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PubMed1989

Protein complexation with DNA phosphates as a cause for DNA duplex destabilization: a thermodynamic model.

Complexation of positively charged sites in a protein with the negative DNA phosphate groups shields the phosphate charges. This diminishes interstrand electrostatic repulsions, which stabilizes the duplex. When phosphate shielding is present in one DNA strand only, the conformation of this strand changes due to a decrease of intrastrand phosphate-phosphate repulsions. This destabilizes the duplex since then the strands differ in conformation. A thermodynamic model is formulated to describe this stabilization/destabilization effect in terms of changed enthalpies and entropies of hybridization. It is found that protein complexation with one DNA strand can indeed lower the TM value of a duplex. The model is applied to the action of helicases (replication), RNA polymerases (transcription), and restriction endonucleases. Mechanisms with unilateral charge shielding are proposed for their duplex-destabilizing properties.

DNA, Single-Stranded↗

PubMed1990

Thermodynamics of polymolecular duplexes between phosphate-methylated DNA and natural DNA.

Phosphate-methylated (P.M.) DNA possesses a very high affinity for complementary natural DNA, as a result of the absence of interstrand electrostatic repulsions. In this study, a model system phosphate-methylated d[Cn] with natural d(Gk) (n less than k) is chosen for an investigation of the thermodynamic properties that determine duplex stability. The enthalpy change of a melting transition is shown to be considerably larger than is observed for corresponding natural DNA duplexes. It is found that delta Hn0 of GG/CC nearest neighbor pairwise interaction equals -15.6 kcal/mol, compared to -11.0 kcal/mol for the natural analog. The entropy change is strongly dependent on the length of the natural DNA strand and the number of phosphate-methylated DNA oligomers hybridized. The results are explained by means of a model in which a cooperative effect for subsequent hybridizations of phosphate-methylated DNA oligomers is assumed, thus giving additional stability.

DNA↗

PubMed1990

Thermodynamic stability of the 5' dangling-ended DNA hairpins formed from sequences 5'-(XY)2GGATAC(T)4GTATCC-3', where X, Y = A, T, G, C.

Expressions for the partition function Q (T) of DNA hairpins are presented. Calculations of Q (T), in conjunction with our previously reported numerically exact algorithm [T. M. Paner, M. Amaratunga, M. J. Doktycz, and A. S. Benight (1990) Biopolymers, 29, 1715-1734], yield a numerical method to evaluate the temperature dependence of the transition enthalpy, entropy, and free energy of a DNA hairpin directly from its optical melting curve. No prior assumptions that the short hairpins melt in a two-state manner are required. This method is then applied in a systematic manner to investigate the stability of the six basepair duplex stem 5'-GGATAC-3' having four-base dangling single-strand ends with the sequences (XY)2, where X, Y = A, T, G, C, on the 5' end and a T4 loop on the 3' end. Results show that all dangling ends of the sample set stabilize the hairpin against melting. Increases in transition temperatures as great as 4.0 degrees C above the blunt-ended control hairpin were observed. The hierarchy of the hairpin transition temperatures is dictated by the identity of the first base of the dangling end adjoining the duplex in the order: purine greater than T greater than C. Calculated melting curves of every hairpin were fit to experimental curves by adjustment of a single parameter in the numerically exact theoretical algorithm. Exact fits were obtained in all cases. Experimental melting curves were also calculated assuming a two-state melting process. Equally accurate fits of all dangling-ended hairpin melting curves were obtained with the two-state model calculation. This was not the case for the melting curve of the blunt-ended hairpin, indicating the presence of a four-base dangling-end drives hairpin melting to a two-state process. Q (T) was calculated as a function of temperature for each hairpin using the theoretical parameters that provided calculated curves in exact agreement with the experimentally obtained optical melting curves. From Q (T), the temperature dependence of the transition enthalpy delta H, entropy delta S, and free energy delta G were calculated for every hairpin providing a quantitative assessment of the effects of dangling ends on hairpin thermodynamics. Comparisons of our results are made with those of the Breslauer group [M. Senior, R. A. Jones, and K. J. Breslauer (1988) Biochemistry 27, 3879-3885] on the T2 5' dangling-ended d(GC)3 duplexes.(ABSTRACT TRUNCATED AT 400 WORDS)

Base Sequence↗

PubMed1991

Solvent reorganization contribution to the transfer thermodynamics of small nonpolar molecules.

