Delta Cp Research Articles

The Relationship between Physiological Characteristics and Durability in Male Professional Cyclists.

This study aimed to determine if durability can be predicted from laboratory measures in a professional cycling population. Data were collected from 10 professional cyclists (age = 19.2 ± 0.8 yr, body mass = 70.4 ± 5.5 kg, height = 182.9 ± 4.0 cm, body mass index = 21.0 ± 1.3 kg·m -2 , V̇O 2max = 74.4 ± 4.8 mL·kg -1 ·min -1 , critical power [CP] = 5.6 ± 0.6 W·kg -1 , W' = 23.7 ± 5.4 kJ). Participants completed a laboratory test and a CP test on two occasions. The second occasion was preceded by a novel fatiguing protocol, which consisted of five bouts of 8-min of exercise at 105%-110% of CP. CP in a fatigued state was expressed as a percentage of the fresh CP and coined delta CP (∆CP). The Pearson product correlation analysis was conducted to determine the relationship between laboratory-based measures and ∆CP. Significant positive relationships were found between ∆CP and relative peak power output ( r = 0.891, P < 0.001), relative maximum oxygen uptake ( r = 0.835, P = 0.003), relative power output at the second ventilatory threshold ( r = 0.738, P = 0.015), power output at the first ventilatory threshold ( r = 0.748, P = 0.013) and relative power output at the first ventilatory threshold ( r = 0.826, P = 0.003), gross efficiency at 300 W ( r = 0.869, P = 0.001), and at 200 W ( r = 0.792, P = 0.006). Significant negative relationships were found between ∆CP and carbohydrate oxidation at 200 W ( r = -0.702, P = 0.024). A multiple linear regression demonstrated that ∆CP can be predicted from laboratory measures ( R2 = 0.96-0.98, P < 0.001). These findings demonstrate the physiological determinants of durability in a professional cycling population.

Steady and dynamic shear rheology of Indian Jujube (Ziziphus mauritiana Lam.) fruit pulp with physiochemical, textural and thermal properties of the fruit

Indian jujube (Ziziphus mauritiana Lam.) is a globally underutilized fruit and is well-known for being a good source of total phenolics and a rich source of ascorbic acid with essential minerals such as calcium, phosphorus and iron. A rheometer was used to investigate the rheological characterization of Indian jujube pulp. It was found that Indian jujube pulp exhibit a shear thinning behaviour as viscosity (η) and shear stress (σ) of pulp significantly (p < 0.05) decreased as the shear rate (γ̇̇) increased. Pulp's behaviour was characterized efficiently by correlating Power-law, Carreau-Yasuda, and Cross models with flow curve. The Cross model was found more efficient (R2:0.99775) than the Power-law (R2:0.85995) and Carreau-Yasuda model (R2:0.97291). Dynamic shear rheology test of the Indian jujube pulp presented weak gel behaviour as storage modulus (G') surpassed loss modulus (G") value. The frequency dependency of both the storage G′ and loss G′′ moduli was evaluated using a power Law model with a determination coefficient (R2) greater than 0.9. The tan(δ) values (0.020 to 0.535) suggested that the Indian jujube pulp is elastic in nature. Textural properties of Indian jujube fruit like hardness, percentage deformation at the hardness and peak stress of Indian jujube fruit were 32.76 ± 5.00 N, 7.15 ± 1.06% and 17780.80 ± 3185.47 N/m2. DSC curve showed glass transition phase of unprocessed Indian jujube fruit flesh at (98.29°C), and specific heat (Delta Cp) was found 193.96 J/g. The findings of this study will aid in the development of handling and processing machinery for the commercial manufacture of Indian jujube pulp.

