Binding of the Bovine and Porcine Pancreatic Secretory Trypsin Inhibitor (Kazal) To Human Leukocyte Elastase: A Thermodynamic Study

Abstract

The effect of pH and temperature on the apparent association equilibrium constant (K_a) for the binding of the bovine and porcine pancreatic secretory trypsin inhibitor (Kazal-type inhibitor, PSTI) to human leukocyte elastase has been investigated. At pH8.0, values of the apparent thermodynamic parameters for human leukocyte elastase: Kazal-type inhibitor complex formation are: bovine PSTT – K_a = 6.3 × 10⁴M^?1, δ5G° = -26.9kJ/mol, δH° = +11.7kJ/mol, and δS° = +1.3 × 10² entropy units; porcine PSTI –K_a = 7.0 × 10³M^?1,δG° = -21.5kJ/mol, δH° = +13.0kJ/mol, and δS° = +1.2 × 10² entropy units (values of K_a δG° and δS° were obtained at 21.0°C; values of δH° were temperature independent over the range (between 5.0°C and 45.0°C) explored). On increasing the pH from 4.5 to 9.5, values of K_a for bovine and porcine PSTI binding to human leukocyte elastase increase thus reflecting the acidic pK-shift of the His57 catalytic residue from ?7.0, in the free enzyme, to ?5.1, in the serine proteinase: inhibitor complexes. Thermodynamics of bovine and porcine PSTI binding to human leukocyte elastase has been analyzed in parallel with that of related serine (pro)enzyme/Kazal-type inhibitor systems. Considering the known molecular models, the observed binding behaviour of bovine and porcine PSTI to human leukocyte elastase was related to the inferred stereochemistry of the serine proteinase/inhibitor contact region(s).     

Binding of the Recombinant Proteinase Inhibitor Eglin c,of the Soybean Bowman-Birk Proteinase Inhibitor and of its Chymotrypsin and Trypsin Inhibiting Fragments to Leu-Proteinase,the Leucine Specific Serine Proteinase from Spinach (Spinacia oleracea L.) Leaves: Thermodynamic Study

Abstract

The effect of pH and temperature on the apparent association equilibrium constant (K_a) for the binding of the recombinant proteinase inhibitor eglin c (eglin c), of the soybean Bowman-Birk proteinase inhibitor (BBI) and of its chymotrypsin and trypsin inhibiting fragments (F-C and F-T, respetively) to Leuproteinase, the leucine specific serine proteinase from spinach (Spinacia oleracea L.) leaves, has been investigated. On lowering the pH from 9.5 to 4.5, values of K_a (at 21°C) for complex formation decrease thus reflecting the acidic pK-shift of the hystidyl catalytic residue from ～6.9, in the free Leu-proteinase, to ～5.1, in the enzyme: inhibitor adducts. At pH 8.0, values of the apparent thermodynamic parameters for the proteinase:inhibitor complex formation are: Leu-proteinase:eglin c - K_a = 2.2 × 10¹¹ M^-1, δG°= - 64kJ/mol, δH° = + 5.9kJ/mol, and δS° = + 240J/molK; Leu-proteinase:BBI - K_a = 3.2 × 10¹⁰ M^-1, δG° = - 59kJ/mol, δH°= + 8.8kJ/mol, and δS° = + 230J/molK; and Leu-proteinase:F-C - K_a = 1.1 × 10⁶ M^-1, δG°= - 34kJ/mol, δH° = + 18J/mol, and δS° = + 180J/molK (values of K_a, δG° and δS° were obtained at 21.0°C; values of δH° were temperature-independent over the range explored, i.e. between 10.0°C and 40.0°C). F-T does not inhibit Leu-proteinase up to an inhibitor concentration of 1.0 × 10^-3 M, suggesting that the upper limit of K_a is 1 × 10² M^-1. Considering the known molecular models, the observed binding behaviour of eglin c, BBI, F-C and F-T to Leu-proteinase has been related to the inferred stereochemistry of the enzyme/inhibitor contact region     

Properties of the Mg2+-induced low-affinity nucleotide binding site of (Na++ K+)-activated ATPase

