Ionization Enthalpies Research Articles

The aminopeptidase from Aeromonas proteolytica can function as an esterase.

The aminopeptidase from Aeromonas proteolytica (AAP) can catalyze the hydrolysis of L-leucine ethyl ester ( L-Leu-OEt) with a rate of 96 +/- 5 s-1 and a Km of 700 microM. The observed turnover number for L-Leu-OEt hydrolysis by AAP is similar to that observed for peptide hydrolysis, which is 67 +/- 5 s-1. The k(cat) values for the hydrolysis of L-Leu-OEt and L-leucine- p-nitroanilide ( L- pNA) catalyzed by AAP were determined at different pH values under saturating substrate concentrations. Construction of an Arrhenius plot from the temperature dependence of AAP-catalyzed ester hydrolysis indicates that the rate-limiting step does not change as a function of temperature and is product formation. The activation energy ( Ea) for the activated ES ester complex is 13.7 kJ mol-1, while the enthalpy and entropy of activation at 25 degrees C calculated over the temperature range 298-338 K are 11.2 kJ mol-1 and -175 J K-1 mol-1, respectively. The free energy of activation at 25 degrees C was found to be 63.4 kJ mol-1. The enthalpy of ionization was also measured and was found to be very similar for both peptide and ester substrates, yielding values of 20 kJ mol-1 for L-Leu-OEt and 25 kJ mol-1 for L- pNA. For peptide and L-amino acid ester cleavage reactions catalyzed by AAP, and 6.07, respectively. Proton inventory data suggest that two protons are transferred in the rate-limiting step of ester hydrolysis while only one is transferred in peptide hydrolysis. The combination of these data with the available X-ray crystallographic, kinetic, spectroscopic, and thermodynamic data for AAP provides new insight into the catalytic mechanism of AAP.

Thermodynamics of Dissociation and Metal Complexation of 3-Nitro-1,5-Diphenylformazan

Thermodynamic parameters for dissociation of 3-nitro-1,5-diphenylformazan and its complexation by some divalent metal ions were determined in a 50%(v/v) dioxane–water mixture at constant ionic strength (0.1 M KCl) using an automatic potentiometric technique. The changes in the standard Gibbs energy ΔGo and enthalpy ΔHo accompanying the complexation were found to decrease with increasing metal ionic radius and to increase with the electronegativity, the ionization enthalpy, and the enthalpy of hydration. The order of −ΔGo and −ΔHo values were found to be Mn2+ Zn2+, in accordance with the Irving–Williams order. The complexes were stabilized by both enthalpy and entropy changes and the results suggest that the complexation is an enthalpy-driven process. The transition-series contraction energy Er(Mn–Zn) and the ligand field stabilization energy δ H were calculated from the enthalpy changes.

Identification of catalytically active groups of lipoxygenase from wheat germ

Catalytically active groups of lipoxygenase in wheat germs have been identified. Blocking of catalytically active groups by specific reagents, dependence of the enzyme activity on pH, enthalpy of ionization of pK1 and pK2, the enzyme photoinactivation allowed to make a conclusion that imidasole and hydroxyl groups take part in lipoxygenase catalytic cycle.

Identification of Catalytically Active Groups of Wheat Germ Lipoxygenase

Dependences of lipoxygenase activity on pH, ionization enthalpies of various chemical groups, photoinactivation of the enzyme, and effects of specific reagents (p-CMB, DPF, and PMSF) were studied to identify catalytically active groups of the enzyme. The data suggest that the catalytic site of lipoxygenase includes imidazole and hydroxyl groups of histidine and serine residues, respectively.