Heat capacity, thermal conductivity and magnetocaloric effect in Heusler alloy Ni-=SUB=-47-=/SUB=-Mn-=SUB=-40-=/SUB=-Sn-=SUB=-13-=/SUB=-

The results of a study of heat capacity, thermal conductivity and magnetocaloric effect of the polycrystalline Ni47Mn40Sn13 alloy depending on temperature (T=80-350 K) and magnetic field (0-8 T) are given. A considerable difference between the values of a sudden change in heat capacity Delta CP in the heating and cooling mode was found near the magnetostructural martensite--austenite phase transition (MSPT), which is related to influence of latent heat of phase transition. Thermal conductivity in the range of T=80-300 K rises with temperature (dkappa/dT&amp;gt;0) and increases in more than three times. Electronic thermal conductivity in the martensitic phase (T=150 K) is 37% of the total value. An anomalous rise of thermal conductivity Deltakappa=kappa(aust)-kappa(mart)=4.2 Wm K was found in the region of MSPT. The contributions of electrons and phonons to the observed sudden change are 63 and 37% respectively and are conditioned both by a rise of mobility of conduction electrons under a martensite--austenite transition and by an increase of phonons' free path length. The magnetocaloric effect in cyclic magnetic fields with the amplitude of 1.8 T was studied. A dependence of reverse effect value on temperature scanning rate was established. The direct measurements of Delta T in the cyclic magnetic field of 1.2 T show a twofold decrease of the effect amplitude near TC at an increase of cyclic magnetic field frequency from 1 to 30 Hz. Most likely, this is related to magnetic and microstructural heterogeneities which act as an additional thermal dissipation channel. Keywords: Heusler alloys, heat capacity, thermal conductivity, magnetocaloric effect, cyclic magnetic fields.

Thermal properties of products based on ABS / PC

Polymer blends are capable of providing materials with extended useful properties beyond the range that can be obtained from single polymers. Acrylonitrile-butadiene-styrene and polycarbonate blends are important industrial materials, especially, in the machine building industry, as ABS improves the processing properties of parts and their toughness, and they also feature positive thermodynamic interaction between PC and styrene-acrylonitrile (SAN) copolymer as a part of ABS. DSC method was used to study the thermal properties of ABS and ABS/PC blends with various ratios. The result from this work shows that abnormal endothermic effect in the ABS glass transition region implies an inadequate manufacturing process for automotive components. The paper studied the influence of ABS/PC blend ratios on the value of glass transition temperature and delta Cp, and it was found that the Tg value for SAN/PC increased with decreasing ABS for ABS/PC blends with mass ratio of 75/25, 60/40, 50/50.

Simulations of the Long Baseline Neutrino Experiment for the Sieroszowice Underground Laboratory (SUNLAB)

The Sieroszowice Underground Laboratory in Poland, SUNLAB, had been studied in the years 2008-2011 within the framework of the FP7 LAGUNA design study as an option for the realization of a next generation large volume neutrino observatory in Europe. However, in order to fully understand its physics capabilities, the feasibility studies of the SUNLAB laboratory have continued after 2011, including sensitivity calculations focused on the delta CP measurement for a large LArTPC detector at a distance of 950 km from CERN in a long baseline neutrino experiment. For this purpose the neutrino beam based on the SPS proton accelerator at CERN was simulated and the LAr data used to simulate the detector response.

Log (TG)/HDL-C ratio as a predictor of decreased islet beta cell function in patients with type 2 diabetes: 6-year cohort study.

