Guido Barone

DSC studies on bovine serum albumin denaturation Effects of ionic strength and SDS concentration

This work analyzed the thermal denaturation process of defatted bovine serum albumin (BSA). DSC measurements were performed on changing the pH, the ionic strength and the sodium dodecyl sulfate (SDS) concentration. These data have been compared with those previously obtained by us and other authors. The purpose of these measurements was to study the correlation between the three-dimensional organization of BSA native protein structure and its thermodynamic stability and to clarify the non-covalent interactions between the globular proteins and amphipathic molecules. These measurements have shown that the thermal denaturation is always irreversible regardless of pH, ionic strength and SDS concentration. The nature of the irreversible process superimposed on the protein unfolding is discussed. The strong stabilizing effect of NaCl on the BSA native structure has been found for the range 0-1.0 M. It is worth noting that the calorimetric curves, confined to the pH region studied, could not be represented by a two-state transition model; they were deconvoluted as the sum of two independent two-state transitions. These transitions were correlated to the domain structure of BSA. Sodium dodecyl sulfate has a net stabilizing effect up to a molar ratio of 10:1 (ligand to protein). In this range of concentrations the presence of SDS cause a biphasic profile of excess heat capacity. A simple thermodynamic model was developed in attempt to reproduce the experimental DSC profiles and collect information regarding the binding equilibrium of SDS.

Enthalpies and entropies of sublimation, vaporization and fusion of nine polyhydric alcohols

The thermodynamics of phase transitions of nine polyhydric alcohols have been studied by different experimental approaches. The enthalpy and entropy changes associated with sublimation of pentaerythritol and myo-inositol, and the vaporization of meso-erythritol, adonitol, D-arabinitol, xylitol, dulcitol, D-mannitol, D-sorbitol and myo-inositol were determined from the temperature dependence of their vapour pressures, as measured by the torsion–effusion method. The enthalpies and temperatures of fusion were measured by differential scanning calorimetry and relevant entropies were also obtained.Enthalpies and entropies of sublimation at 298 K have been derived from the experimental data by considering the appropriate Cp values partly estimated through the group additivity schemes. Results are fairly constant for each class of polyols and in particular ΔsubX0298(tetritols)≈ΔsubX0298(pentitols) < ΔsubX0298(hexitols), but D-sorbitol and myo-inositol show lower values. Sublimation is discussed on the basis of the crystal structure data in the literature.

Theseus: A new software package for the handling and analysis of thermal denaturation data of biological macromolecules

A new software package (THESEUS) has been assembled for the analysis of the DSC data, Concerning the thermal denaturation of biological macromolecules. The system is useful to obtain accurate physico-chemical information, bypassing the casual and systematic errors, very common in these experiments. It can also be used for handling data from other instruments and methodologies giving thermodynamic, spectroscopic or other kind of data as a function of temperature. Because many of the researches in this field are of exploratory nature and continuously new unfolding mechanisms are described or hypothesized in the current literature, we have written and assembled this powerful and flexible program of general applicability, in order to put the operator in a position to control each step of the calculation procedure and use his own experience for choosing the better way to solve unexpected problems.ZusammenfassungBei der Analyse von DSC-Daten der thermischen Zersetzung von biologischen Makromolekülen wurde ein neues Softwarepaket (THESEUS) angewendet. Unter Umgehung der in derartigen Experimenten oft vorkommenden gelegentlichen und systematischen Fehler eignet sich dieses System, um genaue Physikalisch-chemische Informationen zu erhalten. Außerdem kann es zur Verarbeitung von Daten verwendet werden, welche von anderen Instrumenten und Methoden stammen, die thermodynamische, spektroskopische oder andere Daten als Funktion der Temperatur liefern. Da viele der Untersuchungen auf diesem Gebiet Forschungs-Charakter tragen und in der gegen-wärtigen Literatur ständig neue Dekonvolutionsmechanismen beschrieben oder angenommen werden, haben wir dieses leistungsstarke, flexible und allgemein anwendbare Programm geschrieben und umgesetzt, um den Operator in die Lage zu versetzen, jeden Phase des Rechenvorganges kontrollieren und seine eigene Erfahrung benutzen zu können, um den besten Weg zur Lösung unvorhergesehener Probleme zu finden.

Denaturing action of urea and guanidine hydrochloride towards two thermophilic esterases

The stability of two thermophilic esterases, AFEST from Archaeoglobus fulgidus and EST2 from Alicyclobacillus acidocaldarius, against the denaturing action of urea and guanidine hydrochloride has been investigated by means of steady-state fluorescence and circular dichroism measurements. Experimental results indicate that the two enzymes, even though very resistant to temperature and urea, show a resistance to guanidine hydrochloride weaker than expected on the basis of data collected so far for a large set of globular proteins. Structural information available for AFEST and EST2 and ideas that emerged from studies on the molecular origin of the greater thermal stability of thermophiles allow the suggestion of a reliable rationale. The present results may be an indication that the optimization of charge-charge interactions on the protein surface is a key factor for the stability of the two esterases.