The experimental thermodynamic data for the dissolution of five simple hydrocarbon molecules in water were combined with the solute-solvent interaction energy from a computer simulation study to yield data on the enthalpy change of solvent reorganization. Similar data were generated for dissolving these same solute molecules in their respective neat solvents using the equilibrium vapor pressure and the heat of vaporization data for the pure liquid. The enthalpy and the free energy changes upon cavity formation were also estimated using the temperature dependence of the solute-solvent interaction energy. Both the enthalpy and T delta S for cavity formation rapidly increase with temperature in both solvent types, and the free energy of cavity formation can be reproduced accurately by the scaled particle theory over the entire temperature range in all cases. These results indicate that the characteristic structure formation around an inert solute molecule in water produces compensating changes in enthalpy and entropy, and that the hydrophobicity arises mainly from the difference in the excluded volume effect.

Computer Simulation↗

PubMed1991

Importance of oligoelectrolyte end effects for the thermodynamics of conformational transitions of nucleic acid oligomers: a grand canonical Monte Carlo analysis.

Effects of salt concentration on the stabilities of oligonucleotide helices are analyzed directly in terms of delta gamma N----yN identical to gamma denyN - gamma natN, the difference in the salt-nucleotide phosphate preferential interaction coefficients for the denatured state, having yN phosphate charges, and for the native state, having N phosphate charges (y = 1 for hairpin denaturation and y = 0.5 for dimer denaturation). Previous experimental studies of the denaturation of hairpin oligonucleotides (having 18 less than N less than 44) indicate significant differences between delta gamma N----N and delta gamma infinity, the value determined for the denaturation of the corresponding polynucleotide. These differences are thermodynamic manifestations of the oligoelectrolyte end effect. In contrast, the available data on the denaturation of oligonucleotide dimer helices (N less than or equal to 22) imply that differences between delta gamma infinity and delta gamma N----0.5N, and hence oligoelectrolyte end effects, are small or negligible. To determine the origin of these apparently conflicting implications concerning the importance of oligoelectrolyte end effects, we have calculated the N dependence of gamma N from grand canonical Monte Carlo simulations for an idealized model of the structure and charge distribution of each oligomer conformation. Our calculations are in quantitative agreement with the experimental finding for d(TA) hairpin oligomers that -delta gamma N----N decreases linearly as N-1 increases, and with the extant experimental determinations of delta gamma N----0.5N. These results provide an illustration of how the large electrostatic end effects exhibited by the hairpin denaturation data are masked when delta gamma infinity is compared with values of delta gamma N----0.5N for short dimer helices (N less than or equal to 22). For 0.5N greater than 24, -delta gamma N----0.5N is predicted to be a linear function of N-1 whose slope has the opposite sign from, and is more salt-concentration dependent than, the corresponding slope of -delta gamma N----N as a function of N-1. Our calculations also yield predictions about the N dependences of the individual values of gamma N that can be tested by determining Donnan coefficients from membrane dialysis equilibrium experiments. For long enough hairpin and dimer oligonucleotides (yN greater than or equal to 24), in either native or denatured forms, we predict that the (positive) difference gamma infinity - gamma N increases linearly with increasing N-1. For smaller values of N the difference gamma infinity - gamma N continues to increase with increasing N-1.

Base Sequence↗

PubMed1992

Thermodynamic substantiation of water-bridged collagen structure.