The aim of the present study was to explore whether the triglyceride to high density lipoprotein cholesterol ratio [log (TG)/HDL-C] and peripheral blood leukocytes DNA telomere length could predict future islet beta cell function decreased in Chinese type 2 diabetes mellitus (T2DM) during a 6-year cohort. Sixty T2DM patients (without insulin treatment at baseline) were included in the 6-year cohort study. Peripheral blood leukocytes DNA telomere length, HbA1c, blood lipid profile, fatty fat acid, glucose, insulin and C peptide (3 h after a mixed meal) were determined. Delta C peptide area under curve (Delta CP AUC) was used to reflect change in beta cell secretion function (Delta CP AUC = baseline CP AUC - CP AUC after 6 years). Subjects were divided into slow decrease of beta cell function group (Delta CP AUCslow group) and fast decrease group (Delta CP AUCfast group) according to median of Delta CP AUC. Baseline demographic characteristics, clinical variables between two groups were compared. Correlations between baseline data and Delta CP AUC were analyzed. Baseline log (TG)/HDL-C was positively correlated with Delta CP AUC (r = 0.306, P = 0.027); log (TG)/HDL-C in Delta CP AUCfast group was higher than that in Delta CP AUCslow group (0.103 ± 0.033 vs 0.083 ± 0.030, P = 0.027). There was no significant difference in DNA telomere length between the two groups. Change in DNA telomere length over 6 years was not significantly correlated with baseline blood lipid. In Chinese T2DM patients, high baseline log (TG)/HDL-C ratio predicts fast progression of islet beta cell dysfunction. It may be a simple index to predict progression speed of islet beta cell dysfunction.

Novel Use of Pectin as a Microneedle Base

Hydrocolloid pectin formulation was utilized as a novel base for fabricating biodegradable micro-needle (MN) arrays. The pectin MNs were, on average, found to be 897.71 ± 3.48 µm in height and 234.31 ± 2.27 µm in base width, with an inter base spacing of 498.66 ± 1.60 µm, and corresponding to an aspect ratio of 3.83 ± 0.04. Bovine serum albumin (BSA) and pectin gel interaction was found to be dependent on the loaded protein amount. By contrast, regardless of the amount of BSA incorporated, pectin MNs competed with BSA to form a complex with Cu(2+) ions in a bicinchoninic acid (BCA) kit. The glass transition of the pectin MN base was found to be 145.15 ± 12.09, with a delta Cp of 2.60 ± 0.02 J g(-1) K(-1). Because pectin MNs are skin friendly and naturally occurring, with biodegradable and hydrocolloidal features, they are promising vehicle for the controlled release of macromolecules.

Hydration and Heat Stability Effects on Protein Unfolding

In summary, the thermal denaturation of proteins has been elucidated in terms of the chain free energy and the hydration free energy as follows. (1) Method to calculate the unfolding free energy. The free energy of unfolding consists of two contributions: the hydration around the molecule, and the intramolecular interactions. A method to calculate the free energy of hydration from the accessible surface area (ASA) of the constituent atomic groups in a protein has been developed. This assumes a proportionality between the free energy and the ASA, where the proportional constants were determined by least-squares fitting to the experimentally derived thermodynamic data on small molecules. Similarly, the free energy of unfolding for the chain in vacuo can be also calculated from the ASA, using the unfolding thermodynamics derived from the experimental data of the ten proteins. (2) Thermodynamics of protein unfolding predicted from the three-dimensional structures and from the amino acid content in proteins. First, our method is applied to predict the thermodynamics of protein unfolding from the X-ray structure. The predicted values of four test proteins agree well with the experimentally derived values. It also accounts for the temperature dependence of the free energy and of the enthalpy upon unfolding for 14 proteins. Second, this method is applied to the helix-coil transition of short peptides of poly(L-Ala)20 and Ac-(AAAAK)3A-NH2. The calculated enthalpy change is close to the experimental values for poly-L-Lys and poly-L-Glu. Since delta Hcu at 25 degrees C significantly contributes to delta Gu, the helix formation is enthalpy-driven through interactions in the chain. Third, the method is applied to predict the unfolding thermodynamics of a globular protein from its amino acid content. It also accounts for the temperature dependence of the free energy of unfolding for the 14 proteins. The agreement between the experimental and the calculated values by this method for the 14 proteins is not so different from those obtained with the three-dimensional structures. Fourth, the values of delta Cpu for 14 proteins may be closely approximated to the predicted values of delta Cp,hu. The delta Cp,hu value in a protein consists of the major contribution from the hydrophobic and the aromatic residues, and the minor one from the hydrophilic residues. (3) Dominant free energies in protein folding.(ABSTRACT TRUNCATED AT 400 WORDS)

