Casein micelles, sensitized

We have seen earlier in this chapter how the self-assembly of casein systems is sensitively affected by temperature. Another thermodynamic variable that can affect protein-protein interactions in aqueous media is the hydrostatic pressure. Static high-pressure treatment causes the disintegration of casein micelles due to the dismption of internal hydro-phobic interactions and the dissociation of colloidal calcium phosphate. This phenomenon has been used to modify the gelation ability of casein without acidification as a consequence of exposure of hydrophobic parts of the casein molecules into the aqueous medium from the interior of the native casein micelles (Dickinson, 2006). High-pressure treatment leads to a reduction in the casein concentration required for gelation under neutral conditions, especially in the presence of cosolutes such as sucrose (Abbasi and Dickinson, 2001, 2002, 2004 Keenan et al., 2001). 

As discussed in Chapter 4, the casein micelles are stabilized by tc-casein, which represents 12-15% of the total casein and is located mainly on the surface of the micelles such that its hydrophobic N-terminal region reacts hydrophobically with the calcium-sensitive asl-, as2- and j8-caseins while its hydrophilic C-terminal region protrudes into the surrounding aqueous environment, stabilizing the micelles by a negative surface charge and steric stabilization. 

Casein micelle proteins are primarily a8i-, as2-, /3-, and -caseins in approximate proportions 3 .8 3 1. asi-Casein has eight or nine phosphate groups, depending on the genetic variant. aS2-Casein is the most hydrophilic of the caseins. It has two disulfide bonds which, by severe heat treatment, can be caused to interact with those of /3-lactoglobulin. It also has 10 to 13 phosphate groups and is very sensitive to the calcium ion concentration (Kinsella 1984 Swaisgood 1982). 

It is necessary to forewarm milk to impart adequate heat stability to the concentrate to permit it to withstand subsequent sterilization treatments. The heat-induced casein micelle-whey protein complexes in forewarmed milk are less sensitive to heat than native whey proteins and thus provide the required stability to the concentrate. The forewarming treatment also stabilizes the milk mineral system by com-plexing Ca and Mg ions with casein micelles and by converting ionic forms to the less reactive form of colloidal phosphate (Morr 1975). 

The concentrated milk is homogenized at 140 to 210 kg/cm2 (2000 to 3000 lb/in2) at about 48°C (Hall and Hedrick 1966). This process is essential to provide adequate physical stability to the milk fat emulsion system to withstand prolonged storage at room temperature (Brunner 1974). However, homogenization lowers the heat stability of concentrated milk products (Parry 1974), which may be due to increased adsorption of casein micelles onto the newly created milk fat globule surfaces, thus making them more sensitive to heat-induced aggregation. 

Many attempts have been made to locate K-casein in the casein micelle by electron microscopic methods, but the chief problems have been a lack of sensitivity in some of the methods used and doubts about the reliability of the techniques of sample preparation in not altering micellar structure. 

The temperature-sensitive precipitation of unmodified and methylated /3-caseins in the presence of calcium was measured also (see Figure 3). Methylation caused an increase of up to 3°C in the precipitation temperature of calcium /3-caseinate. Results from rennet clotting of an asi-K casein micelle system indicated that replacing native asi-casein with the reductively methylated protein had little influence on clotting time, while replacing K-casein with its reductively methylated derivative re-... 

The direct determination of some major elements (Ca, K, Mg, Na, and P) and Zn by ICP-AES was performed in powdered milk [14]. Samples were diluted with a 5 or 10 percent (v/v) water-soluble, mixed tertiary amine reagent at pH 8. This reagent mixture dissociated casein micelles and stabilized liquid phase cations. No decrease in analyte emission intensities was observed. Reference solutions were prepared in 10 percent (v/v) mixed amine solution, and no internal reference element was needed for ICP-AES. The direct technique is as fast as slurry approaches, without particle size effects or sensitivity losses. 

Acidification to the isolectric pH of casein using lactic acid bacteria or food-grade acids/acidogens, at 20 40 C. and resultant slow quiescent aggregation of the sensitized casein micelles e.g., for cream cheese. [A combination of acidification and rennet-hydrolysis (a smaller quantity of rennet than for rennet-curd cheeses, e.g., 5-100 versus 900-1000 chymosin units per 100 L milk) is normally used for low-fat acid-curd cheeses such as Quark and related varieties (Schulz-Collins and Senge, 2004)]... 

Recently, Slattery and Evard (171) proposed a model for the formation and structure of casein micelles from studies devoted to association products of the purified caseins. They proposed that the micelle is composed of polymer subunits, each 20 nm in diameter. In the micellar subunits the nonpolar portion of each monomer is oriented radially inward, whereas the charged acidic peptides of the Ca2+-sensitive caseins and the hydrophilic carbohydrate-containing portion of K-casein are near the surface. Asymmetric distribution of K-casein in a micelle subunit results in hydrophilic and hydrophobic areas on the subunit surface. In this situation, aggregation through hydrophobic interaction forms a porous micelle (Figure 10). Micelle growth is limited by the eventual concentration, at the micelle surface, of subunits rich in K-casein. 

The size of particles ranges from 20 to 600 nm (2), Casein micelles are mainly responsible for the high sensitivity of milk to physical, chemical and enzymatic treatments (3). Many models have been proposed to explain the structure of the micelle (4). However, the nature of the interactions between caseins which lead to micelle formation remains unclear. One recently proposed model presents the micelle as an aggregate of subunits ( 5) basis of this model, calcium... 

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. 

Mikheeva, L.M., Grinberg, N.V., Grinberg, V.Ya., Khokhlov, A.R., de Kruif, C.G. (2003). Thennodynaniics of micellization of bovine P-casein studied by high-sensitivity differential scanning calorimetry. Langmuir, 19, 2913-2921. 

Casein is very hydrophobic and, therefore, temperature sensitive. Low temperature or removal of calcium causes dissociation of /3-casein from the micelle and destabilizes the remaining micelle (Carpenter and Brown 1985 Dalgleish 1982). Soluble /3-casein can form aggregates of up to 40 monomers when heated. The C-terminal (hydrophobic) portions of /3-casein monomers clump together, and the N-terminal (hydrophilic) portions extend outward into the surrounding aqueous medium (Kinsella 1984). 

In the absence of calcium ions but in the presence of other casein components the normally insoluble K-casein is apparently stabilized by the calcium-sensitive caseins (157). Thus calcium ions are required to coagulate whole casein after treatment with rennin. Therefore in the native milk system the micelle-stabilizing power of K-casein is specifically destroyed by rennin, and in the presence of calcium ions in milk a coagulum is formed (2). This offers a dramatic example of how the functionality of an entire protein system can be altered by specific proteolytic action on a component of that system. 

Based on a new proposed model, each CCP nanocluster is assumed as a core and si-> o s2- ]S-caseins are linked to this core. Since / -casein just contains 1 hydrophobic site, it links to only 1 CCP nanocluster in essence, as soon as ]S-casein links to CCP (core) the growth of micelle in that direction ceases. Contrary, si- and as2-caseins are multi-functional (bi-functional) caseins and own 2 hydrophobic sites. These caseins develop the network via cross-linking since each as-casein that is linked to a CCP is able to interact with the next CCP and in this way nanoclusters link to each other. The tendency of Ca-sensitive caseins to interact with CCP is directly related to their phosphoseryl residues. The multifunctional caseins continually link CCP nanoclusters to each other and this process continues till the end nanocluster links to the first one, and a loop is formed. Bacause multi-functional caseins link randomly to each other a variety range of micelle size is obtained. The location of K-casein and its role in the stability of micelle appear to be unclear in this model [15]. 

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