Re‐evaluation of the inductive effect in isolated molecules and in solution

The classical concept of inductive effect based originally on dissociation constants in water was revisited on isolated molecules in the gas phase. It then has a strict thermodynamic meaning of ΔHo(g) of an isodesmic reaction and can be calculated separately in the neutral molecules of acids (bases) and in anions (cations). Eight different reactions were investigated in which tradition assumes a purely inductive effect. Quantitative estimation was based on the known gas-phase enthalpies of ionization, ΔionHo(g), and gas-phase enthalpies of formation, ΔfHo(g); some lacking values of the latter were substituted by calculations at the MP2/6–31+G** or RHF/6–31+G** level. Substituent effects in neutral molecules are not negligible, particularly in smaller molecules, but are qualitatively different from the common scale of inductive effect and cannot at present be described in terms of any simple theory. The concept of inductive effect is to be reserved just for the energy difference between an ion and the respective neutral molecule. It can be modeled as the energy of interaction between the given substituent and a positive or negative charge but it is quantitatively predicted in terms of electrostatics (Kirkwood–Westheimer theory) only with some considerable disagreements. In water, this effect is strongly attenuated, differently in different classes of compounds; this attenuation can be in a qualitative accord with the electrostatic theory. Copyright © 2000 John Wiley & Sons, Ltd.

Modelling and experimental analysis of vacuum plasma spraying. Part II:prediction of temperatures and velocities of plasma gases and Ti particles ina plasma jet

A numerical model has been developed to calculate the spatial distributions ofplasma gas temperature, enthalpy, velocity and fractions of dissociated andionized species in a vacuum plasma spraying (VPS) plasma jet under a range of plasma current, Ar flowrate, H2 flow rate and chamber pressure, and the trajectories, temperaturesand velocities of Ti particles under typical processing conditions. The modeluses FLUENT V4.2 commercial software, incorporating approximations to describedissociation, ionization and recombination reactions in the plasma jet. Thecalculations show that the spatial distributions of plasma gas temperature,enthalpy, velocity, and degrees of dissociation and ionization in the plasmajet are mainly controlled by the initial boundary values at the plasma gunexit, which are functions of the VPS processing conditions. The model predictsthat the plasma jet length increases with increasing plasma current anddecreasing Ar flow rate and chamber pressure, and shows a maximum with varyingH2 flow rate, agreeing well with measurements. Particle trajectory is largelydetermined by the initial particle position at the plasma gun exit. Particletemperature and velocity increase rapidly in the first 100 mm of the plasmajet and then become nearly constant at axial distances >150 mm. Particletemperature and velocity in the plasma jet decrease with increasing particlesize and initial radial position at the plasma gun exit.

Sequence-specific DNA binding by the glucocorticoid receptor DNA-binding domain is linked to a salt-dependent histidine protonation.

We used isothermal titration calorimetry in the temperature range 21-25 degrees C to investigate the effect of pH on the calorimetric enthalpy (delta H(cal)) for sequence specific DNA-binding of the glucocorticoid receptor DNA-binding domain (GR DBD). Titrations were carried out in solutions containing 100 mM NaCl, 1 mM dithiothreitol, 5% glycerol by volume, and 20 mM Tris, Hepes, Mops, or sodium phosphate buffers at pH 7.5. A strong dependence of delta H(cal) on the buffer ionization enthalpy is observed, demonstrating that the DNA binding of the GR DBD is linked to proton uptake at these conditions. The apparent increase in the pK(a) for an amino acid side chain upon DNA binding is supported by the results of complementary titrations, where delta H(cal) shows a characteristic dependence on the solution pH. delta H(cal) is also a function of the NaCl concentration, with opposite dependencies in Tris and Hepes buffers, respectively, such that a similar delta H(cal) value is approached at 300 mM NaCl. This behavior shows that the DNA-binding induced protonation is inhibited by increased concentrations of NaCl. A comparison with structural data suggests that the protonation involves a histidine (His451) in the GR DBD, because in the complex this residue is located close to a DNA phosphate at an orientation that is consistent with a charged-charged hydrogen bond in the protonated state. NMR spectra show that His451 is not protonated in the unbound protein at pH 7.5. The pH dependence in delta H(cal) can be quantitatively described by a shift of the pK(a) of His451 from approximately 6 in the unbound state to close to 8 when bound to DNA at low salt concentration conditions. A simple model involving a binding competition between a proton and a Na(+) counterion to the GR DBD-DNA complex reproduces the qualitative features of the salt dependence.