A solution of the problem of topology of a hydrogen bond net in a triple helix of collagen is suggested on the basis of an analysis of thermodynamic data on denaturation of phylogenetically different collagen [T. V. Burjanadze (1982), Vol. 21, pp. 1489-1501; T. V. Burjanadze, E. I. Tiktopulo, and P. L. Privalov (1987), Dokl. Akad. Nauk. USSR, Vol. 293, pp. 720-724] as well as on the earlier evaluation of the energy of the OH group of the 4-hydroxyproline bond [A. R. Ward and P. Mason (1973), Journal of Molecular Biology, Vol. 29, pp. 431-435]. It is shown that only the water-bridged collagen structure [G. N. Ramachandran and R. Chandrasekharan (1968), Biopolymers, Vol. 6, pp. 1649-1661; G. N. Ramachandran, M. Bansal, and R. S. Bhatnagar (1973), Biochimica Biophysica Acta, Vol. 322, pp. 166-171; M. Bansal, C. Ramakrishnan, and G. N. Ramachandran (1975), Proceedings of the Indian Academy of Sciences, Vol. 82, pp. 152-164] can explain both the change of thermostability upon proline hydroxylation [J. Rosenbloom, M. Harsch, and S. Jimenez (1973), Archives of Biochemistry and Biophysics, Vol. 158, pp. 478-484] and its phylogenetic change [T. V. Burjanadze (1982)].

Animals↗

PubMed1994

Thermodynamics of cyclophilin catalyzed peptidyl-prolyl isomerization by NMR spectroscopy.

One-dimensional nmr exchange spectroscopy was carried out to determine thermodynamic parameters of cyclophilin-induced cis-trans isomerization of succinyl-Ala-Phe-Pro-Phe-p-nitroanilide. Rate measurements were possible at physiological temperatures. The kc/Km of rat cyclophilin was found to be 12.8 (+/- 0.5) s-1 microM-1 at 37 degrees C, intermediate to previously reported values that used a coupled enzyme assay extrapolated to this temperature. Activation energies (delta G not equal to) for the uncatalyzed and catalyzed reaction at 37 degrees C were found to be 19.7 and 17.1 kcal/mol, respectively, and were primarily due to an enthalpic barrier.

Amino Acid Isomerases↗

PubMed1994

The thermodynamics of solvent exchange.

A model for solvation in mixed solvents, which was developed for the free energy and preferential interaction [J. A. Schellman (1987), Biopolymers, Vol. 26, pp. 549-559; (1990), Biophysical Chemistry, Vol. 37, pp. 121-140; (1993), Biophysical Chemistry, Vol. 45, pp. 273-279], is extended in this paper to cover the thermal properties: enthalpy, entropy, and heat capacity. An important result is that the enthalpy of solvation H(ex)2 responds directly to the fraction of site occupation. This differs from the free energy G(ex)2 and preferential interaction gamma 32, which are measures of the excess binding above a random distribution of solvent molecules. In other words, the enthalpy is governed by K while G(ex)2 and gamma 32 are governed by (K-1) where K is the equilibrium constant on a mole fraction scale [Schellman (1987)]. The solvation heat capacity Cpex2 consists of two term: (1) the intrinsic heat capacity of species in solution with no change in composition, and (2) a term that accounts for the change in composition that accompanies solvent exchange. Binding to biological macromolecules is heterogeneous but experimentalists must use binding isotherms that assume the homogeneity of sites. Equations are developed for the interpretation of the experimental parameters (number of sites nexp, equilibrium constant Kexp, and enthalpy, delta hexp), when homogeneous formulas are applied to the heterogeneous case. It is shown that the experimental parameters for the occupation and enthalpy are simple functions of the moments of the distribution of equilibrium constants over the sites. In general, nexp is greater than the true number of sites and Kexp is greater than the average of the equilibrium constants. The free energy and preferential interaction can be fit to a homogeneous formula, but the parameters of the curve are not easily represented in terms of the moments of distributions over the sites. The strengths and deficiencies of this type of thermodynamic model are discussed.

Chemical Phenomena↗

PubMed2003

Thermodynamic parameters monitoring the equilibrium shift of enzyme-catalyzed hydrolysis/synthesis reactions in favor of synthesis in mixtures of water and organic solvent.