Rosiglitazone preserves islet β-cell function of adult-onset latent autoimmune diabetes in 3 years follow-up study

The newly developed insulin sensitizer-thiazolidinediones have the potential to downregulate inflammation and autoimmune response. The objective of this study was to observe the beneficial effects on β-cell function in the LADA patients treated with rosiglitazone. 54 LADA patients were assigned to oral hypoglycemic agents group (GAD-Ab < 175 U/mL and FCP > 0.3 nmol/L) or early insulin administration group (GAD-Ab ≥ 175 U/mL or GAD-Ab < 175 U/mL and FCP ≤ 0.3 nmol/L). Then, those patients were randomly assigned to receive sulfonylureas (SUs group) or rosiglitazone (RSG group) therapy, or to receive insulin alone (INS group) or rosiglitazone plus insulin (INS + RSG group). Plasma glucose, HbA1c, fasting C-peptide (FCP) and C-peptide after 2 h 75-g glucose load (PCP) were determined every 6 months. The levels of PCP and delta CP were higher in RSG group compared with those in SUs group after the 18th month. The PCP level (after the 12th month) and delta CP level (after the 18th month) in INS ± RSG group were higher than those in INS group. Rosiglitazone combined with insulin wherever or not preserved β-cell function in LADA patients after 3 years.

Effect of the central disulfide bond on the unfolding behavior of elongation factor Ts homodimer from Thermus thermophilus.

Functionally active elongation factor Ts (EF-Ts) from Thermus thermophilus forms a homodimer. The dimerization interface of EF-Ts is composed of two antiparallel beta-sheets that can be connected by an intermolecular disulfide bond. The stability of EF-Ts from T. thermophilus in the presence and absence of the intermolecular disulfide bond was studied by differential scanning calorimetry and circular dichroism. The ratio of the van't Hoff and calorimetric enthalpies, delta H(vH)/delta H(cal), indicates that EF-Ts undergoes thermal unfolding as a dimer independently of the presence or absence of the disulfide bond. This can be concluded from (1) the presence of residual secondary structure above the thermal transition temperature, (2) the absence of concentration dependence, which would be expected for dissociation of the dimer prior to unfolding of the monomers, and (3) a relatively low heat capacity change (delta Cp) upon unfolding. The retained dimeric structure of the thermally denatured state allowed for the determination of the effect of the intermolecular disulfide bond on the conformational stability of EF-Ts, which is deltadelta G(S-S,SH HS) = 10.5 kJ/mol per monomer at 72.5 degrees C. The possible physiological implications of the dimeric EF-Ts structure and of the intersubunit disulfide bond for the extreme conformational stability of proteins in thermophiles are discussed.

C-peptide and glucagon profiles in minority children with type 2 diabetes mellitus.

The present study was conducted to determine the extent of insulin deficiency and glucagon excess in the hyperglycemia of type 2 diabetes in children. The incidence of type 2 diabetes mellitus in children and adolescents has increased substantially over the past several years. Because insulin and glucagon action both regulate blood glucose concentration, we studied their responses to mixed meals in children with type 2 diabetes. Subjects were 24 patients with type 2 diabetes compared with 24 controls, aged 9--20 yr (predominantly African-Americans), matched for body mass index and sexual maturation. All of those with diabetes were negative for antibodies to glutamic acid decarboxylase. Plasma glucose, glucagon, and serum C-peptide concentrations were measured at 0, 30, 60, 90, and 120 min after a mixed liquid meal (Sustacal) ingestion (7 mL/kg body weight; maximum, 360 mL). The area under the curve (AUC) was calculated by trapezoidal estimation. The incremental C-peptide (Delta CP) in response to the mixed meal was calculated (peak -- fasting C-peptide). The plasma glucose AUC was significantly greater in patients than in controls (mean +/- SEM, 1231 +/- 138 vs. 591 +/- 13 mmol/L x min; P < 0.001). The Delta CP was significantly lower in those with diabetes than in controls (1168 +/- 162 vs. 1814 +/- 222 pmol/L; P < 0.02). Glucagon responses did not differ between the two groups. Hyperglycemia is known to inhibit glucagon secretion. Therefore, our patients with substantial hyperglycemia would be expected to have decreased glucagon responses compared with controls and are thus relatively hyperglucagonemic. Patients were divided into poorly and well controlled subgroups (glycosylated hemoglobin A(1c), > or =7.2% and <7.2%, respectively). There were no significant differences in the Delta CP and glucagon responses between these two subgroups. We next analyzed the data in terms of duration of diabetes (long term, > or =1 yr; short term, <1 yr). The CP was significantly lower in long- vs. short-term patients (768 +/- 232 vs. 1407 +/- 199 pmol/L; P < 0.05). The plasma glucagon AUC was significantly higher in the long- vs. short-term patients (9029 +/- 976 vs. 6074 +/- 291 ng/L x min; P < 0.001). Hemoglobin A(1c) did not differ between long- vs. short-term patients. Our results indicate that relative hypoinsulinemia and hyperglucagonemia represent the pancreatic beta- and alpha-cell dysfunctions in children with type 2 diabetes. The severity of both beta- and alpha-cell dysfunctions appears to be determined by the duration of diabetes.

