of studying formation pathways to methyl formate in the gas phase. The rate constant and branching fractions are reported for the gas-phase reaction between protonated methanol and formic acid to form protonated trans-methyl formate and water as well as adduct ion: Rate constants were experimentally determined using a flowingmore » afterglow-selected ion flow tube apparatus at 300 K and a pressure of 530 mTorr helium. The results indicate a moderate overall rate constant of (3.19 {+-} 0.39) Multiplication-Sign 10{sup -10} cm{sup 3} s{sup -1} ({+-} 1{sigma}) and an average branching fraction of 0.05 {+-} 0.04 for protonated trans-methyl formate and 0.95 {+-} 0.04 for the adduct ion. These experimental results are reinforced by ab initio calculations at the MP2(full)/aug-cc-pVTZ level of theory to examine the reaction coordinate and complement previous density functional theory calculations. This study underscores the need for continued observational studies of trans-methyl formate and for the exploration of other gas-phase formation routes to complex organic molecules.« less

Solubility and modeling acid-base properties of adrenaline in NaCl aqueous solutions at different ionic strengths and temperatures.

PubMed

Bretti, Clemente; Cigala, Rosalia Maria; Crea, Francesco; De Stefano, Concetta; Vianelli, Giuseppina

2015-10-12

Solubility and acid-base properties of adrenaline were studied in NaCl aqueous solutions at different ionic strengths (0