Authors:G. Barone, P. Del Vecchio, D. Fessas, C. Giancola, and G. Graziano
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.
Authors:G. Barone, S. Capasso, P. Del Vecchio, C. De Sena, D. Fessas, C. Giancola, G. Graziano, and P. Tramonti
In a previous paper, we report a preliminary DSC study on bovine (BSA) and human (HSA) serum albumins. However, at accurate
HPLC analysis the commercial proteins show three peaks: Fraction V-I, probably globulins (as declared by the producers), Fraction
V-II (about 15–18% of the product) and Fraction V-III that represents pure BSA or HSA. A hypothesis is that the Fraction II
is a covalent dimer, or trimer or a mixture of both, generated during the scalf-life of the commercial product.
Denaturation enthalpies of the purified Fraction V-III and Fraction V-II of BSA, have been determined calorimetrically, at
changing thepH, and the results of both compared with those obtained on the untreated protein. Few calorimetric experiments have been also
carried on a BSA monomer derivative with sulphidril group protected. Computer program have been developed for the deconvolution
of exo- and endothermic effects and for the analysis of thermal denaturation profiles.
Authors:Jianan Sun, Toshihiko Matsuo, and Tetsuya Uchida
-glucose molecules, has been known to protect proteins against chemical and thermal denaturation, and hence, against loss of the function [ 4 ]. For instance, trehalose stabilizes native structure of alpha-crystallin and also inhibits alpha
Authors:G. Barone, P. Del Vecchio, D. Fessas, C. Giancola, G. Graziano, and A. Riccio
DSC measurements have been accomplished in aqueous solutions of bovine pancreatic ribonuclease A (RNAase A) in the presence
of subsaturating amounts of 3′ cytidine monophosphate (3′ CMP) and 2′ cytidine monophosphate (2′ CMP) atpH 5.0 and 5.5. In these conditions the experimental profiles do not conform to a one-step unfolding process. It can be emphasized,
as a general phenomenon, that a strong linkage between the temperature-induced protein unfolding and the ligand binding, when
the ligand is less than the saturation level, causes marked distortions from a two-state transition. A purely equilibrium
thermodynamic analysis gives a correct account of this behaviour and allows to simulate calorimetric curves. It is thus possible
to obtain, in an indirect manner, information about the thermodynamic parameters concerning the binding process, namely the
association constant and the binding enthalpy. The values ofKb and ΔbH for 3′CMP and 2′CMP, so determined, are consistent with the literature data.
It is well established that the reversible thermal denaturation of small globular proteins is a cooperative two-state transition,
analogous to a first-order phase transition in a finite-size system. Finite-size effects on the cooperativity of the reversible
thermal denaturation become more important when the polypeptide chain is very small, as in the case of some synthesized mini-proteins.
The analysis of two specific examples of mini-proteins, by means of a statistical mechanical approach, leads to the conclusion
that their thermal denaturation, in view of its broadness and energetics, cannot be considered a cooperative first-order phase
The thermal denaturation of DNA and tRNAs was investigated by differential scanning calorimetry (DSC). Endothermic peaks were
observed in the DSC curves when the samples were heated. The denaturation temperatures (Td) and denaturation enthalpies (ΔHd) of samples of lyophilized powder and of samples in 4 mol/dm3 urea solution were obtained. The experimental results showed that the thermal stabilities of DNA and tRNAs were obviously
different under these conditions.
Authors:Cs. Mohácsi-Farkas, J. Farkas, L. Mészáros, O. Reichart, and É. Andrássy
Thermal stability of vegetative cells of Listeria monocytogenes, Escherichia coli and Lactobacillus plantarum was studied by counting viable fractions and determining DSC curves of their suspensions. DSC curves in the 5–99°C range showed a series of endothermic transitions between 50 and 60°C, where the heat destruction of cells occurred. Heat denaturation of DNA required a higher temperature than cell killing. Thermal death was strongly influenced by the pH, composition and NaCl content of the suspending buffer. A mathematical model developed by us enabled comparison of DSC peak temperatures and temperatures required for loss of viability.
Differential scanning calorimetry (DSC) has been employed to study the thermal denaturation processes of the main protein
fractions of blood serum. These processes have been compared for albumins (nondefatted (HSA) and fatty acid free (HSAf)),
α,β-globulins, γ-globulins, and their mixtures in aqueous (pH 6.5) and buffer (pH 7.2) solutions. The results have indicated
that α,β-globulins inhibit γ-globulins’ aggregation in buffer solutions. The decrease of stability of HSA and HSAf aqueous
solutions has been observed in the presence of γ-globulins. The mixtures of albumins and γ-globulins have revealed the tendency
to ready aggregation in water. Moreover, the results have suggested that neither γ-globulins nor albumins severely change
the stability of α,β-globulins.