7 Isothermal Titration Calorimetry in Drug Discovery

Isothermal titration calorimetry (ITC) follows the heat change when a test compound binds to a target protein. It allows precise measurement of affinity. The method is direct, making interpretation facile, because there is no requirement for competing molecules. Titration in the presence of other ligands rapidly provides information on the mechanism of action of the test compound, identifying the intermolecular complexes that are relevant for structure-based design. Calorimetry allows measurement of stoichiometry and so evaluation of the proportion of the sample that is functional. ITC can characterize protein fragments and catalytically inactive mutant enzymes. It is the only technique which directly measures the enthalpy of binding (delta H degree). Interpretation of delta H degree and its temperature dependence (delta Cp) is usually qualitative, not quantitative. This is because of complicated contributions from linked equilibria and a single change in structure giving modification of several physicochemical properties. Measured delta H degree values allow characterization of proton movement linked to the association of protein and ligand, giving information on the ionization of groups involved in binding. Biochemical systems characteristically exhibit enthalpy-entropy compensation where increased bonding is offset by an entropic penalty, reducing the magnitude of change in affinity. This also causes a lack of correlation between the free energy of binding (delta G degree) and delta H degree. When characterizing structure-activity relationships (SAR), most groups involved in binding can be detected as contributing to delta H degree, but not to affinity. Large enthalpy changes may reflect a modified binding mode, or protein conformation changes. Thus, delta H degree values may highlight a potential discontinuity in SAR, so that experimental structural data are likely to be particularly valuable in molecular design.

Thermodynamic molecular switch in sequence‐specific hydrophobic interactions

AbstractThis communication will demonstrate the existence of a thermodynamic molecular switch in the pairwise, sequence‐specific hydrophobic interaction of Ile–Ile, Leu–Ile, Val–Leu, or Ala–Leu over the temperature range of 273–333 K reported by Nemethy and Scheraga in 1962. Based on Chun's development of the Planck–Benzinger methodology, the change in inherent chemical bond energy at 0 K, ΔH°(T0), is 3.0 kcal mol−1 for Ile–Ile, 2.4 for Leu–Ile, 1.8 for Val–Leu, and 1.2 kcal mol−1 for Ala–Leu. The value of ΔH°(T0) decreases as the length of the hydrophobic side chain decreases. It is clear that the strength and stability of the hydrophobic interaction is determined by the packing density of the side chains, with Ala–Leu being the most stable. At 〈Tm〉, the thermal agitation energy, \documentclass{article}\pagestyle{empty}\begin{document}$\int^{T}_{0}\Delta Cp^{\circ}(T)\,dT$\end{document}, is about five times greater than ΔH°(T0) in each case. Additionally, the thermal agitation energy for the same series, evaluated at 〈Tm〉, decreases in the same order, that is, as the length of the side chain decreases. This pairwise, sequence‐specific hydrophobic interaction is highly similar in its thermodynamic behavior to that of other biological systems, except that the negative Gibbs free energy change minimum at 〈Ts〉 occurs at a considerably higher temperature, 355 K compared to about 300 K. The melting temperature, 〈Tm〉, is also high, 470 K compared to 343 K in a biological system. The implication is that the negative Gibbs free energy minimum at a well‐defined 〈Ts〉 has it origin in the hydrophobic interactions, which are highly dependent on details of molecular structure. In addition to the four specific dipeptide interactions described, we have shown in our unpublished work the existence of a thermodynamic molecular switch in the interactions of 32 dipeptides wherein a change of sign in ΔCp°(T)reaction leads to a true negative minimum in the Gibbs free energy of reaction, and hence, a maximum in the related Keq. Indeed, all interacting biological systems that we have thus far examined using the Planck–Benzinger approach point to the universality of thermodynamic molecular switches. © 2001 John Wiley &amp; Sons, Inc. Int J Quantum Chem, 2001

