Denaturation
Denaturation refers to the process in which a protein's structure is altered, leading to loss of its biological activity. This can be caused by factors such as heat, pH changes, or exposure to certain chemicals. Denaturation disrupts the protein's native conformation, affecting its ability to function properly within a biological system.
5 Key excerpts on "Denaturation"
eBook - ePub- Srinivasan Damodaran(Author)
- 2017(Publication Date)
- CRC Press(Publisher)
...Cheftel et al. [ 8 ] defined protein Denaturation more specifically as any modification in conformation (secondary, tertiary, or quartenary) not accompanied by the rupture of peptide bonds involved in primary structure. The difficulty in defining Denaturation is in deciding to what extent certain modifications of the protein molecule can be included in the concept of Denaturation since a minute fraction of the protein conformation can undergo change without affecting the functional property of the protein. A more restrictive definition of Denaturation specifies the loss of one or more of the most characteristic properties of the protein, e.g., loss of solubility in solvents in which the protein was previously soluble, loss of enzymatic activity, or changes in molecular weight of the protein. Depending on the protein and conditions, Denaturation may be confined to a region of the protein or may involve the complete molecule in an “all-or-none” reaction, which reflects the cooperative nature of the transition from the native conformation to the least structured state [ 5 ]. One of the most common methods of denaturing proteins is to heat them in solution. Heat treatment of globular proteins in water or solvent increases their thermal motion, leading to the rupture of various intermolecular and intramolecular bonds stabilizing the native protein structure. This results in reorganization of both the secondary and tertiary configuration where previously inward-oriented “hydrophobic” amino acid residues become exposed to solvent. This process normally results in the formation of new short-lived intermediary conformations. Thermal coagulation is the random interaction of protein molecules by heat treatment, leading to formation of aggregates that could be either soluble or insoluble (precipitates) [ 9 ]. Thermal gelation (to be covered in a separate chapter), on the other hand, is the formation of a three-dimensional network exhibiting certain degree of order [ 9 ]...
eBook - ePub- Pieter Walstra(Author)
- 2002(Publication Date)
- CRC Press(Publisher)
...The stability, defined as the free energy difference between the folded and unfolded states, is fairly small. It is, however, the sum of two very large terms, one promoting and the other opposing unfolding. This means that small changes in conditions can already lead to unfolding. The bonds involved are for the greater part H-bonds, but these can only be strong in an apolar environment, implying that the presence of hydrophobic residues is essential in obtaining a folded, i.e., globular, conformation. Unfolding generally occurs at high or very low temperature, at extreme pH, at very high pressure, and upon adsorption onto hydrophobic surfaces (solid, oil, or air). Several solutes may cause unfolding due to altering the solvent quality, such as salts that are “high” in the Hofmeister series. Other solutes have specific effects, such as the breakage of—S—S—bridges. Denaturation of a globular protein may be equated to the unfolding of its peptide chain; it can also be related to the effects it has, such as loss of biological (e.g., enzyme) activity, or aggregation. If these changes are to be permanent, refolding of the peptide chain into its native conformation should be prevented. Several reactions, which especially occur at high temperature or high pH, can cause changes in configuration that do prevent refolding. The kinetics of Denaturation, particularly of heat Denaturation, is of great practical importance. In general, the kinetics is intricate, though in many cases the Denaturation rate is controlled by the unfolding reaction. Then, the reaction is first-order and has a very steep temperature dependence. The latter is due to the very large activation enthalpy (numerous bonds have to be broken simultaneously). The very large entropy change (e.g., the increase in conformational entropy) causes the reaction nevertheless to proceed at a reasonable rate at moderate temperatures (mostly 50-80°C)...
