The Casein Micelle

Colloidal calcium phosphate (CCP) acts as a cement between the hundreds or even thousands of submicelles that form the casein micelle. Binding may be covalent or electrostatic. The casein micelles are not static there are three dynamic equilibria between the micelle and its surroundings  

The following factors must be considered when assessing the stability of the casein micelle The role of Ca++ is very significant in milk. More than 90% of the calcium content of skim milk is associated in some way or another with the casein micelle. The removal of Ca++ leads to reversible dissociation of P-casein without micellular disintegration. The addition of Ca++ leads to aggregation. The same reaction occurs between the individual caseins in the micelle, but not as much because there is no secondary structure in casein proteins. 

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. 

Electrostatic interactions Some of the subunit interactions may be the result of ionic bonding, but the overall micellar structure is very loose and open. 

Van der Waals forces There has been some success in relating these forces to micellar stability. However, the steric stabilization has been found to be also of some importance. Especially, the hairy layer interferes with the interparticle approach. There are several factors that will affect the stability of the casein micelle system  

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Destruction of the casein micelles in the milk with subsequent precipitation of the casein can be accomplished in a number of ways. The action of heat or the action of alcohols, acids, salts and the enzyme rennet all bring about precipitation. In commercial practise the two techniques used employ either acid coagulation or rennet coagulation mechanisms. 

Dalgleish, D. G., Spagnuolo, P. A., and Douglas Goff, H. (2004). A possible structure of the casein micelle based on high-resolution field emission scanning electron microscopy. Int. Dairy. 14,1025-1031. 

The most abundant milk protein is casein, of which there are several different kinds, usually designated a-, (1-, and K-casein. The different caseins relate to small differences in their amino acid sequences. Casein micelles in milk have diameters less than 300 nm. Disruption of the casein micelles occurs during the preparation of cheese. Lactic acid increases the acidity of the milk until the micelles crosslink and a curd develops. The liquid portion, known as whey, containing water, lactose and some protein, is removed. Addition of the enzyme rennet (chymosin) speeds up the process by hydrolysing a specific peptide bond in K-casein. This opens up the casein and encourages further cross-linking. 

The Daily Industiy. The first step in cheese manufacture is the coagulation of milk. Coagulation can be divided into two distinct phases, enzymatic and the non-enzymatic. In the primary enzymatic phase a proteol ic enzyme such as chymosin (rennet), or less effectively, pepsin, carries out an extremely specific and limited proteolysis, cleaving a phenylalanine-methionine bond of /c-casein, making the casein micelle metastabie. In the second, non-enzymatic phase, the... 

Caseins are able to aggregate if the surface of the micelle is reactive. Although the casein micelle is fairly stable, there are four major ways in which aggregation can be induced ... 

A) Caserns cross-linked by die enzyme transglutaminase within the casein micelle ... 

Nature itself gives us a spectacular example of a biopolymer-based delivery system in the form of the native casein micelle of mammalian milk (Lemay et al, 2007). This is primarily a colloidal delivery system for calcium, where the micronutrient is in the form of calcium phosphate, which does not give a bitter taste, and which provides good bioavailability owing to its colloidal size, amorphous state and quick dissolution in gastric conditions (pH 1-2). Nevertheless, the casein micelle structure is unique there are no other readily available natural delivery systems for most nutraceuticals. Therefore some new designs are clearly required (Velikov and Pelan, 2008 McClements et al, 2008, 2009). 

The casein responsible for colloidal stabilization of the casein micelle is K-casein. This glycoprotein has a molecular weight of 19 kDa and is composed of 169 amino acids (Swaisgood, 2003). K-Casein is special in being the only casein component that is insensitive to calcium ions (up to concentrations of 400 mM). 

Figure 6.1 The dual binding model of the structure of the casein micelle as built up from the individual caseins (0 1, o 2, P and k) in combination with calcium phosphate (CaP). Reproduced from Home (1998) with permission.

Fox, P.F., Brodkorb, A. (2008). The casein micelle historical aspects, current concepts and significance. International Dairy Journal, 18, 677-684. 

Marchin, S., Putaux, J.L., Pignon, F., Leonil, J. (2007). Effects of the environmental factors on the casein micelle structure studied by cryo-transmission electron microscopy and small-angle X-ray scattering/ultra-small-angle X-ray scattering. Journal of Chemical Physics, 126, 45-101. 