Thermodynamic analyses of triplex formation with homopurine oligonucleotide.

We analyzed the thermodynamics of purine motif triplex formation by isothermal titration calorimetry. The signs of calorimetric enthalpy change, delta Hcal, and entropy change, delta S, of the triplex formation were negative in the temperature range between 15 and 35 degrees C. Since an observed negative delta S was unfavorable for the triplex formation, the triplex formation was driven by a large negative delta Hcal. delta Hcal decreased with increasing temperature, yielding a negative heat capacity change, delta Cp, of approximately -1.2 kcal mol-1 K-1. We found that the binding constant, Ka, increased with increasing temperature, leading to an apparent positive van't Hoff enthalpy change, delta Hvh, which was in sharp contrast with the large negative delta Hcal. The analyses of the observed temperature dependence of Ka and delta Hcal and the negative delta Cp suggest that the purine motif triplex formation near room temperature is not a simple two-state binding process but exhibits multiple states, which was previously observed for the pyrimidine motif triplex formation near room temperature.

Microcalorimetric determination of thermodynamic parameters for ionophore-siderophore host-guest complex formation.

Thermodynamic parameters (delta H, delta S, and delta G) were determined by microcalorimetry in wet chloroform for host-guest assembly formation involving second-sphere complexation of the siderophore ferrioxamine B by crown ether (18-crown-6, cis-dicyclohexano-18-crown-6, benzo-18-crown-6) and cryptand (2.2.2 cryptand) hosts. Similar data were also collected for the same hosts with the pentylammonium ion guest, which corresponds to the pendant pentylamine side chain of ferroxamine B. Host-guest assembly formation constants (Ka) obtained from microcalorimetry agree with values obtained indirectly from chloroform/water extraction studies in those cases where comparable data are available. On the basis of a trend established by the pentylammonium guest, an enhanced stability relative to the crown ethers is observed for the assembly composed of ferrioxamine B and 2.2.2 cryptand that is due to entropic effects. Trends in delta H and delta S with changes in host and guest structure are discussed and attributed directly to host-guest complex formation, as solvation effects were determined to be insignificant (delta Cp = 0).

Probing the "two-pronged plug two-holed socket" model for the mechanism of binding of the Src SH2 domain to phosphotyrosyl peptides: a thermodynamic study.