The main strategy developed to shift the equilibrium state of a hydrolase-catalyzed hydrolysis/synthesis reaction consists in reducing water activity by addition of organic solvents in the reaction medium. We have used several mixtures of water and 1,4-butanediol, ranging from pure water to pure 1,4-butanediol, to study the hydrolysis/synthesis reaction of the N-Cbz-L-tryptophanyl-glycineamide dipeptide, catalyzed by alpha-chymotrypsin. In the presence of 1,4-butanediol, alpha-chymotrypsin also catalyzed the esterification reaction between this diol and N-Cbz-L-tryptophan; this ester hydrolysis/synthesis reaction has thus also been examined. The dipeptide and ester equilibrium concentrations increase when the water content of the reaction medium is decreased. Using our experimental data, we have determined the equilibrium constants of the hydrolysis/synthesis equilibria involving the nonionized forms of the protected amino acids, the estimated values of which are Ksp = 8 10(5) for the dipeptide and Kse = 78 for the ester respectively. They are true thermodynamic equilibrium constants, each related to a single, well-defined reaction equilibrium and with water activity being taken into account. If an organic solvent is added to the reaction medium these equilibria can be shifted towards synthesis by decreasing the water activity but also by modifying the ionization/neutralization equilibrium constant of the ionizable groups. These two effects depend both on the water content and on the nature of the organic solvent used, and, in particular, on its dielectric constant. Because of the importance of this parameter in our study, we discuss using it as an indicator to select an appropriate organic solvent to perform an enzyme-catalyzed synthesis.

Bioreactors↗

PubMed2007

Influence of pH, temperature, and urea molar flowrate on Arthrospira platensis fed-batch cultivation: a kinetic and thermodynamic approach.

Arthrospira platensis was cultivated photoautotrophically at 6.0 klux light intensity in 5.0-L open tanks, using a mineral medium containing urea as nitrogen source. Fed-batch experiments were performed at constant flowrate. A central composite factorial design combined to response surface methodology (RSM) was utilized to determine the relationship between the selected response variables (cell concentration after 10 days, X(m), cell productivity, P(X), and nitrogen-to-cell conversion factor, Y(X/N)) and codified values of the independent variables (pH, temperature, T, and urea flowrate, K). By applying the quadratic regression analysis, the equations describing the behaviors of these responses as simultaneous functions of the selected independent variables were determined, and the conditions for X(m) and P(X) optimization were estimated (pH 9.5, T = 29 degrees C, and K = 0.551 mM/day). The experimental data obtained under these conditions (X(m) = 749 mg/L; P(X) = 69.9 mg/L.day) were very close to the estimated ones (X(m) = 721 mg/L; P(X) = 67.1 mg/L.day). Additional cultivations were carried out under the above best conditions of pH control and urea flowrate at variable temperature. Consistently with the results of RSM, the best growth temperature was 29 degrees C. The maximum specific growth rates at different temperatures were used to estimate the thermodynamic parameters of growth (DeltaH* = 59.3 kJ/mol; DeltaS* = -0.147 kJ/mol.K; DeltaG* = 103 kJ/mol) and its thermal inactivation (DeltaH(D) (o) = 72.0 kJ/mol; DeltaS(D) (o) = 0.144 kJ/mol.K; DeltaG(D) (o) = 29.1 kJ/mol).

Biomass↗

PubMed2007

Thermodynamic electron equivalents model for bacterial yield prediction: modifications and comparative evaluations.

Modifications are made to an earlier thermodynamic model (TEEM1) for prediction of maximum microbial yields from aerobic and anaerobic as well as heterotrophic and autotrophic growth. The revised model (TEEM2) corrects for lower yields found with aerobic oxidations of organic compounds where an oxygenase is involved and with growth on single-carbon (C1) compounds. TEEM1 and TEEM2 are based on energy release and consumption as determined from the reduction potential or Gibbs free energy of (1/2)-reaction reduction equations together with losses of energy during energy transfer. Energy transfer efficiency is a key parameter needed to make predictions with TEEM2, and was determined through evaluations with extensive data sets on aerobic heterotrophic yield available in the literature. For compounds following normal catabolic pathways, the best-fit value for energy transfer efficiency was 0.37, which permitted accurate predictions of growth with a precision of 15%-20% as determined by standard deviation. Using the same energy transfer efficiency, a similar precision, but somewhat less accuracy was found for organic compounds where oxidation involves an oxygenase (estimates 8% too high) and for C1 compounds (estimates 17% too high). In spite of the somewhat lower accuracy, the TEEM2 modifications resulted in improved predictions over TEEM1 and the comparison models.