Thermodynamic dissection of the binding energetics of KNI-272, a potent HIV-1 protease inhibitor.

KNI-272 is a powerful HIV-1 protease inhibitor with a reported inhibition constant in the picomolar range. In this paper, a complete experimental dissection of the thermodynamic forces that define the binding affinity of this inhibitor to the wild-type and drug-resistant mutant V82F/184V is presented. Unlike other protease inhibitors, KNI-272 binds to the protease with a favorable binding enthalpy. The origin of the favorable binding enthalpy has been traced to the coupling of the binding reaction to the burial of six water molecules. These bound water molecules, previously identified by NMR studies, optimize the atomic packing at the inhibitor/protein interface enhancing van der Waals and other favorable interactions. These interactions offset the unfavorable enthalpy usually associated with the binding of hydrophobic molecules. The association constant to the drug resistant mutant is 100-500 times weaker. The decrease in binding affinity corresponds to an increase in the Gibbs energy of binding of 3-3.5 kcal/mol, which originates from less favorable enthalpy (1.7 kcal/mol more positive) and entropy changes. Calorimetric binding experiments performed as a function of pH and utilizing buffers with different ionization enthalpies have permitted the dissection of proton linkage effects. According to these experiments, the binding of the inhibitor is linked to the protonation/deprotonation of two groups. In the uncomplexed form these groups have pKs of 6.0 and 4.8, and become 6.6 and 2.9 in the complex. These groups have been identified as one of the aspartates in the catalytic aspartyl dyad in the protease and the isoquinoline nitrogen in the inhibitor molecule. The binding affinity is maximal between pH 5 and pH 6. At those pH values the affinity is close to 6 x 10(10) M(-1) (Kd = 16 pM). Global analysis of the data yield a buffer- and pH-independent binding enthalpy of -6.3 kcal/mol. Under conditions in which the exchange of protons is zero, the Gibbs energy of binding is -14.7 kcal/mol from which a binding entropy of 28 cal/K mol is obtained. Thus, the binding of KNI-272 is both enthalpically and entropically favorable. The structure-based thermodynamic analysis indicates that the allophenylnorstatine nucleus of KNI-272 provides an important scaffold for the design of inhibitors that are less susceptible to resistant mutations.

Carbon-related defects in proton-irradiated, n-type epitaxial Si1−xGex

C i and CiCs defects, created by proton irradiation of n-type, strain-relaxed, epitaxial Si1−xGex of 0.005⩽x⩽0.5, have been studied using deep-level transient spectroscopy (DLTS). The ionization enthalpies of the two defects relative to the conduction band edge, ΔH, are found to increase linearly with increasing Ge content. It is shown that the corresponding levels are not pinned to any of the band edges. Furthermore, it is shown that, for both defects, the slopes, δΔH/δx, as well as the full width at half maximum (FWHM) of the corresponding DLTS peaks, are similar. These observations are in agreement with conclusions deduced from previous electron-paramagnetic resonance (EPR) measurements in pure silicon, stating that, for both defects, the trapped electron is preferentially located at the Ci atom because of its larger electronegativity as compared to those of silicon and germanium. The anneal temperature of the Ci defect, and correspondingly the in-growth temperature of the CiCs complex, increase with increasing Ge content. This is equivalent to an increasing retardation of the diffusion of Ci in Si1−xGex with increasing Ge content.