eBook - ePubBioseparations of Proteins
Unfolding/Folding and Validations
- Ajit Sadana, Satinder Ahuja(Authors)
- 1997(Publication Date)
- Academic Press(Publisher)
...In that case, it may be suggested that the bioseparation of the protein should be considered in its denatured state prior to steps that would facilitate its refolding to the native and active form. In some cases the Denaturation of the protein or other biological macromolecule may be a “blessing in disguise” in that powerful techniques may be utilized for the separation. Of course, in these cases the basic premise is that the protein can be renatured–refolded to its native and active form. Thus, we note the importance of: (1) the further development of refolding techniques for a wide variety of applications; and (2) the need for a better understanding of the parameters that influence the different stages in protein refolding especially as applied to obtaining the native structure vis-à-vis the inactive aggregative form. It is quite possible that as the technique of purposely unfolding a protein prior to its bioseparation becomes popular and more researchers become involved in it, newer avenues may open up that exhibit the potential to increase the efficiency (with regard to the quantity and quality of the product separated) of the bioseparation process. So that one may obtain better insights into the refolding process, the next section analyzes the mechanisms that are involved in the refolding process. III IN VITRO FOLDING MECHANISMS OF PROTEINS Protein folding is constrained both by kinetics and by thermodynamics (Jaenicke, 1991). The kinetic nature arises due to the vectorial nature of the protein synthesis, and the thermodynamic nature arises due to the necessity of energy minimization of the different states. The driving force of the three-dimensional protein structure formation is the minimization of the free energy of stabilization. The author commented on the hierarchical nature of the three-dimensional structure formation (Go, 1984 ; Jaenicke, 1991). In brief, short-ranged interactions lead to secondary structure elements...
eBook - ePub- Athel Cornish-Bowden(Author)
- 2013(Publication Date)
- Wiley-Blackwell(Publisher)
...Chapter 11 Temperature Effects on Enzyme Activity 11.1 Temperature Denaturation In principle, the theoretical treatment discussed in Section 1.8 of the temperature dependence of simple chemical reactions applies equally to enzyme-catalyzed reactions, but in practice there are several complications that must be properly understood if any useful information is to be obtained from temperature-dependence measurements. § 1.8, pages 15–21 First, almost all enzymes become denatured if they are heated much above physiological temperatures, and the conformation of the enzyme is altered, often irreversibly, with loss of catalytic activity. Denaturation is chemically a complicated and only partly understood process, and only a simplified account will be given here. In this section I shall limit it to reversible Denaturation, assuming that an equilibrium exists at all times between the active and denatured enzyme and that only a single denatured species needs to be taken into account. However, I emphasize that limiting it to reversible Denaturation is for the sake of simplicity, not because irreversible effects are unimportant in practice. Denaturation does not involve rupture of covalent bonds, but only of hydrogen bonds and other weak interactions that are involved in maintaining the active conformation of the enzyme. Although an individual hydrogen bond is far weaker than a covalent bond (about 20 kJ · mol −1 for a hydrogen bond compared with about 400 kJ · mol −1 for a covalent bond), Denaturation generally involves the rupture of many of them. More exactly, it involves the replacement of many intramolecular hydrogen bonds with hydrogen bonds between the enzyme molecule and solvent molecules...
eBook - ePubPhysical Biochemistry
Principles and Applications
- David Sheehan(Author)
- 2013(Publication Date)
- Wiley(Publisher)
...protein/nucleic acid Denaturation; hydrolysis of covalent bonds between building blocks of which biopolymers are composed). Functional effects are frequently more subtle and may be reversible (e.g. deprotonation of chemical groups in the biomolecule resulting in ionization; partial unfolding of proteins). A detailed treatment of these effects on the main classes of biomolecules is outside the scope of the present volume but a working knowledge of the likely effects of these conditions can be very useful in deciding conditions for separation or analytical manipulation. 1.2.1 pH Effects pH is defined as the negative log of the proton concentration: (1.1) Because both the H + and OH − concentrations of pure water are 10 –7 M, this scale runs from a maximum of 14 (strongly alkaline) to a minimum of 0 (strongly acidic). As it is a log scale, one unit reflects a 10- fold change in proton concentration. Most biomacromolecules are labile to alkaline or acid-catalyzed hydrolysis at extremes of the pH scale but are generally stable in the range 3–10. It is usual to analyse such biopolymers at pH values where they are structurally stable and this may differ slightly for individual biopolymers. For example, proteins normally expressed in lysosomes (pH 4) are quite acid-stable while those from cytosol (pH 7) may be unstable near pH 3. Aqueous solutions in which sample molecules are dissolved usually comprise a buffer to prevent changes in pH during the experiment. These are described in more detail in Section 1.3 below. Many biomolecules are amphoteric in aqueous solution that is they can accept or donate protons. Some chemical groups such as inorganic phosphate or acidic amino acid side-chains (e.g...