Bacteria↗

PubMed2003

Protein stabilisation by compatible solutes: effect of mannosylglycerate on unfolding thermodynamics and activity of ribonuclease A.

Differential scanning calorimetry, optical spectroscopy, and activity measurements were used to investigate the effect of mannosylglycerate, a negatively charged osmolyte widely distributed among thermophilic and hyperthermophilic archaea and bacteria, on the thermal unfolding of ribonuclease A (RNase A). For comparison, assays in the presence of trehalose, a canonical solute in mesophiles, and potassium chloride were also carried out. A thermodynamic analysis was performed by using differential scanning calorimetry data. The changes in the heat capacity for unfolding were similar for the different solutes examined. Mannosylglycerate was an efficient thermostabiliser of RNase A and induced an increase of 6 degrees C mole(-1) in the melting temperature. Moreover, the performance of mannosylglycerate as a stabiliser depended on the net charge of the molecule, with the maximal effect being observed at pH values above 4.5. Analysis of the enthalpic and entropic contributions to unfolding, derived from calorimetric data, revealed that the stabilisation rendered by mannosylglycerate is primarily achieved through a decrease in the unfolding entropy. Also, the number of protons taken up by RNase A upon denaturation in the presence of mannosylglycerate was considerably higher than with other solutes, a result consistent with a more rigid structure of the native protein. Mannosylglycerate (potassium salt) inhibited the activity of RNase A, albeit to a smaller extent than KCl, and acted as an efficient suppressor of aggregation of the denatured protein, thereby having a remarkable beneficial effect on the inactivation of RNase A upon thermal denaturation. The results are discussed in view of the physiological role of this charged compatible solute.

Animals↗

PubMed2003

GdIII complexes with fast water exchange and high thermodynamic stability: potential building blocks for high-relaxivity MRI contrast agents.

On the basis of structural considerations in the inner sphere of nine-coordinate, monohydrated Gd(III) poly(aminocarboxylate) complexes, we succeeded in accelerating the water exchange by inducing steric compression around the water binding site. We modified the common DTPA(5-) ligand (DTPA=(diethylenetriamine-N,N,N',N",N"-pentaacetic acid) by replacing one (EPTPA(5-)) or two (DPTPA(5-)) ethylene bridges of the backbone by propylene bridges, or one coordinating acetate by a propionate arm (DTTA-prop(5-)). The ligand EPTPA(5-) was additionally functionalized with a nitrobenzyl linker group (EPTPA-bz-NO(2) (5-)) to allow for coupling of the chelate to macromolecules. The water exchange rate, determined from a combined variable-temperature (17)O NMR and EPR study, is two orders of magnitude higher on [Gd(eptpa-bz-NO(2))(H(2)O)](2-) and [Gd(eptpa)(H(2)O)](2-) than on [Gd(dtpa)(H(2)O)](2-) (k(ex)298=150x10(6), 330x10(6), and 3.3x10(6) s(-1), respectively). This is optimal for attaining maximum proton relaxivities for Gd(III)-based, macrocyclic MRI contrast agents. The activation volume of the water exchange, measured by variable-pressure (17)O NMR spectroscopy, evidences a dissociative interchange mechanism for [Gd(eptpa)(H(2)O)](2-) (DeltaV(not equal sign)=(+6.6+/-1.0) cm(3) mol(-1)). In contrast to [Gd(eptpa)(H(2)O)](2-), an interchange mechanism is proved for the macrocyclic [Gd(trita)(H(2)O)](-) (DeltaV (not equal sign)=(-1.5+/-1.0) cm(3) mol(-1)), which has one more CH(2) group in the macrocycle than the commercial MRI contrast agent [Gd(dota)(H(2)O)](-), and for which the elongation of the amine backbone also resulted in a remarkably fast water exchange. When one acetate of DTPA(5-) is substituted by a propionate, the water exchange rate on the Gd(III) complex increases by a factor of 10 (k(ex)298=31x10(6) s(-1)). The [Gd(dptpa)](2-) chelate has no inner-sphere water molecule. The protonation constants of the EPTPA-bz-NO(2) (5-) and DPTPA(5-) ligands and the stability constants of their complexes with Gd(III), Zn(II), Cu(II) and Ca(II) were determined by pH potentiometry. Although the thermodynamic stability of [Gd(eptpa-bz-NO(2))(H(2)O)](2-) is reduced to a slight extent in comparison with [Gd(dtpa)(H(2)O)](2-), it is stable enough to be used in medical diagnostics as an MRI contrast agent. Therefore both this chelate and [Gd(trita)(H(2)O)](-) are potential building blocks for the development of high-relaxivity macromolecular agents.