(1) The Mg²⁺-induced low-affinity nucleotide binding by (Na⁺ + K⁺)-ATPase has been further investigated. Both heat treatment (50–65°C) and treatment with N-ethylmaleimide reduce the binding capacity irreversibly without altering the K_d value. The rate constant of inactivation is about one-third of that for the high-affinity site and for the (Na⁺ + K⁺)-ATPase activity. (2) Thermodynamic parameters (ΔH° and ΔS°) for the apparent affinity in the ATPase reaction (K_m ATP) and for the true affinity in the binding of AdoPP[NH]P (K_d and K_i) differ greatly in sign and magnitude, indicating that one or more reaction steps following binding significantly contribute to the K_m value, which thus is smaller than the K_d value. (3) Ouabain does not affect the capacity of low-affinity nucleotide binding, but only increases the K_d value to an extent depending on the nucleotide used. GTP and CTP appear to be most sensitive, ATP and ADP intermediately sensitive and AdoPP[NH]P and least sensitive to ouabain. Ouabain reduces the high-affinity nucleotide binding capacity without affecting the K_d value. (4) The nucleotide specificity of low-affinity binding site is the same for binding (competition with AdoPP[NH]P) and for the ATPase activity (competition with ATP): AdoPP[NH]P > ATP > ADP > AMP. (5) The low-affinity nucleotide binding capacity is preserved in the ouabain-stabilized phosphorylated state, and the K_d value is not increased more than by ouabain alone. (6) It is inferred that the low-affinity site is Iocated on the enzyme, more specifically its α-subunit, and not on the surrounding phospholipids. It is situated outside the phosphorylation centre. The possible functional role of the low-affinity binding is discussed.     

The First Example of a Hoogsteen Basepaired DNA Duplex in Dynamic Equilibrium with a Watson-Crick Basepaired Duplex—A Structural (NMR), Kinetic and Thermodynamic Study

Abstract

A single-point substitution of the O4′ oxygen by a CH₂ group at the sugar residue of A ⁶ (i.e. 2′-deoxyaristeromycin moiety) in a self-complementary DNA duplex, 5′- d(C¹G²C³G⁴A⁵A⁶T⁷T⁸C⁹G¹⁰C¹¹G¹²)₂ ^?3, has been shown to steer the fully Watson-Crick basepaired DNA duplex (1A), akin to the native counterpart, to a doubly A ⁶:T⁷ Hoogsteen basepaired (1B) B-type DNA duplex, resulting in a dynamic equilibrium of (1A)→←(1B): K_eq = k₁/k_-1 = 0.56±0.08. The dynamic conversion of the fully Watson-Crick basepaired (1A) to the partly Hoogsteen basepaired (1B) structure is marginally kinetically and thermodynamically disfavoured [k₁ (298K) = 3.9± 0.8 sec^?1; δH°? = 164±14 kJ/mol;-TδS°? (298K) = ?92 kJ/mol giving a δG₂₉₈°? of 72 kJ/mol. Ea (k1) = 167±14 kJ/mol] compared to the reverse conversion of the Hoogsteen (1B) to the Watson-Crick (1A) structure [k-1 (298K) = 7.0±0.6 sec-1, δH°? = 153±13 kJ/mol;-TδS°? (298K) = ?82 kJ/mol giving a δG₂₉₈°? of 71 kJ/mol. Ea (k-1) = 155±13 kJ/mol]. A comparison of δG₂₉₈°? of the forward (k1) and backward (k-1) conversions, (1A)→←(1B), shows that there is ca 1 kJ/mol preference for the Watson-Crick (1A) over the double Hoogsteen basepaired (1B) DNA duplex, thus giving an equilibrium ratio of almost 2:1 in favour of the fully Watson-Crick basepaired duplex. The chemical environments of the two interconverting DNA duplexes are very different as evident from their widely separated sets of chemical shifts connected by temperature-dependent exchange peaks in the NOESY and ROESY spectra. The fully Watson-Crick basepaired structure (1A) is based on a total of 127 intra, 97 inter and 17 cross-strand distance constraints per strand, whereas the double A ⁶:T⁷ Hoogsteen basepaired (1B) structure is based on 114 intra, 92 inter and 15 cross-strand distance constraints, giving an average of 22 and 20 NOE distance constraints per residue and strand, respectively. In addition, 55 NMR-derived backbone dihedral constraints per strand were used for both structures. The main effect of the Hoogsteen basepairs in (1B) on the overall structure is a narrowing of the minor groove and a corresponding widening of the major groove. The Hoogsteen basepairing at the central A ⁶:T⁷ basepairs in (1B) has enforced a syn conformation on the glycosyl torsion of the 2′- deoxyaristeromycin moiety, A ⁶, as a result of substitution of the endocyclic 4′-oxygen in the natural sugar with a methylene group in A ⁶. A comparison of the Watson-Crick basepaired duplex (1A) to the Hoogsteen basepaired duplex (1B) shows that only a few changes, mainly in α, σ and γ torsions, in the sugar-phosphate backbone seem to be necessary to accommodate the Hoogsteen basepair.     

