Denatured protein isolate

The denatured protein isolate (DPI) was prepared as follows. Acid precipitated curd is washed, resuspended at pH 7.0, heated at temperatures above 90 C, and spray dried. DSC measurements indicate this isolate to be totally denatured. 

Soybean Protein Isolates. Soybean protein isolates, having a protein content of >90 wt%, are the only vegetable proteins that are widely used in imitation dairy products (1). Most isolates are derived from isoelectric precipitation, so that the soybean protein isolates have properties that are similar to those of casein. They are insoluble at thek isoelectric point, have a relatively high proportion of hydrophobic amino acid residues, and are calcium-sensitive. They differ from casein in that they are heat-denaturable and thus heat-labile. The proteins have relatively good nutritional properties and have been increasingly used as a principal source of protein. A main deterrent to use has been the beany flavor associated with the product. Use is expected to increase in part because of lower cost as compared to caseinates. There has been much research to develop improved soybean protein isolates. 

WPI, whey protein isolates. Properties of nonextmded WPI pH 6.8, protein 88.9%, insoluble (denatured) 28.0%, and digestibility 87.7%. Means with different letters within a column are significantly (p < 0.05) different. 

Proteins unfolded by GdmHCl or urea will have a dominant conformation, Pn- At low temperatures we find about one-third of the residues in chemically denatured proteins in the Pn-helix conformation, with two-thirds in the form of the high-temperature ensemble. Since at least one-third of the residues in this ensemble are isolated Pn residues or in Pn helices of two or three residues, the total Pn content will be 50% or greater. The Pn content of cold- and acid-denatured proteins will be substantial, probably >40%, but not as large as in chemically denatured proteins. 

Roff, C.F., Foegeding, E.A. (1996). Dicationic-induced gelation of pre-denatured whey protein isolate. Food Hydrocolloids, 10, 193-198. 

Figure 3.2 Evolution of the microstructure of phase-separated biopolymer emulsion system containing pectin and 0.5 vt% heat-denatured (HD) whey protein isolate (WPI) stabilized oil droplets, (a) Composition 1U 3L (one-to-three mass ratio of upper and lower phases). The large circles are the water droplets (W), while the small circles are the oil droplets (O). This system forms a W2/W1-O/W1 emulsion, where O is oil, Wi is HD-WPI-rich and W2 is pectin-rich, (b) Composition 2U 2L. This system forms an 0/Wi/W2 emulsion, where O is oil, Wi is HD-WPI-rich and W2 is pectin-rich, (c) Composition 3U 1L. This system forms an 0/W]/W2 emulsion, where O is oil, Wi is HD-WPI-rich and W2 is pectin-rich. Reproduced from Kim et al. (2006) with permission.

Kinsella (13, 14) summarized present thinking on foam formation of protein solutions. When an aqueous suspension of protein ingredient (for example, flour, concentrate, or isolate) is agitated by whipping or aeration processes, it will encapsulate air into droplets or bubbles that are surrounded by a liquid film. The film consists of denatured protein that lowers the interfacial tension between air and water, facilitating deformation of the liquid and expansion against its surface tension. 

The upper aqueous phase containing nucleic acids is then separated and the DNA precipitated by addition of ethanol. Because of the ionic nature of DNA, it becomes insoluble if the aqueous medium is made less polar by addition of an organic solvent. The DNA forms a threadlike precipitate that can be collected by spooling onto a glass rod. The isolated DNA may still be contaminated with protein and RNA. Protein can be removed by dissolving the spooled DNA in saline medium and repeating the chloroform-isoamyl alcohol treatment until no more denatured protein collects at the interface. 

A good example of its use applied to a protein associated with food chemistry is that of P-lactoglobulin. Ikeuchi et al. (2001) used ANS as a probe to follow P-lactoglobulin de-naturation under high pressure and its subsequent renaturation on release of pressure (Fig. B3.6.11). The denaturation was shown to be completely reversible at pH 2 but not at neutral pH, explaining why whey protein isolates subjected to high pressures form a gel at pH 7 but not at acid pH. 

The isolation of bacterial DNA described in this experiment, patterned after the work of Marmur (1961), accomplishes these objectives. Bacterial cells are disrupted by initial treatment with the enzyme, egg-white lysozyme, which hydrolyzes the peptidoglycan that makes up the structural skeleton of the bacterial cell wall. The resultant cell walls are unable to withstand osmotic shock. Thus, the bacteria lyse in the hypotonic environment. The detergent, sodium dodecyl sulfate, (SDS, sodium do-decyl sulfate) then completes lysis by disrupting residual bacterial membranes. SDS also reduces harmful enzymatic activities (nucleases) by its ability to denature proteins. The chelating agents, citrate and EDTA (ethylenediamine tetraacetic acid), also inhibit nucleases by removing divalent cations required for nuclease activity. 

The formation of beads is a two-step process based on the cold gelation of whey proteins in the presence of divalent cations, such as Ca2+ [67], Briefly, the whey protein isolate (WPI) solution (10% w/v in deionized water) was (i) adjusted at pH 7 to favor the apparition of negative charges implied in ionic bounds with Ca2+ ions and (ii) heated (80 °C, 45 min) to denaturate the proteins. Recombinant cells in the beginning of their stationnary growth phase were suspended in a sterile solution of... 

Figure 19. Bacillus thuringiensis crystal toxin protein. Isolated protein crystals were solubilized as indicated and analyzed by denaturing polyacrylamide gel electrophoresis. Solubilization in Ellis buffer yields the 134,000 dalton protoxin.

The water-extractability of both the 7S and 11S proteins decreases with aging of the soy meal (4). During isoelectric precipitation 15-30% of the globular fraction is denatured and fails to redissolve in neutral buffer (4, 5). As the pH is lowered to 2-3 the 11S component becomes increasingly sensitive to irreversible denaturation. This acid-sensitive fraction not only limits the solubility of such isoelectric protein isolates, and therefore their functional uses, but this fraction may also be responsible for retention of undesirable flavors (5). 

Solubility. The results from the solubility experiments are given in Figures 4 and 5. The broken curve is the solubility curve for the original soy protein isolate (Purina 500 E) the nitrogen solubility of this product at neutral pH is below 40%, which indicates that the isolate has been partly denatured during its processing. The sample denoted as DH = 1.0% is the control and it is clearly demonstrated that the acid treatment has caused further aggregation and denaturation. The definitely positive DH-value... 

In protein-stabihzed foams, protein flexibility is critical to the molecule functionality in stabilizing interfaces (Hailing 1981 Lemeste et al. 1990). This has important consequences in the development and stability of dairy foams and emulsions, where the heat treatment received by the material can define its foamability and dispersion properties. A symbiotic effect between native and denatured proteins on the emulsifying properties of whey proteins isolate blends has been observed by (Britten et al. 

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