Chelating Agents↗

PubMed2004

Thermodynamic analysis of receptors based on guanidinium/boronic acid groups for the complexation of carboxylates, alpha-hydroxycarboxylates, and diols: driving force for binding and cooperativity.

The thermodynamics of guanidinium and boronic acid interactions with carboxylates, alpha-hydroxycarboxylates, and diols were studied by determination of the binding constants of a variety of different guests to four different hosts (7-10). Each host contains a different combination of guanidinium groups and boronic acids. The guests included molecules with carboxylate and/or diol moieties, such as citrate, tartrate, and fructose, among others. The Gibbs free energies of binding were determined by UV/Vis absorption spectroscopy, by use of indicator displacement assays. The receptor based on three guanidinium groups (7) was selective for the tricarboxylate guest. The receptors that incorporated boronic acids (8-10) had higher affinities for guests that included alpha-hydroxycarboxylate and catechol moieties over guests containing only carboxylates or alkanediols. Isothermal titration calorimetry revealed the enthalpic and entropic contributions to the Gibbs free energies of binding. The binding of citrate and tartrate was investigated with hosts 7-10, for which all the binding events were exothermic, with positive entropy. Because of the selectivity of hosts 8-10, a simple boronic acid (14) was also investigated and determined to be selective for alpha-hydroxycarboxylates and catechols over amino acids and alkanediols. Further, the cooperativity of 8 and 9 in binding tartrate was also investigated, revealing little or no cooperativity with 8, but negative cooperativity with 9. A linear entropy/enthalpy compensation relationship for all the hosts 7-10, 14, and the carboxylate-/diol-containing guests was also obtained. This relationship indicates that increasing enthalpy of binding is offset by similar losses in entropy for molecular recognition involving guanidinium and boronic acid groups.

Binding Sites↗

PubMed2004

Complexing mechanism of the lanthanide cations Eu3+, Gd3+, and Tb3+ with 1,4,7,10-tetrakis(carboxymethyl)-1,4,7,10-tetraazacyclododecane (dota)-characterization of three successive complexing phases: study of the thermodynamic and structural properties of the complexes by potentiometry, luminescence spectroscopy, and EXAFS.

Complexation of the lanthanides Eu3+, Gd3+, and Tb3+ with 1,4,7,10-tetrakis(carboxymethyl)-1,4,7,10-tetraazacyclododecane (dota) has been studied in solution by using potentiometry, luminescence spectrometry, and EXAFS. Three series of successive complexes were characterized by at least two of these methods: the immediate [LnHn(dota)](n-1)+** and intermediate [LnHn(dota)](n-1)+* complexes with 0 </= n </= 2, and the final [Ln(dota)]- complexes. The formation constants of the intermediate and final complexes were determined by using potentiometry. From the results, a complexation mechanism involving three steps has been proposed. In the [LnHn(dota)](n-1)+** complexes that are instantaneously formed, the lanthanide is bound to four oxygen atoms of the carboxylate groups and to five water molecules. These species evolve rapidly: the lanthanide moves into the macrocycle cavity, two new bonds are formed with two nitrogen atoms diametrically opposed in the tetraaza cycle and only three water molecules remain bound to the lanthanide in the [LnHn(dota)](n-1)+* (0 </= n </= 2) complexes, which appear after a two-day wait. These compounds are stable for about four days. After 4-8 weeks, a concerted rearrangement occurs which leads to the formation of thermodynamically stable [Ln(dota)]- complexes in which the lanthanide is bound to four nitrogen atoms, four carboxylate oxygen atoms, and one water molecule.