Asymptotical Tests on the Equivalence,Substantial Difference and Non‐inferiority Problems with Two Proportions

Let d = p₂ ? p₁ be the difference between two binomial proportions obtained from two independent trials. For parameter d, three pairs of hypothesis may be of interest: H₁: d ≤ δ vs. K₁: d > δ; H₂: d ? (δ₁, δ₂) vs. K₂: d ∈ (δ₁, δ₂); and H₃: d ∈ [δ₁, δ₂] vs. K₃: d ? [δ₁, δ₂], where H_i is the null hypothesis and K_i is the alternative hypothesis. These tests are useful in clinical trials, pharmacological and vaccine studies and in statistics generally. The three problems may be investigated by exact unconditional tests when the sample sizes are moderate. Otherwise, one should use approximate (or asymptotical) tests generally based on a Z‐statistics like those suggested in the paper. The article defines a new procedure for testing H₂ or H₃, demonstrates that this is more powerful than tests based on confidence intervals (the classic TOST – two one sided tests – test), defines two corrections for continuity which reduce the liberality of the three tests, and selects the one that behaves better. The programs for executing the unconditional exact and asymptotic tests described in the paper can be loaded at http://www.ugr.es/~bioest/software.htm. (© 2004 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Effect of Age on the Kinetics of the Binding of Iodocyanopindolol to a Membrane Preparation of Rat Lungs

Abstract

The association (k₊₁) and dissociation (k_-1) rate constants, and the equilibrium thermodynamic binding parameters (ΔG°, ΔH° and ΔS°) of the β-adrenergic ligand [¹²⁵Iodo]cyanopindolol (ICYP) were studied in a crude lung membrane preparation of rats of different ages. There was no difference in k₊₁-values for the different age groups, while the k_-1-values were in all cases difficult to measure: almost no dissociation of ICYP from its binding site occurs. The thermodynamic properties were not affected by age. It is concluded that, in these experimental conditions, age has no effect on the kinetic parameters of the binding of ICYP to the β-adrenoceptors in rat lung.     

Comparative thermodynamics of opioid receptor ligand interaction in the bovine adrenal medulla membranes—Evidence of opioid site heterogeneity

1. A marked dependence on temperature of agonist binding δ, μ and κ₁₋₃, opioid sites in the bovine adrenal medulla was observed, at the range of 0 to 37°C. These changes concern kinetic (k₁) and equilibrium constants (K_d), but not binding capacities (B_max).2. These dependences are different for each ligand and each opioid receptor, suggesting their molecular heterogeneity.3. The comparative thermodynamics indicates that the interaction of opioid agonists with their receptor is exergonic (ΔG° < 0) and entropy driven (ΔS° > 0).4. The comparison of Van't Hoff and Arrhenius plots indicates a discrete mechanism in the binding of each opioid receptor.     

Interaction Between Yeast Eukaryotic Initiation Factor eIF4E and mRNA 5′ Cap Analogues Differs from That for Murine eIF4E

Abstract

Measurements of interaction of 7-methyl-GTP eIF4E from S. cerevisiae were performed by means of two methods: Isothermal Titration Calorimetry (ITC) and fluorescence titration. The equilibrium association constants (K_as) derived from the two methods show significantly different affinity of yeast eIF4E for the mRNA 5′ cap than those of the murine and human proteins. The observed differences in the K_as values and the enthalpy changes of the association (ΔH°) suggest some dissimilarity in the mode of binding and stabilization of cap in the complexes with eIF4E from various sources.     