Complete basis set and gaussian ab initio computational study of lithium and sodium insertion reactions with hydrogen molecule

High level complete basis set and gaussian ab initio methods are applied in exploring various potential energy surfaces for hydrogen, lithium, and sodium reactions with hydrogen molecules. The potential energy surface was explored for neutral, as well as charged reactants. Computed ionization potentials, electron affinities, and enthalpies for electron transfers were compared to experimental data when available, and for other chemical systems our best evaluated values were suggested for further experimental exploration of these reaction surfaces. It was demonstrated that lithium and sodium actually have different reaction pathways when reacting with hydrogen. Lithium abstracts hydrogen while sodium inserts into the hydrogen molecule. In any case, the computed activation barriers are substantially higher than in the case of the hydrogen radical reaction with a hydrogen molecule. It was predicted that in their excited states, the reaction barriers are decreasing, and the reactions should be experimentally feasible, which is supported by many experimental evidences.

Energetics of Ca 2+–EDTA interactions: calorimetric study

The interaction between Ca 2+ and EDTA has been studied using isothermal titration calorimetry to elucidate the detailed mechanism of complex formation and to relate the apparent thermodynamic parameters of calcium binding to the intrinsic effects of ionization. It has been shown that Ca 2+ binding to EDTA is an exothermic process in the temperature range 5–48°C and is highly dependent on the buffer in which the reaction occurs. Calorimetric measurements along with pH-titration of EDTA under different solvent conditions shows that the apparent enthalpy effect of the binding is predominantly from the protonation of buffer. Subtraction of the ionization effect of buffer from the total enthalpy shows that the enthalpy of binding Ca 2+ to EDTA is −0.56 kcal mol −1 at pH 7.5. The Δ H value strongly depends on solvent conditions as a result of the degree of ionization of the two amino groups in the EDTA molecule, but depends little on temperature, indicating that the heat capacity increment for metal binding is close to zero. At physiological pH values where the amino groups of EDTA with p K a=6.16 and p K a=10.26 are differently ionized, the coordination of the Ca 2+ ion into the complex leads to release of one proton due to deprotonation of the amino group having p K a=10.26. Increasing the pH up to 11.2, where little or no ionization occurs, leads to elimination of the enthalpy component due to ionization, while its decrease to pH 2 leads to its increase, due to protonation of the two amino groups. The heat effect of Ca 2+/EDTA interactions, excluding the deprotonation enthalpy of the amino groups, i.e. that associated with the intrinsic enthalpy of binding, is higher in value (Δ b H o=−5.4 kcal mol −1) than the apparent enthalpy of binding. Thus, the large Δ G value for Ca 2+ binding to EDTA arises not only from favorable entropic but also enthalpic changes, depending on the ionization state of the amino groups involved in coordination of the calcium. This explains the great variability in Δ H obtained in previous studies. The ionization enthalpy is always unfavorable, and therefore dramatically decreases Ca 2+ affinity by reduction of the enthalpy term of the stability function. The origin of the enthalpy and entropy terms in the stability of the Ca 2+–EDTA complex is discussed.

Optimum temperature range for positive ion production from metal halide molecules incident upon heated metal catalysts

When the surface temperature ( T) of catalytic metal (Re, W, Mo, Ta or Nb) was gradually increased in a high vacuum (∼10 −5–10 −3 Pa), the positive ionization efficiency ( β +) of diatomic halide molecule YX (e.g., LiI, KF or TlCl) impinging with a constant flux (∼10 12–10 14 molecules cm −2 s −1) upon the surface steeply increased up to unity at the first boundary temperature ( T 1). Above T 1, β +=1 continued until the second boundary temperature ( T 2), above which β + decreased as T increased. Theoretical analysis of β + around the optimum temperature range ( T 1− T 2) yields the empirical formulae of T 1=( D 1+ I 1− φ 1 +)/ R 1 k and T 2=( φ 2 +− I 2)/ R 2 k. Here, D, I, and φ + are the dissociation enthalpy of YX, the ionization enthalpy of Y and the effective work function for the ionization on each metal surface employed, respectively, in eV at T 1 or T 2 in K,and k is Boltzmann's constant. The empirical constants ( R 1=28.4±1.3 and R 2=5.47±0.30) thus determined always hold for any system, irrespective of the difference in species of both sample and catalyst.

