Proteins cosolvents

The present paper is devoted to the local composition of liquid mixtures calculated in the framework of the Kirkwood—Buff theory of solutions. A new method is suggested to calculate the excess (or deficit) number of various molecules around a selected (central) molecule in binary and multicomponent liquid mixtures in terms of measurable macroscopic thermodynamic quantities, such as the derivatives of the chemical potentials with respect to concentrations, the isothermal compressibility, and the partial molar volumes. This method accounts for an inaccessible volume due to the presence of a central molecule and is applied to binary and ternary mixtures. For the ideal binary mixture it is shown that because of the difference in the volumes of the pure components there is an excess (or deficit) number of different molecules around a central molecule. The excess (or deficit) becomes zero when the components of the ideal binary mixture have the same volume. The new method is also applied to methanol + water and 2-propanol -I- water mixtures. In the case of the 2-propanol + water mixture, the new method, in contrast to the other ones, indicates that clusters dominated by 2-propanol disappear at high alcohol mole fractions, in agreement with experimental observations. Finally, it is shown that the application of the new procedure to the ternary mixture water/protein/cosolvent at infinite dilution of the protein led to almost the same results as the methods involving a reference state. 

Protein Cosolvent Molality pH Preferential binding parameter p(m) [mol/mol] Reference... 

In the present paper, a connection between the preferential binding parameter of a protein and its solubility in an aqueous solvent was established. The preferential binding parameter is a measure of the protein / water and protein / cosolvent interaction at molecular level [6,19]. Regarding the preferential binding parameter, Timasheff subdivided the cosolvents into several groups [6] When a protein molecule is immersed into a solvent consisting of water and another chemical species (a cosolvent), the interactions between the protein and the... 

Experimental data regarding 7 and V2 are available in the literature for many water/protein/cosolvent systems [4-14,18-20,22]. The Kirkwood-Buff integrals Gu and G13 are for the binary mixture water/cosolvent and ean be ealeulated as de-seribed in the literature [24-28]. The Kirkwood-Buff integrals Gi2 and G23 can be calculated Ifom Eqs. (3) and (6) using experimental data for 7 Tj, Vi and V3. 

It is noteworthy to point out that the preferential binding parameter provides an interconnection between the local and bulk properties in water+protein+cosolvent mixtures. Indeed, when the preferential binding parameter 1 23 is negative, a protein is preferentially hydrated (water is in excess), the protein is additionally stabilized and its solubility is decreased by the cosolvent. It seems that there is no exception to this rule. 

Protein Cosolvent Experimental data used Do die criteria Eqs. (10), (11) or (13), (14) work ... 

This approximation implies that the nonidealities between the protein and the constituents of the mixed solvent are much stronger than those between the constituents of the mixed solvent. In other words, in this case, the main contribution to the nonideality of the very dilute mixture protein + mixed solvent stems from the nonideality of the protein with the mixed solvent and not from the nonideality of the mixed solvent itself. This means that the activity coefficients and their derivatives with respect to the concentrations in the pairs protein—water and protein— cosolvent are much larger than for the pair water—cosolvent. 

The simplest approximation is to consider that all the sites are identical and independent, an approximation that was applied by Timasheff to several water + protein + cosolvent mixtures. In this case... 

An analysis of the cosolvent concentration dependence of the osmotic second virial coefficient (OSVC) in water—protein—cosolvent mixtures is developed. The Kirkwood—Buff fluctuation theory for ternary mixtures is used as the main theoretical tool. On its basis, the OSVC is expressed in terms of the thermodynamic properties of infinitely dilute (with respect to the protein) water—protein—cosolvent mixtures. These properties can be divided into two groups (1) those of infinitely dilute protein solutions (such as the partial molar volume of a protein at infinite dilution and the derivatives of the protein activity coefficient with respect to the protein and water molar fractions) and (2) those of the protein-free water—cosolvent mixture (such as its concentrations, the isothermal compressibility, the partial molar volumes, and the derivative of the water activity coefficient with respect to the water molar fraction). Expressions are derived for the OSVC of ideal mixtures and for a mixture in which only the binary mixed solvent is ideal. The latter expression contains three contributions (1) one due to the protein—solvent interactions which is connected to the preferential binding parameter, (2) another one due to protein/protein interactions (B p ), and (3) a third one representing an ideal mixture contribution The cosolvent composition dependencies of these three contributions... 

FIGURE 11.7 Cosolvent-induced equilibrium shift in the heat denaturation. A comparison between cf 9ln /9lna, (black) and-AG (meshed) of Equation 11.10. The eontribution from the protein-cosolvent interaction changes (meshed) dominate. (Data from S. Shimizu, 2011, Molecular Origin of the Cosolvent-Induced Changes in the Thermal Stability of Proteins, Chemical Physics Letters, 514, 156.)... 