Thermodynamic Study of Iodocyanopindolol β-Adrenoceptors Interactions with Rat Lung and Cerebral Cortex

Abstract

(±)¹²⁵ I-cyanopindolol (±) I CYP) was used to characterize β-adrenoceptors on rat lung and cerebral cortex membranes. The affinity of (±) ICYP was higher for lung (K^d = 64.3 pM) at 37°C. The association reaction of (±) ICYP was faster with lung (k₊₁ = 1.52 × 10⁹ M^?1.min^?1) than with cerebral cortex β-adrenoceptors (k₊₁ = 1.75 × 10⁸ M^?1.min^?1). In both tissues, the dissociation reaction followed a biphasic process with a fast (t ½ = 15.4 min and 5.6 min for lung and cerebral cortex respectively) and a slow component (t ½ = 474 min and 255 min for lung and cerebral cortex respectively). The thermodynamic parameters for (±) ICYP - β-adrenoceptors binding have been determined from kinetics and equilibrium studies, for the two tissues, at several temperatures between 0° and 44° C. For lung and cerebral cortex, Arrhenius plots were linear with different energies of activation. Van't Hoff plot was not linear for lung and the standard enthalpy and entropy changes of (±) ICYP - β-adrenoceptors interaction decreased linearly with temperature : the binding occured with a negative heat capacity change (ΔCp° = -368.9 cal. moles^?1. K^?1) at 25° C. Thermodynamic and kinetic results show that binding of (±) ICYP to lung β-adrenoceptors could involve two successive equilibria with a conformational change of the β-adrenergic receptor.     

Investigations on the interactions of DiAmsar with serum albumins: Insights from spectroscopic and molecular docking techniques

Diamine‐sarcophagine (DiAmsar) binding to human serum albumin (HSA) and bovine serum albumin (BSA) was investigated under simulative physiological conditions. Fluorescence spectra in combination with Fourier transform infrared (FT‐IR), UV‐visible (UV–vis) spectroscopy, cyclic voltammetry (CV), and molecular docking method were used in the present work. Experimental results revealed that DiAmsar had an ability to quench the HSA and BSA intrinsic fluorescence through a static quenching mechanism. The Stern–Volmer quenching rate constant (K_sv) was calculated as 0.372 × 10³ M^‐1 and 0.640 × 10³ M^‐1 for HSA and BSA, respectively. Moreover, binding constants (K_a), number of binding sites (n) at different temperatures, binding distance (r), and thermodynamic parameters (?H°, ?S°, and ?G°) between DiAmsar and HSA (or BSA) were calculated. DiAmsar exhibited good binding propensity to HSA and BSA with relatively high binding constant values. The positive ?H° and ?S° values indicated that the hydrophobic interaction is main force in the binding of the DiAmsar to HSA (or BSA). Furthermore, molecular docking results revealed the possible binding site and the microenvironment around the bond. Copyright © 2014 John Wiley & Sons, Ltd.     

Proton magnetic resonance studies of the conformational changes of dideoxynucleoside ethyl phosphotriesters

PMR investigations on the diastereomeric phosphate methyl protons of the dinucleoside ethyl phosphotriesters Tp(C₂H₅)T, dA, and dIp(C₂H₅)dI have been used to study the conformational changes of these dimersin solution. In D₂O (273°K), the diastereomeric phosphate-methly groups of Tp(C₂H₅)T appear as a triplet. The methyl resonances of dIp(C₂H₅)dI and dAp(C₂H₅)dA appear as two sets of triplets and their chemical shift differences (δ₁ ? δ₂), decrease with increasing temperature, finally becoming zero at 292°K and 333°K, respectively. The same phenomenon is observed for dAp(C₂H₅)dA in CD₃OD; in this detacking solvent, the difference (δ₁ ? δ₂) diminishes to zero at a lower temperature (261°K). At room temperature in D₂O, the chemical shift of the phosphate methyl of Tp(C₂H₅)T appears at lower field than those of dIp(C₂H₅)dI or dAp(C₂H₅)dA. The differences between the chemical shifts of these groups (δ_I ? δ_T or δ_A ? δ_T) increase with increasing temperature, and reach maximal values at 301°K and 333°K, respectively. The results suggest that at low temperature the largest fraction of the dimer population exists in a stacked state, with the phosphate-ethyl groups outside the stack. Increasing temperature causes an oscillation of the bases and a shift in the dimer population away from the stacked state. Finally at high temperature, the planar bases rorate with respect to one another and in the case of dIp(C₂H₅)dI and dAp(C₂H₅)dA, the ethyl groups experience shielding by the anisotropic ring current of the five-membered ring of the bases. Thus, the current pmr studies and those reported earlier from our laboratory support an “oscillation-rotation model” for the unstacking process of the dimers. The relationship of this model and the “two-state model” is discussed.     

