On the stabilizing action of protein denaturants: acetonitrile effect on stability of lysozyme in aqueous solutions

Abstract

Stability of hen lysozyme in the presence of acetonitrile (MeCN) at different pH values of the medium was studied by scanning microcalorimetry with a special emphasis on determination of reliable values of the denaturational heat capacity change. It was found that the temperature of denaturation decreases on addition of MeCN. However, the free energy extrapolation showed that below room temperature the thermodynamic stability increases at low concentrations of MeCN in spite of the general destabilizing effect at higher concentrations and temperatures. Charge-induced contribution to this stabilization was shown to be negligible (no pH-dependence was found); therefore, the most probable cause for the phenomenon is an increase of hydrophobic interactions at low temperatures in aqueous solutions containing small amounts of the organic additive. The difference in preferential solvation of native and denatured states of lysozyme was calculated from the stabilization free energy data. It was found that the change in preferential solvation strongly depends on the temperature in the water-rich region. At the higher MeCN content this dependence decreases until, at 0.06 mole fractions of MeCN, the difference in the preferential solvation between native and denatured lysozyme becomes independent of the temperature over a range of 60 K. The importance of taking into account non-ideality of a mixed solution, when analyzing preferential solvation phenomena was emphasized.

Unfolding of Ervatamin C in the Presence of Organic Solvents: Sequential Transitions of the Protein in the O-state

“…Intra-molecular hydrogen bonds are important at determining these structures, but hydrophobic interactions make the largest single contribution to stabilizing the native conformations of these macromolecules in aqueous solution. In the absence of these interactions, the stability of a molecule reduces, as evidence by thermal transitions in the presence of acetonitrile; a similar case of thermal destabilization has been reported in lysozyme in presence of acetonitrile (Kovrigin, 2000). Several hypotheses have been proposed to account for the denaturation/ inactivation of enzymes in organic solvents (Ogino and Ishikawa, 2001); (a) water molecules in the enzyme are stripped away or replaced by solvent molecules thereby causing deformation or denaturation, (b) organic molecules bind to specific enzyme sites, (c) interfacial or surface tension of solvents destroys the tertiary structure of enzymes in two phase systems.…”

Amphiphile self-aggregation: An attempt to reconcile the agreement–disagreement between the enthalpies of micellization determined by the van’t Hoff and Calorimetry methods

“…The processes on which such nonagreements appeared were mainly biological in origin, essentially those dealing with interaction of biopolymers with small molecules. A number of reports [59,[93][94][95][96][97][98][99][100][101] have recorded discrepancies between VH and calorimetric enthalpy changes for various biological interactions. The direct method of calorimetry takes care of all involved processes including adsorption, dilution, binding, desolvation, macromolecular rearrangement, etc., whereas VH only speaks for a reversible equilibrium process, and fails to record protein confor- [33], DeTAB (Ref.…”

Reproducible and efficient separation of aggregatable zein proteins by CZE using a volatile background electrolyte

“…For small ions, it has already been shown that the inverse of the effective mobility, assuming fully charged species, depends linearly on t/eZ, where Z, t and e are the solvent viscosity, the Debye dielectric relaxation time and the permittivity, respectively [34,35]. However, such general equation does not apply to proteins, since the ACN content will modify not only the pK a of the various acidic and basic sites [36] but will also modify the 3-D structure of the zein proteins [37,38] and thus their electrophoretic mobilities.…”