Excess enthalpies of aqueous solutions of mono- and oligo-saccharide at 25°

Abstract The heats of dilution in water of d -xylose, d -fructose, d -galactose, d -mannose, lactose, and raffinose have been determined calorimetrically at 25°. The calorimetric data, expressed in terms of excess enthalpy, lead to an evaluation of pair- and triplet-interaction coefficients. Osmotic data, where known, permit the analogous coefficients of the excess free energy to be obtained and thence those of the excess entropy. Analysis of the excess functions and comparison with spectroscopic properties permits some qualitative hypotheses to be formulated on the molecular interactions occurring in these solutions.

Chiral recognition between enantiomeric α-aminoacids. A calorimetric study at 25°C

Homotactic and heterotactic enthalpic pair interaction coefficients were determined at 25°C from the enthalpies of dilution of binary and ternary aqueous solutions containing the L and D forms of α-aminoacids bearing alkyl chains of increasing length (alanine, α-aminobutyric acid, valine, norvaline, leucine, isoleucine and norleucine). Significant differences were found between the values of the homotactic (hDD or hLL) and heterotactic (hDL) pairwise coefficients, for the enantiomers of leucine and its isomers. The role of the zwitterionic interactions is discussed to explain the nature of the chiral recognition.

Vapour pressures and sublimation enthalpies of urea and some of its derivatives

Abstract The sublimation enthalpy and entropy of urea and its derivatives monomethylurea ( mmu ), monoethylurea ( meu ), monopropylurea ( mpu ), dimethyl-1,1-urea (1,1- dmu ), dimethyl-1,3-urea (1,3- dmu ), and diethyl-1,3-urea (1,3- deu ) were determined from the dependence of their vapour pressures on temperature. The vapour pressures were measured by the torsion-effusion method. The corresponding pressure against temperature equations are: urea, lg ( p kPa ) = 10.30-4750( K T ) ; mmu , lg ( p kPa ) = 10.72-4562( K T ) ; meu , lg ( p kPa ) = 10.20-4496( K T ) mpu , lg ( p kPa ) = 10.83-4608( K T ) ; 1,1- dmu , lg ( p kPa ) = 11.13-4655( K T ) ; 1,3- dmu , lg ( p kPa ) = 10.78-4454( K T ) ; 1,3- deu , lg ( p kPa ) = 12.20-5047( K T ) ; where the errors associated with the slopes (±100) and intercepts (±0.18), (±0.30 for urea), were estimated.

The hydrophobic effect in aqueous solutions of nonelectrolytes. I. Self-interactions of alkylureas

In order to clarify some aspects of the hydrophobic interactions, the enthalpies of dilution of monoethylurea, 1,3-dimethylurea, and 1,3-diethylurea have been determined calorimetrically at 25°C. The calorimetric data, expressed in terms of excess enthalpy, permit the evaluation of the pair and triplet interaction coefficients. The analyses of these and of the analogous coefficientsgxx andgxxx, derived from osmotic data, indicate a driving force favorable to the interactions among the hydrated solute molecules. Nevertheless, the positive values of thehxx andhxxx coefficients seem to suggest that the source of the effect is a rearrangement of the water molecules rather than a direct association of the solute molecules. There are evidences of a strict correlation between the enthalpic and the entropic effects.

Enthalpies of solvation for N-alkylamides in water and in carbon tetrachloride at 25°C

Molar enthalpies of solution at infinite dilution have been determined at 25°C for several N-alkyl and N,N-dialkylamides in water and in carbon tetrachloride, using a Calvet-type rotating calorimeter, and solution concentrations below 5×10−2 molal. Relevant enthalpies of transfer between the two solvents also have been derived. Molar enthalpies of solvation have been obtained by adding enthalpies of vaporization to solution values. Results are compared with those of other laboratories on other substituted amides, and their dependence on the number of carbon atoms in the chain is discussed. A possible computation of solvation enthalpies of functional groups is suggested and results for hydration of peptide or similar groups present in the compounds examined are discussed in terms of current models of their hydration and hydrogen bond formation.

Interactions in aqueous solutions of urea-like compounds. Heats of mixing of urea, monomethylurea and thiourea at 298.15 K

The heats of mixing of aqueous solutions of monomethylurea, urea and thiourea were determined at 298.15 K and the excess enthalpies obtained were expressed in terms of pair- and triplet-interaction coefficients.The validity of the empirical rule: hij2≈hij hjj, relating the self- and cross-interaction coefficients, was tested for the solutes under examination, for which both hii, hjj < 0.