Estimation of intracellular pH in muscle of fishes from different thermal environments

A technique based on homogenisation of rapidly frozen tissue was used to investigate the regulation of intracellular pH (pH_i) in freshwater and marine fish from diverse environmental temperatures. The following species were held at ambient temperatures of ca. 1°C (Notothenia coriiceps; Antarctica), 5°C (Pleuronectes platessa, Myoxocephalus scorpius; North Sea), and 26°C (Oreochromis niloticus; African lakes). The effects of seasonal acclimatisation to 4, 11 and 18°C were also examined in rainbow trout in the winter, autumn and summer, respectively. Extracellular (whole blood) pH (pH_e) did not follow the constant relative alkalinity relationship, where pH⁺=pOH⁻ for any particular temperature, over a range of 1–26°C (overall δpH_e/δT=0.009±0.002 U °C⁻¹; P<0.001), apparently being regulated by ionic fluxes and ventilation. Intracellular pH (pH_i) was also regulated independently of pN(=0.5 pK water) in all species of fish examined. The inverse relationship between pH_i and environmental temperature gave an overall δpH_i/δT of −0.010±0.001 U °C⁻¹ (for both white and red muscle) and −0.004±0.003 U °C⁻¹ (cardiac muscle). However, between 1 and 11°C δpH_i/δT was much higher (P<0.001), −0.022±0.003 U °C⁻¹ (white muscle) and −0.022±0.004 U °C⁻¹ (red muscle). The possible adaptive roles for these different acid–base responses to environmental temperature variation among tissues and species, and the potential difficulties of estimating pH_i, are discussed.     

Discontinuities in the Temperature Function of Transmembrane Water Transport in Chara: Relation to Ion Transport

The NMR (nuclear magnetic resonance) method of Conlon and Outhred (1972) was used to measure diffusional water permeability of the nodal cells of the green alga Chara gymnophylla. Two local minima at 15 and 30°C of diffusional water permeability (P _d ) were observed delimiting a region of low activation energy (E _a around 20 kJ/mol) indicative of an optimal temperature region for membrane transport processes. Above and below this region water transport was of a different type with high E _a (about 70 kJ/mol). The triphasic temperature dependence of the water transport suggested a channel-mediated transport at 15–30°C and lipid matrix-mediated transport beyond this region. The K⁺ channel inhibitor, tetraethylammonium as well as the Cl⁻ channel inhibitor, ethacrynic acid, diminished P _d in the intermediate temperature region by 54 and 40%, respectively. The sulfhydryl agent p-(chloromercuri-benzensulfonate) the water transport inhibitor in erythrocytes also known to affect K⁺ transport in Chara, only increased P _d below 15°C. In high external potassium (`K-state') water transport minima were pronounced. The role of K⁺ channels as sensors of the optimal temperature limits was further emphasized by showing a similar triphasic temperature dependence of the conductance of a single K⁺ channel also known to cotransport water, which originated from cytoplasmic droplets (putatively tonoplast) of C. gymnophylla. The minimum of K⁺ single channel conductance at around 15°C, unlike the one at 30°C, was sensitive to changes of growth temperature underlining membrane lipid involvement. The additional role of intracellular (membrane?) water in the generation of discontinuities in the above thermal functions was suggested by an Arrhenius plot of the cellular water relaxation rate which showed breaks at 13 and 29°C. Received: 12 August 1998/Revised: 13 November 1998     