Cations↗

PubMed2006

Kinetic, thermodynamic, and mechanistic patterns for free (unbound) cytochrome c at Au/SAM junctions: impact of electronic coupling, hydrostatic pressure, and stabilizing/denaturing additives.

Combined kinetic (electrochemical) and thermodynamic (calorimetric) investigations were performed for an unbound (intact native-like) cytochrome c (CytC) freely diffusing to and from gold electrodes modified by hydroxyl-terminated self-assembled monolayer films (SAMs), under a unique broad range of experimental conditions. Our approach included: 1) fine-tuning of the charge-transfer (CT) distance by using the extended set of Au-deposited hydroxyl-terminated alkanethiol SAMs [-S-(CH(2))(n)-OH] of variable thickness (n=2, 3, 4, 6, 11); 2) application of a high-pressure (up to 150 MPa) kinetic strategy toward the representative Au/SAM/CytC assemblies (n=3, 4, 6); 3) complementary electrochemical and microcalorimetric studies on the impact of some stabilizing and denaturing additives. We report for the first time a mechanistic changeover detected for "free" CytC by three independent kinetic methods, manifested through 1) the abrupt change in the dependence of the shape of the electron exchange standard rate constant (k(o)) versus the SAM thickness (resulting in a variation of estimated actual CT range within ca. 15 to 25 A including ca. 11 A of an "effective" heme-to-omega-hydroxyl distance). The corresponding values of the electronic coupling matrix element vary within the range from ca. 3 to 0.02 cm(-1); 2) the change in activation volume from +6.7 (n=3), to approximately 0 (n=4), and -5.5 (n=6) cm(3) mol(-1) (disclosing at n=3 a direct pressure effect on the protein's internal viscosity); 3) a "full" Kramers-type viscosity dependence for k(o) at n=2 and 3 (demonstrating control of an intraglobular friction through the external dynamic properties), and its gradual transformation to the viscosity independent (nonadiabatic) regime at n=6 and 11. Multilateral cross-testing of "free" CytC in a native-like, glucose-stabilized and urea-destabilized (molten-globule-like) states revealed novel intrinsic links between local/global structural and functional characteristics. Importantly, our results on the high-pressure and solution-viscosity effects, together with matching literature data, strongly support the concept of "dynamic slaving", which implies that fluctuations involving "small" solution components control the proteins' intrinsic dynamics and function in a highly cooperative manner as far as CT processes under adiabatic conditions are concerned.

Alkanes↗

PubMed2006

Homochirality as a consequence of thermodynamic equilibrium?

Recent results on the crystallisation/dissolution equilibrium of enantiomorphic crystals of NaClO3 lead to the conclusion that liquid-phase systems composed of achiral or fast racemising compounds yielding enantiomorphic solid phases (racemic conglomerates) can derive spontaneously to a single homochiral solid phase. This is a thermodynamically driven total resolution, which can only occur when the system is so perturbed that chiral recognition between the species of the system becomes feasible.

Chlorates↗

PubMed1995

Folding of aminosuccinyl peptides: thermodynamic data from temperature dependent circular dichroism measurements.

The conformational equilibrium of aminosuccinyl peptides between extended conformations and an intramolecularly hydrogen bonded type II' beta-turn conformation has been studied on the peptide Boc-L-Asu-Gly-L-Ala-OMe (Asu = aminosuccinyl residue) by means of temperature dependence of circular dichroism spectra. Owing to the peculiar chiroptical and conformational properties of the Asu residue, this technique proved to be very useful for deriving thermodynamic data for the above folding process. The value of delta H0 (-6.6 kJ mol-1), obtained for the peptide studied in a chloroformacetonitrile mixture, shows that the lower energy of the folded conformer is primarily due to the characteristic intramolecular hydrogen bond of the beta turns.

Amino Acid Sequence↗