Ionization thermodynamics of methylbenzoic acids in EtOH–H 2O mixtures

Ionization enthalpies of o-, m- and p-methylbenzoic acids in ethanol–water mixtures have been determined. Combined with corresponding free energies, the ionization entropies have been calculated. The substituent and solvent effects on the ionization thermodynamics are discussed in terms of solute–solvent interactions and the structure of ethanol–water mixtures.

Formation and characterization of methoxy isothiocyanate (CH3ONCS) and methyl cyanate N-sulfide (CH3OCN+–S−) as radical cations and neutrals in the gas phase

Dissociative ionization of heterocyclic precursors has provided a convenient source of isomeric [CH3O,C,N,S]˙+ radical cations. Metastable ion (MI), collisional activation (CA), neutralization–reionization (NR) spectra, and ion–molecule reactions, performed in a hybrid tandem mass spectrometer of sectors–quadrupole–sectors configuration, have demonstrated the isothiocyanate, CH3ONCS˙+ a, and the nitrile N-sulfide, CH3OCN–S˙+ b, connectivities. For the sake of comparison, a potential precursor of the isocyanate, CH3SNCO˙+ c, was also investigated. The gas phase stability of the corresponding neutral molecules is indicated by the NR and NR/CA spectra. In addition, consecutive collisional activation processes (MS/MS/MS spectra) were used to characterize the structures of metastable ions which, in several cases, were found to be different from the corresponding ions generated in the ion source. Calculated ionization energies and enthalpies of reaction with NO˙ (at the G2(MP2,SVP) level) support the experimental characterization of ions a and b.

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

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

Thermodynamics of ionization of o-fluoro and o-chloro benzoic acids in water–ethanol mixtures at 298.15 K

The ionization enthalpies of Δ H° of o-fluoro and o-chloro benzoic acids were measured by LKB 2277 BioActivity Monitor at 298.15 K in water–ethanol mixtures from pure water up to 0.8 weight fraction ethanol. The ionization-free enthalpies Δ G° of the two acids in the same mixtures were obtained from published literature. The corresponding entropies Δ S° of ionization were calculated from the values of Δ H° and Δ G°.

Thermodynamics of ionization processes for fluoro-substituted benzoic acids in water–dimethylformamide mixtures at 298.15 K

The ionization enthalpies, Δ H 0, of benzoic acid, o- and p-fluorobenzoic acids were measured calorimetrically at 298.15 K in water–dimethylformamide (DMF) mixtures ranging from pure water up to 0.7 weight-fraction DMF. The ionization Gibbs energies, Δ G 0, of the acids in the same solutions were measured by the conductance method. The corresponding entropies, Δ S 0, of ionization were calculated. The effect of solvent and substituent on the thermodynamic properties of ionization were considered.

Can O=NOOH undergo homolysis?

Recent thermodynamic calculations of Merenyi and Lind [(1997) Chem. Res. Toxicol. 10, 1216-1220] suggest that O=NOOH can undergo homolysis to form the hydroxyl radical and nitrogen dioxide. This result is based in part on our statement that the enthalpy of ionization of O=NOOH is close to zero [Koppenol et al. (1992) Chem. Res. Toxicol. 5, 834-842]. As the ionization of O=NOOH is sensitive to the milieu and the rate of isomerization (to nitrate) to the total concentration of O=NOOH and O=NOO- [Kissner et al. (1997) Chem. Res. Toxicol. 10, 1285-1292], we reinvestigated the temperature dependence of the ionization constant and determined a deltaHo of 4+/-2 kcal mol(-1). This results in a standard Gibbs energy of homolysis of 16 kcal mol(-1) and a rate of homolysis of 1 x 10(-2) s[-1]. Given the uncertainty in the Gibbs energy of homolysis, upper and lower rates are 1 x 10(-4) and 0.6 s(-1), slower than the rate of isomerization, 1.2 s(-1) at 25 degrees C. The recombination of the homolysis products NO2. and HO. is known to lead to mainly peroxynitrous acid. If one assumes that a few percent of the recombinations lead to nitrate instead, then the rate of homolysis must be much higher than the rate of isomerization. We conclude therefore that homolysis is unlikely.