Identification and Characterization of Specific Binding Sites for Somatostatin in Cytosol of Bovine Cystic Duct Mucosa

Abstract

Specific binding sites for somatostatin have been detected in cytosolic fraction of bovine cystic duct mucosa. At 37°C, the interaction of ¹²⁵I-Tyr¹¹-somatostatin with cytosolic fraction was rapid, reversible, specific and saturable. At equilibrium, the binding of tracer was competitively inhibited by native peptide in the 1 nM to 2 µ M range of concentrations. Scatchard analysis of binding data suggested the presence of two distinct classes of somatostatin binding sites: a class with a high affinity (K_d = 7.8 ± 0.3 nM) and a low capacity (1.3 ± 0.3 pmol somatostatin/mg protein) and a class with a low affinity (K_d = 129.1 ± 2.0 nM) and a high capacity (43.5 ± 6.7 pmol somatostatin/mg protein). The binding sites were shown to be highly specific for somatostatin since neuropeptides present in cystic duct such as Leu-enkephalin, neurotensin, substance P and vasoactive intestinal peptide did practically not show competition. These findings suggest that somatostatin could contribute to the regulation of the functions of the cystic duct mucosa in physiological and pathological conditions.     

Application of isothermal titration calorimetry in bioinorganic chemistry

The thermodynamics of metals ions binding to proteins and other biological molecules can be measured with isothermal titration calorimetry (ITC), which quantifies the binding enthalpy (ΔH°) and generates a binding isotherm. A fit of the isotherm provides the binding constant (K), thereby allowing the free energy (ΔG°) and ultimately the entropy (ΔS°) of binding to be determined. The temperature dependence of ΔH° can then provide the change in heat capacity (ΔC _p°) upon binding. However, ITC measurements of metal binding can be compromised by undesired reactions (e.g., precipitation, hydrolysis, and redox), and generally involve competing equilibria with the buffer and protons, which contribute to the experimental values (K _ITC, ΔH _ITC). Guidelines and factors that need to be considered for ITC measurements involving metal ions are outlined. A general analysis of the experimental ITC values that accounts for the contributions of metal–buffer speciation and proton competition and provides condition-independent thermodynamic values (K, ΔH°) for metal binding is developed and validated.     

Reconstructing the δ18O of atmospheric water vapour via the CAM epiphyte Tillandsia usneoides: seasonal controls on δ18O in the field and large‐scale reconstruction of δ18Oa

Using both oxygen isotope ratios of leaf water (δ¹⁸O_L) and cellulose (δ¹⁸O_C) of Tillandsia usneoides in situ, this paper examined how short‐ and long‐term responses to environmental variation and model parameterization affected the reconstruction of the atmospheric water vapour (δ¹⁸O_a). During sample‐intensive field campaigns, predictions of δ¹⁸O_L matched observations well using a non‐steady‐state model, but the model required data‐rich parameterization. Predictions from the more easily parameterized maximum enrichment model (δ¹⁸O_L–M) matched observed δ¹⁸O_L and observed δ¹⁸O_a when leaf water turnover was less than 3.5 d. Using the δ¹⁸O_L–M model and weekly samples of δ¹⁸O_L across two growing seasons in Florida, USA, reconstructed δ¹⁸O_a was ?12.6 ± 0.3‰. This is compared with δ¹⁸O_a of ?12.4 ± 0.2‰ resolved from the growing‐season‐weighted δ¹⁸O_C. Both of these values were similar to δ¹⁸O_a in equilibrium with precipitation, ?12.9‰. δ¹⁸O_a was also reconstructed through a large‐scale transect with δ¹⁸O_L and the growing‐season‐integrated δ¹⁸O_C across the southeastern United States. There was considerable large‐scale variation, but there was regional, weather‐induced coherence in δ¹⁸O_a when using δ¹⁸O_L. The reconstruction of δ¹⁸O_a with δ¹⁸O_C generally supported the assumption of δ¹⁸O_a being in equilibrium with precipitation δ¹⁸O (δ¹⁸O_ppt), but the pool of δ¹⁸O_ppt with which δ¹⁸O_a was in equilibrium – growing season versus annual δ¹⁸O_ppt – changed with latitude.     