Src homology 2 (SH2) domains are protein modules that specifically bind to tyrosyl phosphorylated peptides on signaling proteins. X-ray crystallographic studies of the SH2 domain of the Src kinase have probed the mechanism of binding, leading to the "two-pronged plug two-holed socket" mechanism whereby binding is hypothesized to resemble a two-pronged plug (the peptide) inserting into a two-holed socket (the SH2 domain). This binding model predicts (1) a hydrophobic basis for high-affinity binding largely determined by the level of insertion of the third residue C-terminal to the phosphotyrosine in the peptide into a primarily hydrophobic pocket (the +3 binding pocket) of the SH2 domain, and (2) a binding mechanism involving no significant conformational changes in the SH2 domain. In this study, we have probed these predictions by using isothermal titration calorimetry to extract complete thermodynamic profiles (Delta G degrees, Delta H degrees, Delta S degrees, Delta Cp degrees) for the binding of the Src SH2 domain to two series of tyrosyl phosphopeptides. One series consisted of peptides that have been determined by X-ray crystallography to have different levels of insertion of the peptide's +3 position into the +3 binding pocket. The other series consisted of peptides with progressively smaller hydrophobic side chains (I, L, V, and A) at the +3 position. Consistent with a binding mechanism that does not involve substantial conformational changes, the Delta Cp degrees values for all peptides were small and, at least for the high-affinity interactions, similar to the Delta Cp degrees values predicted from surface area calculations. However, unexpectedly, this study reveals that high-affinity binding was only partially determined by the interactions between the +3 residue in the peptide and the +3 binding pocket. Furthermore, the Delta Cp degrees values for all peptides studied were similar, implying similar degrees of desolvation of the +3 binding pocket upon binding. These results indicate that the "two-pronged plug two-holed socket" model is an oversimplification of the Src SH2 domain binding mechanism.

Thermodynamics of the interaction of human immunoglobulin E with its high-affinity receptor Fc epsilon RI.

We have employed isothermal titration calorimetry (ITC) and circular dichroism (CD) spectroscopy to characterize the binding of soluble fragments of IgE (IgE-Fc and Fc epsilon 3-4) to a soluble fragment of the high-affinity receptor Fc epsilon RI alpha-chain (sFc epsilon RI alpha). The thermodynamic parameters for the interaction of IgE-Fc and Fc epsilon 3-4 with sFc epsilon RI alpha, determined using ITC, confirm the earlier conclusion that the C epsilon 2 domain is not involved in the interaction and that the stoichiometry of both complexes is 1:1. For both IgE-Fc and Fc epsilon 3-4, the value of Delta H degrees is -36.9 +/- 4.6 kcal mol-1 at 37.3 degreesC and Delta Cp degrees is -820 +/- 120 cal mol-1 K-1. The temperature at which DeltaS degrees is zero is 284 +/- 1 K, indicating that the entropy contribution to the thermodynamics of association is unfavorable at physiological temperature. Of particular interest is the large value of Delta Cp degrees. The large surface area of IgE and Fc epsilon RI alpha that is implicated in complex formation from previous mutagenesis studies on the two proteins may account in part for the magnitude of Delta Cp degrees. Additional contributions may arise from hydration within the binding site and changes in tertiary structure of the individual components of the complex. However, the CD spectra of IgE, IgE-Fc, and Fc epsilon 3-4 complexes with sFc epsilon RI alpha are merely the sum of the spectra of their individual components, indicating that the secondary structure of the immunoglobulin domain folds are preserved on complex formation. Thus, any change in tertiary structure must be limited to the relative disposition of the immunoglobulin domains C epsilon 3 and C epsilon 4 in IgE and the two immunoglobulin-like domains in the alpha-chain of Fc epsilon RI.

Energetics of two-step binding of a chromophoric reaction product, trans-3-indoleacryloyl-CoA, to medium-chain acyl-coenzyme-A dehydrogenase.