Temperature dependence of histidine ionization constants in myoglobin

The standard enthalpy of ionization of six titratable histidines in horse metaquomyoglobin was determined by repeating proton NMR titrations as a function of temperature and using the van't Hoff relationship. It was found that deltaH degrees varies between 16 and 37 kJ mol(-1) in the protein, compared with a value of 29 kJ mol(-1) in free histidine. The standard entropy change was evaluated by combining the enthalpy and free energy changes derived from the pKa values. Although the entropy change could not be precisely and accurately obtained by this method, it could be established that it spans a wide range, from -60 to 0 J K(-1) mol(-1), about the value of -23 J K(-1) mol(-1) for the free histidine. The entropy change was used within the framework of enthalpy-entropy compensation to partition the solvation component from the standard thermodynamic quantities for each of the titrating residues. It was shown that the partitioning of the values in the protein is not readily understood in terms of solvent accessibility or electrostatic interactions. The contribution of solvation effects to the temperature response appeared to be significant only in the case of His-119 and His-48. The standard quantities were also used to explore the energetics of proton binding in the native state at temperatures below the onset of thermal denaturation.

Isothermal titration microcalorimetric studies for the binding of octenoyl-CoA to medium chain acyl-CoA dehydrogenase.

We investigated the binding of octenoyl-CoA to pig kidney medium chain acyl-CoA dehydrogenase (MCAD) by isothermal titration microcalorimetry under a variety of experimental conditions. At 25 degrees C in 50 mM phosphate buffer at pH 7.6 (ionic strength of 175 mM), the binding is characterized by the stoichiometry (n) of 0.89 mole of octenoyl-CoA/(mole of MCAD subunit), delta G = -8.75 kcal/mol, delta H = -10.3 kcal/mol, and delta S = -5.3 cal mol(-1) K(-1), suggesting that formation of MCAD-octenoyl-CoA is enthalpically driven. By employing buffers with various ionization enthalpies, we discerned that formation of the MCAD-octenoyl-CoA complex, at pH 7.6, accompanies abstraction (consumption) of 0.52 +/- 0.15 proton/(MCAD subunit) from the buffer media. We studied the effects of pH, ionic strength, and temperature on the thermodynamics of MCAD-octenoyl-CoA interaction. Whereas the ionic strength does not significantly influence the above interaction, the pH of the buffer media exhibits a pronounced effect. The pH dependence of the association constant of MCAD +octenoyl-CoA <==> MCAD-octenoyl-CoA yields a pKa for the free enzyme of 6.2. Among thermodynamic parameters, whereas delta G remains invariant as a function of temperature, delta H and deltaS(standard) both decrease with an increase in temperature. At temperatures of < 25 degrees C, delta G is dominated by favorable entropic contributions. As the temperature increases, the entropic contributions progressively decrease, attain a value of zero at 23.8 degrees C, and then becomes unfavorable. During this transition, the enthalpic contributions become progressively favorable, resulting in an enthalpy-entropy compensation. The temperature dependence of delta H yields the heat capacity change (delta Cp(0)) of -0.37 +/- 0.05 kcal mol(-1) K(-1), attesting to the fact that the binding of octenoyl-CoA to MCAD is primarily dominated by the hydrophobic forces. The thermodynamic data presented herein are rationalized in light of structural-functional relationships in MCAD catalysis.