Pressure denaturation of protein

Hummer, G., Garde, S., Garcia, A. E., Paulaitis, M. E., and Pratt, L. R. (1998b). The pressure dependence of hydrophobic interactions is consistent with the observed pressure denaturation of proteins. Proc. Natl. Acad. Sci. USA 95, 1552-1555. Hummer, G., Garde, S., Garcia, A. E., Pohorille, A., and Pratt, L. R. (1996). An information theory model of hydrophobic interactions. Proc. Natl. Acad. Sci. USA 93, 8951-8955. 

Pressure denaturation of protein has been one of the problems in the focus of protein research due not only to its significance in science [47-49], but also to its importance in industrial applications, including food processing [50], The molecular mechanism of the process has not been clarified for a long time, especially concerning the role played by water or hydration. We have applied the RISM/3D-RISM theory to this problem to clarify the molecular mechanism behind the thermodynamics process [51]. 

An analysis of computer simulations of water at different pressures by Hummer et al. (110) suggested that hydrophobic contact pairs become increasingly destabilized with increasing pressure. The proposed scenario could explain the pressure denaturation of proteins as a swelling in terms of water molecules that enter the hydrophobic core by creating water-separated hydrophobic contacts. Additional support for the validity of Hummer s IT-model analysis has been achieved by pressure-dependent computer simulation studies of isolated pairs of hydrophobic particles, as well as rather concentrated solutions of hydrophobic particles (111, 112). Recently, the pressure-induced swelling of a polymer composed of apolar particles at low temperatures can be observed (113). 

Hummer G, Garde S, Garca AE, Paulaitis ME, Pratt LR. The pressure dependence of hydrophobic interactions is consistent with the observed pressure denaturation of proteins. Proc. Natl. Acad. Sci. U.S.A. 1998 95 1552-1555. 

Pressure denaturation of proteins. For a typical protein, folding can be regarded as involving two states, native N) and denatured (D),... 

Korhonen, H., Pihlanto-Leppala, A., Rantamaki, P., and Tupasela, T. (1998). Impact of processing on bioactive proteins and peptides. Trends Food Sci. Technol. 9,307-319. Kunugi, S. and Tanaka, N. (2002). Cold denaturation of proteins under high pressure. Biochim. Biophys. Acta 1595, 329-344. 

The simplicity and accuracy of such models for the hydration of small molecule solutes has been surprising, as well as extensively scrutinized (Pratt, 2002). In the context of biophysical applications, these models can be viewed as providing a basis for considering specific physical mechanisms that contribute to hydrophobicity in more complex systems. For example, a natural explanation of entropy convergence in the temperature dependence of hydrophobic hydration and the heat denaturation of proteins emerges from this model (Garde et al., 1996), as well as a mechanistic description of the pressure dependence of hydrophobic... 

In this equation Ap is the compressibility factor difference (P =PV) and Aa the difference of the thermal expansion factor (a =aV) of the denatured and native states of proteins. An important assumption in the derivation of this equation is the temperature and pressure independence of Aa, AP and ACp. The AG=0 curve is an ellipse on the P-T plane and it describes the equilibrium border between the native and denatured state of the protein. This curve is known as the phase or stability diagram. This is visualized in Fig. 2. The diagram illustrates the interconnection between the cold, heat and pressure unfolding of proteins. 

Inverse melting, another expression for the same phenomena, has now been computer modelled with a statistical mechanical model [89]. However, the challenge remains to simulate the crucial role of water in the phase diagram of proteins. A possible mechanism has been proposed for the cold denaturation of proteins at high pressure [90]. 

The stabilizing (destabilizing) effect of pressure on the thermal denaturation of proteins has been associated with the presence (absence) of aggregation of the unfolded protein [79]. Intermolecular aggregation is indeed one of the most commonly observed effects of thermal denaturation. The temperature of unfolding may however be lowered considerably by a... 

The partial molar volume is a thermodynamic quantity that plays an essential role in the analysis of pressure effects on chemical reactions, reaction rate as well as chemical equilibrium in solution. In the field of biophysics, the pressure-induced denaturation of protein molecules has continuously been investigated since an egg white gel was observed under the pressure of 7000 atmospheres [60]. The partial molar volume is a key quantity in analyzing such pressure effects on protein conformations When the pressure in increased, a change of the protein conformation is promoted in the direction that the partial molar volume reduces. A considerable amount of experimental work has been devoted to measuring the partial molar volume of a variety of solutes in many different solvents. However, analysis and interpretation of the experimental data are in many cases based on drastically simplified models of solution or on speculations without physical ground, even for the simplest solutes such as alkali-halide ions in aqueous solution. Matters become more serious when protein molecules featuring complicated conformations are considered. 

The principle of steam sterilisation for medical devices, pharmaceutical products and utensils is based on heat transfer by hot condensing steam under pressure. The steam condenses in the autoclave to pure water, releasing at that moment its heat content. This is a very effective means of heat transfer. Furthermore, the mechanism of inactivation by saturated steam (denaturation of proteins) is also very effective. Therefore, steam sterilisatirai in an autoclave is the preferred method for medical devices, utensils and some pharmaceutical products. It is of critical importance that the steam in a steam autoclave is completely saturated and not superheated, because only then the sterilisation is effective. For the details of steam sterilisation reference is made to other textbooks and guidance, such as [4,5]. The pressure of saturated steam at different temperatures is shown in Table 30.2. 

Pressure-induced protein denaturation. Figure 2.5.28 shows a (p, T) phase diagram for the denaturation of proteins (1 b l = 1 atm). 

R. K. Cho, J. H. Lee, Y. Ozaki, M. Iwamoto. FT-NIR monitoring system for the study of denaturation of proteins at elevated pressures and temperatures. In Leaping ahead with Near Infrared Spectroscopy, Proceedings of the Sixth International Conference on Near-Irfrared Spectroscopy. G. D. Batten, P. C. Flinn, L. A. Welsh, A. B. Blakeney, co-eds. The NIR Spectroscopy Group, Royal Australian Chemical Institute, 1/21 Vale Street, North Melbourne, Australia, 1995, p. 483-486. 

The denaturation of proteins does not just occur during thermal processing of foods, but also can be the result of low temperatures (when food is exposed to freezing temperatures of between 0 and -15 °C). As a result, crystals of ice are produced, which can break the cell membranes, followed by denaturation due to surface phenomena at the interface of the two phases (protein solution and ice crystals), or possibly due to the freezing of water needed to maintain the native conformation of the protein. Loss of water also leads to an increase in the salt concentration and osmotic pressure, which can accelerate protein denaturation. Lipoproteins are particularly sensitive to denaturation (e.g. those in egg yolk). The extent of the denaturation is Umited in rapid freezing at very low temperatures, however, as smaller ice crystals are formed. 

It is important to point out that during the pressure-assisted cold denaturation many proteins show significant residual secondary structure which parallels the structure of folding intermediates. It appears that pressure denaturation and pressure-assisted cold denaturation of proteins studied by high resolution NMR techniques provide novel information about the folding of proteins. In addition, these experiments allow the determination of phase diagrams for proteins. Figure 12 shows the... 

Quantitative protein analyses can be performed by applying a technique of resolution enhancement such as Fourier self-deconvolution and by referring to empirical rules on the correspondence between the wavenumbers of amide I bands and secondary structures [42,43]. Figure 2.9 shows, as a typical example, the deconvolved amide I band of lysozyme recorded in D2O solution [43], Noticeably, the denaturation of proteins as a consequence of changes in temperature, pH and pressure can be studied by measuring changes in the amide I band. 

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