The presence of casein-polysaccharide interactions is commonly invoked to explain the mechanistic stabilizing role of food hydrocolloids in dairy colloids (Dickinson, 1998a). Thus, for example, under conditions where the casein micelles and K-carrageenan both carry a net negative... 

Co-preclpltate is an insoluble milk protein product that is produced by heating skinimllk to high temperatures ( > 90 C) to denature the whey proteins and complex them with the casein micelles. The heated system is subsequently adjusted to isoelectric point conditions of pH 4.5-5 to precipitate the complexed whey protein-casein micelles, centrifuged or filtered to recover the precipitate, washed and dryed. The resulting product, which is virtually insoluble, exhibits only minor functionality in most typical emulsification applications. 

These changes in the concentration of Ca2 + and pH lead to destabilization of the casein micelles. 

The casein micelles are retained by fine-pore filters. Filtration through large-pore ceramic membranes is used to purify and concentrate casein on a laboratory scale. Ultrafiltration (UF) membranes retain both the caseins... 

Since the micelles are of colloidal dimensions, they are capable of scattering light and the white colour of milk is due largely to light scattering by the casein micelles the white colour is lost if the micelles are disrupted, e.g. by removing colloidal calcium phosphate (by citrate, ethylene... 

Principal micelle characteristics. The structure of the casein micelles has attracted the attention of scientists for a considerable time. Knowledge of micelle structure is important because the stability and behaviour of the micelles are central to many dairy processing operations, e.g. cheese manufacture, stability of sterilized, sweetened-condensed and reconstituted milks and frozen products. Without knowledge of the structure and properties of the casein micelle, attempts to solve many technological problems faced by the dairy industry will be empirical and not generally applicable. From the academic viewpoint, the casein micelle presents an interesting and complex problem in protein quaternary structure. 

Since the pioneering work of Waugh in 1958, a considerable amount of research effort has been devoted to elucidating the structure of the casein micelle, and several models have been proposed. This work has been reviewed in the references cited in the next section. The principal properties of the casein micelles are listed below and the models which best meet these requirements discussed briefly in the next section. 

Figure 4.20 Submicelle model of the casein micelle (from Walstra and Jenness, 1984).

Although the submicellar model of the casein micelle readily explains many of the principal features and physicochemical reactions undergone by the micelles and has been widely supported, it has never enjoyed unanimous support and two alternative models have been proposed recently. Visser (1992) proposed that the micelles are spherical conglomerates of individual casein molecules randomly aggregated and held together partly by salt bridges in the form of amorphous calcium phosphate and partly by other forces, e.g. hydrophobic bonds, with a surface layer of K-casein. Holt (1992, 1994) depicted the casein micelle as a tangled web of flexible casein... 

Figure 4.21 Model of the casein micelle (modified from Holt, 1994).

Although CCP represents only about 6% of the dry weight of the casein micelle, it plays an essential role in its structure and properties and hence has major effects on the properties of milk it is the integrating factor in the casein micelle without it, milk is not coagulable by rennet and its heat and calcium stability properties are significantly altered. In fact, milk would be a totally different fluid without colloidal calcium phosphate. 

As discussed in Chapter 2 (p. 38), crystallization of lactose as x-monohyd-rate exacerbates the situation. The combination of increased concentrations of Ca2 + and reduced pH causes destabilization of the casein micelles. 

Denaturation of whey proteins. Whey proteins are denatured very rapidly at 140°C as discussed in section 9.6.3, the denatured proteins associate with the casein micelles, via sulphydryl-disulphide interactions with K-casein, and probably with as2-casein, at pH values below 6.7. The whey proteins can be seen in electron photomicrographs as appendages on the casein micelles. 

Association and shattering of micelles. Electron microscopy shows that the casein micelles aggregate initially, then disintegrate and finally aggregate into a three-dimensional network. 

Changes in hydration. As would be expected from many of the changes discussed above, the hydration of the casein micelles decreases with the duration of heating at 140CC. The decrease appears to be due mainly to the fall in pH - if samples are adjusted to pH 6.7 after heating, there is an apparent increase in hydration on heating. 

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