Casein surface hydrophobicity

The molecule must be capable of adsorbing at the oil-water or air-water interface and hence must have relatively high surface hydrophobicity the caseins, especially /3-casein, meet this requirement very well. 

The micelles on the fat surfaces cannot be coirpletely intact, because the original hydrophilic K-casein surface of the micelle is unlikely to bind to the fat surface. Homogenization must cause partial disruption of the micelles (H), allowing hydrophobic points of contact with the freshly exposed fat surfaces. There is no evidence that casein micelles interact with polystyrene latices to form a model system, for exanple (Dalgleish, unpublished results). Thus, although the micelles v ich bind to the fat in homogenized milk appear to be intact, their surfaces must have suffered some distortion, particularly of the sterically stabilizing K-casein molecules diich are near to the point of interaction of the micelle and the fat surface. 

This conclusion compares favorably with what is known from studies of the adsorption of P-casein on to air/liquid, liquid/liquid, and solid/liquid interfaces using a range of other techniques. It has generally been found that the adsorbed amount of P-casein on hydrophobic surfaces is between 2 and 3 mg m over a wide range of bulk concentrations. This is the case for planar air/water and planar oil/water interfaces (59), for hydrocarbon oil/water interfaces in emulsions (64), and for interfaces between water... 

Gatti, C.A. Risso, P.H. Pires, M.S. Spectrofluorometric study on surface hydrophobicity of bovine casein micelles in suspension and during enzymic coagulation. J. Agric. Food Chem. 1995, 48, 2339—2344. 

Amebrant and Nylander used the technique to study the sequential and competitive adsorption behavior of " C-labeled jS-lactoglobulin with KT-casein on hydrophobic and hydrophilic chromium surfaces. The adsorption was also followed by in situ ellipsometry measurements, providing a basis for comparison of the two techniques as described in Section II. 1. 

Avseenko et al. (2001) immobilized antigens onto aluminum-coated Mylar films by electrospray (ES) deposition. Various surface modifications of the metallized films were studied to determine their abilities to enhance sensitivity. The plastic surfaces were firsf cleaned by plasma discharge treatment, followed by coating with proteins (BSA and casein) or polymers such as poly (methyl methacrylate) or oxidized dextran, or they were exposed to dichlorodimethyl silane to create hydrophobic surfaces. Protein antigen was prepared in 10-fold excess sucrose and sprayed onto the surfaces to form arrays with spot diameters between 7 and 15 pm containing 1 to 4 pg protein. 

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... 

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. 

Dosaka, S., Kaminogawa, S., Taneya, S. and Yamauchi, K. 1980A. Hydrophobic surface areas and net charge of asi-, K-casein, and asi-casein /t-casein complex. J. Dairy Res. 47, 123-129. 

Like as2-casein, /e-casein has two disulfide bonds which can form cross-links with /3-lactoglobulin. The N-terminal two-thirds of the molecule is hydrophobic and contains the two disulfide bonds. The C-termi-nal end is hydrophilic, polar, and charged. It varies in the number of attached carbohydrate moieties and has only one phosphate group. These characteristics make /c-casein ideal for the surface of casein micelles, where it is most often found. It is not susceptible to calcium ion binding, as the other caseins are, and when present on the surface of micelles, it protects the other caseins from calcium (McMahon and Brown 1984A Swaisgood 1982). 

ELISA of sheep IgM (antigen) was conducted in a PDMS chip. The capture antibody (rabbit anti-sheep IgM-HRP) was added. Then the fluorogenic substrate (HPPA) was added for fluorescent detection. The conventional blocking reagents (0.5% w/v BSA, 0.5% w/v casein, 0.5% v/v Tween-20), which normally worked for the polystyrene ELISA plate, did not work with the hydrophobic PDMS surface. To solve this problem, 4% PEG and 7% normal rabbit serum were included in the above solution for effective blocking [173]. 

Historically, ideas of casein micelle structure and stability have evolved in tandem. In the earlier literature, discussions of micellar stability drew on the classical ideas of the stability of hydrophobic colloids. More recently, the hairy micelle model has focused attention more on the hydrophilic nature of the micelle and steric stabilization mechanisms. According to the hairy micelle model, the C-terminal macropeptides of some of the K-casein project from the surface of the micelle to form a hydrophilic and negatively charged diffuse outer layer, which causes the micelles to repel one another on close approach. Aggregation of micelles can only occur when the hairs are removed enzymatically, e.g., by chymosin (EC 3.4.23.4) in the renneting of milk, or when the micelle structure is so disrupted that the hairy layer is destroyed, e.g., by heating or acidification, or when the dispersion medium becomes a poor solvent for the hairs, e.g., by addition of ethanol. 

In asi-casein, it is arguable that because the hydrophobic interaction surfaces are well separated from Ca2+ ion-binding sites, the electrostatic and hydrophobic free energies of association can be treated as separate and additive, leading to the Z2 dependence of the rate of aggregation under many circumstances. Likewise, the nearly bifunctional nature of the aggregation reaction is consistent with the formation of linear polymers, as observed in the absence of Ca2+ (Thurn etal., 1987b), and may involve the apposition of hydrophobic surfaces formed from the N- and C-terminal peptides. 

---