Role in protein stability

Shortle, D. 1996. The denatured state (the other half of the folding equation) and its role in protein stability. Faseb J 10 27-34. 

Mourad-Terzian, T. Steghens, J.P. Min, K.L. Collombel, C. Bozon, D. Creatine kinase isoenzymes specificities histidine 65 in human CK-BB, a role in protein stability, not in catalysis. FEBS Lett., 475, 22-26 (2000)... 

Electrostatic interactions play a minor role in protein stability [76,91], albeit they can be of importance to the specificity of protein structure or protein interactions [32],... 

In particular, Fujita and coworkers [92,112,124,125] evaluated the effect of dhp-containing ILs on the activity and stability of metalloproteins. They showed that cytochrome c, horseradish peroxidase, and ascorbate oxidase retain their activity in hydrated choline dhp ILs. Moreover, strnctnral studies (ATR-FTIR, Raman, and CD) have shown that the surroundings of the active site and the secondary structure of proteins dissolved in these media are similar to those in buffer solution. The positive effect of third-generation ILs on enzyme activity and stability was also confirmed for hydrolases, such as lipase, protease, lysozyme, and a-chymotrypsin [78, 126-129]. Recently, SAXS studies of lysozyme in choline dihydrogen phosphate IL have indicated that charge effects on the surface of the protein play a role in protein stability in this medium [130]. 

The native luciferase having a molecular weight of 106,000 probably consists of two units of the functional 19 kDa protein and two units of the 35 kDa protein. The value of A28o,icm for a solution containing 1 mg/ml of the native luciferase is calculated to be about 0.9 from the inferred amino acid sequence. The function of the 35 kDa protein remains unclear, although it might have a role in the stabilization of the 19 kDa protein. 

We present a molecular theory of hydration that now makes possible a unification of these diverse views of the role of water in protein stabilization. The central element in our development is the potential distribution theorem. We discuss both its physical basis and statistical thermodynamic framework with applications to protein solution thermodynamics and protein folding in mind. To this end, we also derive an extension of the potential distribution theorem, the quasi-chemical theory, and propose its implementation to the hydration of folded and unfolded proteins. Our perspective and current optimism are justified by the understanding we have gained from successful applications of the potential distribution theorem to the hydration of simple solutes. A few examples are given to illustrate this point. 

The aliphatic amino acids (class 1) include glycine, alanine, valine, leucine, and isoleucine. These amino acids do not contain heteroatoms (N, 0, or S) in their side chains and do not contain a ring system. Their side chains are markedly apolar. Together with threonine (see below), valine, leucine, and isoleucine form the group of branched-chain amino acids. The sulfurcontaining amino acids cysteine and methionine (class 11), are also apolar. However, in the case of cysteine, this only applies to the undissociated state. Due to its ability to form disulfide bonds, cysteine plays an important role in the stabilization of proteins (see p. 72). Two cysteine residues linked by a disulfide bridge are referred to as cystine (not shown). 

No cysteine residues are found for alpha(sl) and P-caseins do. If any S-S bonds occur within the micelle, they are not the driving force for stabilization. Caseins are among the most hydrophobic proteins, and there is some evidence to suggest that they play a role in the stability of the micelle. It must be remembered that hydrophobic interactions are very temperature sensitive. 

In the case of enzymes, the rate of the catalysed reaction increases regularly with increasing temperature. However, the probability of the unfolding of the threedimensional conformation of the protein molecule also increases, as there is more energy available to split the non-covalent interactions between side chains. In some cases it has been demonstrated that such noncovalent interactions play a dominant role in the stability of the native conformation. For example, Brosnan, Kelly and Fogarty (1992) demonstrated that the irreversible thermoinactivation of -amylase of Bacillus stearothermophilus at 90°C is related to changes of the hydrophobic interactions in the molecule. 

Phospholipase C from B. cereus contains two zinc ions per molecule of protein (molecular weight 23 000), the two metal ions being about 5 A apart. The zinc appears to have a particular role in the stabilization of the protein structure.585 Added Zn11 protects the enzyme against denaturation and induces the refolding of the denatured enzyme. Other metals are much less effective than zinc in carrying out this function.586... 

In addition to their role in primary stabilization related to viscosity increase, some hydrocolloids (particularly carrageenan) are traditionally used as secondary stabilizers. Many of the primary stabilizing hydrocolloids, including locust bean gum and carboxy methyl cellulose induce precipitation of the milk proteins in the mix. This phenomenon in ice cream mix is known as wheying-off, and may be due to direct protein-polysaccharide binding and/or protein-polysaccharide incompatibility in the water phase40. The latter phenomenon may be due to decreased solvent quality due to the competition between protein and polysaccharide for solubilisation. 

In this article we shall examine the main achievements of microcalorimetric studies of protein denaturation and of the dissolution of nonpolar substances in water. This analysis has led us to reconsider the popular point of view on the mechanism of hydrophobic interaction and its role in the stabilization of protein structures. 

Here, the charge density of the metal ion plays a pivotal role in the stability constants. The first series contains metal ions of similar size but different charges, while the ions in the second series increase with size from Mg2+ to Ba2+. In both cases, ions with higher charge densities form stronger complexes. Sometimes ions with similar sizes and charge densities can be transported by the same transport proteins. For example, Cd2+ (0.97 A) can be transported via Ca2+ (0.99 A) channels. 

Among wine polysaccharides, mannoproteins play an important role in protein haze stabilisation (Waters et al. 1994 Dupin et al. 2000). Gelatin fining of a wine phenolic extract in wine-like solution resulted in a much higher precipitation rate than when the same treatment was applied on the original wine. After addition of wine polysaccharides at the concentration normally encountered in wines, precipitation was reduced back to the level measured in wine, confirming the stabilizing effect of polysaccharides (Cheynier et al. 2006). 

Recently, it has been found that, in addition to its detoxification function and its function as a biomarker for up-regulation of other phase II enzymes, up-regulation of quinone reductase by monofunctional inducers may play a role in the stabilization of p53, the protein product of a tumor suppressor gene, which induces growth arrest and apoptosis. Sulforaphane has also been shown to mediate growth arrest and induce cell cycle arrest and apoptosis in many cancer cell lines, including those of human prostate, colon, and T-cell leukemia origin. - The exact mechanisms, and whether all the bioactivities of sulforaphane involve the ARE, are not yet understood. 

M. J. Stone, NMR relaxation studies of the role of conformational entropy in protein stability and ligand binding,... 

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