We previously demonstrated that the UV/visible spectrum of a chromophoric ligand, trans-3-indoleacryloyl-coenzyme-A (IACoA), is red-shifted (due to polarization of its carbonyl group) upon binding to pig kidney medium-chain acyl-CoA dehydrogenase (MCAD). The transient kinetic data revealed that the overall binding occurred in two steps. The first (fast) step involved the formation of an MCAD-IACoA collision complex in which the electronic structure of IACoA remained unchanged (the "colorless" complex), followed by a slow isomerization step with a concomitant red-shift in the IACoA spectrum (the "colored" complex) [Johnson, J. K., Wang, Z. X., and Srivastava, D. K. (1992) Biochemistry 31, 10564-10575]. To ascertain the energetics of the above two-step process, we investigated the temperature dependence of the spectral changes, binding constant of the MCAD-IACoA complex, and the rate constants for the conversion between the colorless and colored complexes. The data revealed that as the temperature of the incubation mixture of MCAD and IACoA ([IACoA] >> [MCAD] > Kd) increases from 12 to 35 degrees C, the resultant spectral peak of the MCAD-IACoA complex (lambda max = 417 nm) decreases. However, in this temperature range, the equilibrium constant for the second (isomerization) step remains unaffected. Isothermal titration calorimetric studies for the binding of IACoA to MCAD reveal that the overall binding energy at 25 degrees C (delta G degree = -7.4 kcal/mol) is contributed almost equally by the enthalpic (delta H degree = -3.7 kcal/mol) and entropic (T delta S degree = 3.7 kcal/mol) changes. As the temperature increases, both delta H degree and T delta S degree decrease proportionately, resulting in a strong enthalpy-entropy compensation effect. The temperature dependence of delta H degree yields a delta Cp degree value of -0.24 kcal/mol. The data presented herein throw light on the energetic consequences for the binding of IACoA to MCAD, the apparent similarity between the van't Hoff and calorimetric enthalpies, enthalpic and entropic contributions during the polarization of the carbonyl group of IACoA, and the overall structural-functional features of the enzyme-ligand complex as well as enzyme catalysis.

Conformational stability of muscle acylphosphatase: the role of temperature, denaturant concentration, and pH.

The conformational stability (delta G) of muscle acylphosphatase, a small alpha/beta globular protein, has been determined as a function of temperature, urea concentration, and pH. A combination of thermally induced and urea-induced unfolding, monitored by far-UV circular dichroism, was used to define the conformational stability over a wide range of temperature. Through analysis of all these data, the heat capacity change upon unfolding (delta Cp) could be estimated, allowing the determination of the temperature dependence of the main thermodynamic functions (delta G, delta H, delta S). Thermal unfolding in the presence of urea made it possible to extend such thermodynamic analysis to examine these parameters as a function of urea concentration. The results indicate that acylphosphatase is a relatively unstable protein with a delta G(H2O) of 22 +/- 1 kJ mol-1 at pH 7 and 25 degrees C. The midpoints of both thermal and chemical denaturation are also relatively low. Urea denaturation curves over the pH range 2-12 have allowed the pH dependence of delta G to be determined and indicate that the maximum stability of the protein occurs near pH 5.5. While the dependence of delta G on urea (the m value) does not vary with temperature, a significant increase has been found at low pH values, suggesting that the overall dimensions of the unfolded state are significantly affected by the number of charges within the polypeptide chain. The comparison of these data with those from other small proteins indicates that the pattern of conformational stability is defined by individual sequences and not by the overall structural fold.

14] Prediction of binding energetics from structure using empirical parameterization

We have presented an empirical method that can be used to predict the binding energetics for protein-protein or protein-peptide interactions from three-dimensional structures. The approach differs from other empirical methods in yielding a thermodynamic description of the binding process, including delta Cp, delta H degree, and delta S degree, rather than predicting delta G degree alone. These thermodynamic terms can provide a wealth of detail about the nature of the interaction, and, if sufficient experimental data are available for comparison, a greater assessment of the accuracy of the calculations. A recurring theme throughout this article is the need for more complete thermodynamic and structural characterizations of protein-ligand interactions. This includes not only characterization of the binding delta H degree, delta S degree, and delta Cp, but a thorough investigation into equilibria linked to binding, such as protonation, ion binding, and conformational changes. Sufficient data will allow parameterization on binding data rather than protein unfolding data. Further inclusion of information obtained from unfolding studies is not likely to generate significant improvement in the accuracy of the calculations. As additional binding data become available, the parameterization can be further extended to include relationships derived from analyses of these data. Not only will this increase accuracy and thus confidence, but allow extension of the method of additional types of interactions.