What is even more powerful is that the molecular crowding approximation (Equation 11.7) is even reasonably valid for chemical denaturation by urea and guani-dinium ions. The protein-cosolvent interaction thus seems to be a driving force for a wider range of biochemical processes, including protein-ligand interaction, cosolvent-induced denaturation, and the cosolvent-induced modulation of thermal denaturation. 

PROTEIN-WATER AND PROTEIN-COSOLVENT INTERACTIONS 11.6.1 Theory... 

A linear correlation between preferential hydration parameter and G23, but not with G21. This suggests that the protein-cosolvent interaction is responsible for the variation of preferential hydration phenomenon. 

G23 values for the Hofmeister salts and PEGs fall onto the same line, suggesting a unifying principle underlying both the Hofmeister effect and molecular crowding namely, the variation of the protein-cosolvent interaction. 

Here, again, the protein-cosolvent interaction has been shown to be a crucial driving force describing the observed diversity of the cosolvent effect (Shimizu, McLaren, and Matubayasi 2006). 

Cosolvents ana Surfactants Many nonvolatile polar substances cannot be dissolved at moderate temperatures in nonpolar fluids such as CO9. Cosolvents (also called entrainers, modifiers, moderators) such as alcohols and acetone have been added to fluids to raise the solvent strength. The addition of only 2 mol % of the complexing agent tri-/i-butyl phosphate (TBP) to CO9 increases the solubility ofnydro-quinone by a factor of 250 due to Lewis acid-base interactions. Veiy recently, surfac tants have been used to form reverse micelles, microemulsions, and polymeric latexes in SCFs including CO9. These organized molecular assemblies can dissolve hydrophilic solutes and ionic species such as amino acids and even proteins. Examples of surfactant tails which interact favorably with CO9 include fluoroethers, fluoroacrylates, fluoroalkanes, propylene oxides, and siloxanes. 

Product recoveiy from reversed micellar solutions can often be attained by simple back extrac tion, by contacting with an aqueous solution having salt concentration and pH that disfavors protein solu-bihzation, but this is not always a reliable method. Addition of cosolvents such as ethyl acetate or alcohols can lead to a disruption of the micelles and expulsion of the protein species, but this may also lead to protein denaturation. These additives must be removed by distillation, for example, to enable reconstitution of the micellar phase. Temperature increases can similarly lead to product release as a concentrated aqueous solution. Removal of the water from the reversed micelles by molecular sieves or sihca gel has also been found to cause a precipitation of the protein from the organic phase. 

Timasheff, S.N. (1982). Preferential interactions in protein-water-cosolvent systems. In Biophysics of Water, ed. F. Franks, pp. 70-2. London John Wiley. 

Properly folded native proteins tend to aggregate less than when unfolded. Solution additives that are known to stabilize the native proteins in solution may inhibit aggregation and enhance solubility. A diverse range of chemical additives are known to stabilize proteins in solution. These include salts, polyols, amino acids, and various polymers. Timasheff and colleagues have provided an extensive examination of the effects of solvent additives on protein stability [105]. The unifying mechanism for protein stabilization by these cosolvents is related to their preferential exclusion from the protein surface. With the cosolvent preferentially excluded, the protein surface is... 

M. Buck, Trifluoroethanol and colleagues cosolvents come of age. recent studies with peptides and proteins. Q. Rev. Biophys. 31, 297 355 (1998). 

The above observations provide a clear demonstration that cosolvents in selected ranges of concentration create reversible perturbations of protein similar to those induced by other modifiers. The reversibility of the cosolvent effect is a prerequisite to cosolvent use and will depend on the concentration of cosolvent, which in turn will vary markedly with the type of solvent used. For instance, polyols can be used at concentrations up to 8 Af while methanol at 3 M causes the appearance of a new absorption band (410 nm) and, after further increases in concentration, an irreversible conversion of cytochrome P-450 into P-420. Other aliphatic alcohols cause denaturation at much lower concentrations. 

As a matter of fact, cosolvents such as primary alcohols, polyols, di-methylformamide and dimethyl sulfoxide are now almost routinely used to perturb the overall reactions and elementary equilibria or rate processes of the highly organized systems carrying out DNA, RNA, and protein synthesis. However, in spite of the fact that such systems respond well and in a reversible way to these perturbations, cosolvent effects remain relatively poor probes of reaction mechanisms (Hamel, 1972 Voigt et al., 1974 Ballesta and Vasquez, 1973 Crepin et ai, 1975 Nakanishi et al., 1974 Brody and Leautey, 1973). The most common result reported upon addition of increasing amounts of cosolvents is a bell-shaped curve equilibria and rate processes are first stimulated and... 

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