Determination on the binding of chlortetracycline to bovine serum albumin using spectroscopic methods

In this work, the interaction of chlortetracycline with bovine serum albumin (BSA) was investigated by fluorescence spectroscopy, circular dichroism (CD) spectroscopy, and molecular docking. Results indicated that chlortetracycline quenches BSA fluorescence mainly by a static quenching mechanism. The quenching constants (K_SV) were obtained as 5.64 × 10⁴, 4.49 × 10^4/, and 3.44 × 10^4/ M_?1 at 283, 295, and 307 K, respectively. The thermodynamic parameters of enthalpy change Δ H°, entropy change Δ S°, and free energy change Δ G° were ?5.12 × 10^4/ J mol?1, ?97.6 J mol?1 K?1, and ?2.24 × 10^4/ J mol?1 (295 K), respectively. The association constant (K_A) and the number of binding sites (n) were 9.41 × 10^3/ M?1 and 0.86, respectively. The analysis results suggested that the interaction was spontaneous, and van der Waals force and hydrogen‐bonding interactions played key roles in the reaction process. In addition, CD spectra proved secondary structure alteration of BSA in the presence of chlortetracycline. © 2012 Wiley Periodicals, Inc. J Biochem Mol Toxicol 26:331–336, 2012; View this article online at wileyonlinelibrary.com . DOI 10:1002/jbt.21424     

Kinetic and Thermodynamic Studies of Phenolic Compounds’ Adsorption on River Sediment

Phenolic compounds have increasingly attracted global concerns in recent years due to their strong bioaccumulation and potential toxicity. In this study, the adsorption of 2, 4-Dichlorophenol (2, 4-DCP), 2, 4-Dinitrophenol (2, 4-DNP) and 2, 4-Dimethyphenol (2, 4-DMP) on sediment at different temperatures was studied. Adsorption isotherms fitted well to the Freundlich model and the adsorption capacity was increased when the temperature increased from 5°C to 25°C. It was found that the adsorption process could be modeled well using pseudo-second-order kinetic equation. The thermodynamic parameter ΔG° for 2, 4-DCP, 2, 4-DNP and 2, 4-DMP varied between ?8.82 and ?13.68, ?4.9 and ?8.05, and ?7.52 and ?10.55 kJ/mol, respectively, with ΔH° (kJ/mol) and ΔS°(J/(mol·K)) measuring 55.397 and 0.2263, 40.121 and 0.1585, and 38.012 and 0.16, respectively. The calculated thermodynamic parameters suggested that adsorption of the three selected phenolic compounds was a spontaneous (ΔG°< 0), endothermic (ΔH°> 0), and entropy-driven reaction (ΔS° > 0). The thermodynamic data also suggested that the three selected phenolic compounds to sediment were closed to chemisorption because ΔH° was around 40.     

Determination of the Group Electronegativity of CF3 Group in 3′-O-CF3-Thymidine by 1-NMR

Abstract

The interplay of enthalpy of the gauche effect (ΔH°_GE) of the [X3′-C3′-C4′-O4′] fragment in various 3′-substituted (X) 2′,3′-dideoxythymidine derivatives 1–7 and the inherent anomeric effect drives the two-state North ? South equilibrium in the constituent sugar moiety. The group electronegativity of 3′-OCF₃ substituent in Marriott's, Inamoto's and Mullay's scales has been determined from simple calibration graphs correlating the group electronegativity of various 3′-substituents (X) in 2′,3′-dideoxythymidine derivatives 1–7 with the experimental strength (ΔH°_GE) of the [X3′-C3′-C4′-O4′] gauche effect. ΔH°_GE has been experimentally determined from pseudorotational analyses of temperature-dependent ³J_HH coupling constants, and can be used as an unambiguous tool for direct experimental estimation of the group electronegativity of a specific substituent covalently attached to 3′-carbon of 2′,3′-dideoxythymidine, which can be compared, in turn, with the theoretical estimation carried out according to Marriott's or Inamoto's procedure. Inconsistency found between theoretical values in Marriott's and Inamoto's scales, on the one hand, and between our experimental estimate and the theoretical value in Marriott's scale, on the other, have been solved by refining the electronegativity scale using our experimental data for 1–7.