Caseins secondary structures

It is believed that high level of proline comprising 17 of 199 amino acid residues in a i-casein, 10 of 207 amino acid residues in as2-casein, 35 of 209 amino acid residues in ]S-casein and 20 of 169 amino acid residues in ic-casein prevents from formation of secondary structure in caseins and thus a minute amount of a-helix and p-sheet have been observed in caseins [2], According to investigations on jc-casein, secondary structure is determined in this protein [11] in particular, helix structure in macropeptide portion and sheet structure in para-/c-casein portion of the molecule [12]. Recent publications have reported the presence of a-helix and j -sheet structures in as2-casein molecule [6, 8]. It is predicted that loop structure might be found in Ogi-, as2- and ) -caseins [8, 13],... 

Fig. 9. Comparison of FTIR absorption spectra of four proteins in H20 (left, amide I + II) and D20 (right, amide F + IF). Comparison between protein spectra for dominant secondary structure contributions from a-helix (myoglobin, MYO, top), /Fsheet (immunoglobin, IMUN), from both helix and sheet (lactoferrin, LCF) and from no long-range order (o -casein, CAS, bottom). The comparisons emphasize the high similarity, differing mostly by small frequency shifts of the amide I with the changes in secondary structure.

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. 

Caseins are highly disordered proteins having rather limited secondary structure. This is mainly due to the unusually high proline content, which is fairly uniformly distributed along the polypeptide chain. This feature leads to an open extended structure of the casein molecules which differentiates them from the globular whey proteins like a-lactalbumin and (3-lactoglobulin. The caseins have been described as rheomorphic ... 

Holt, C., Sawyer, L. (1993). Caseins as rheomorphic proteins interpretation of the primary and secondary structures of the otsi-, p- and K-caseins. Journal of the Chemical Society, Faraday Transactions, 89, 2683-2692. 

Caessens, P.W.J.R., de Jongh, H.H.J., Norde, W., Gruppen, H. (1999). The adsorption-induced secondary structure of p-casein and of distinct parts of its sequence in relation to foam and emulsion properties. Biochimica et Biophysica Acta, 1430, 73-83. 

At pH 12, the disulfide and noncovalent bonds are both broken, and the monomer with a sedimentation constant of 1.45 Svedberg units is released. From frictional ratios, the monomer appears to exist as a coil with a diameter of 16 A and a length of 150 A. Analysis of the primary structure of K-casein (Loucheux-Lefebvre et al. 1978) suggests considerable secondary structure in the monomer. 23% a-helix, 31% /3-sheets, and 24% 0-turns. In contrast, other investigators, using several different approaches, obtained a-helix contents ranging from 0 to 20.8% (Bloomfield and Mead 1975). Circular dichroism spectra on the monomer indicated 14 and 31% for a-helix and / -sheet, respectively (Loucheux-Lefebvre et al 1978). An earlier study of the optical rotatory dispersion of the K-casein monomer yielded values for the a-helix content ranging from 2 to 16% (Herskovits 1966). 

In addition to these problems, the classical CD spectrum exhibited by many proteins with a high content of P-sheet secondary structure (Fig. B3.5.6D) is quite different from those for another group of P-structure-containing proteins, of which the first identified was the WW domain (Koepf et al., 1999 not shown), and another is the mushroom inhibitor clitocypin (Fig. B3.5.6C see Kidric et al., 2002, for further examples). P-casein (Fig. B3.5.6B) shows a similar spectrum, but originating from a more complex mix of conformations (Farrell et al., 2001). 

Farrell, H.M. Jr., Wickham, E.D., Unmh, J.J., Qi, P.X., and Hoagland, P.D. 2001. Secondary structural studies of bovine caseins Temperature dependence of J-casein structure as analyzed by circular dichroism and FTIR spectroscopy and correlation with micellization. Food Hydrocolloids 15 341-354. 

A. Properties of Individual Caseins 1. Secondary Structures of Caseins... 

Haga et al. (1983) have measured the ORD and CD spectra of as2-casein in Ca2+-free phosphate buffer, pH 7.2, and estimated the content of a helix to be about four times as large as in aSi-casein, which they attribute to the lower proline content of the more phosphorylated protein. There are marked differences in the primary and predicted secondary structures of aS2-type caseins from different species. One notable constant feature, however, is a predicted a helix... 

A variety of secondary structure prediction methods has been applied to (3-casein. Regions of a helix around residues 24, 94, and 133 and (3 strands near residues 83, 147, and 190 are widely predicted (Creamer etal., 1981 Graham et al., 1984 Holt and Sawyer, 1988a,b). The predicted a helix in the N-terminal phosphopeptide region may only be stable at low pH, causing the increase in apparent helix content at pH 1.5, compared to neutrality (Creamer et al., 1981). 

Methodologies to assess interface properties of amphiphiles are surface tension measurements (Phan et al. 2006 Golding and Sein 2004 Miller et al. 2004), ellip-sometry (Dickinson 2003a Murray 2002) and Brewster angle microscopy (Grigoriev et al. 2006 Rodrguez Patino et al. 2001). Both and P NMR have been applied in order to study the conformation and dynamics of P-casein at the oil-water interface of emulsions (ter Beek et al. 1996). Their NMR results showed that the protein at the interface has mobile regions with httle secondary structure in which the motions are rather slow. 

A rapid FTIR method for the direct determination of the casein/whey ratio in milk has also been developed [26]. This method is unique because it does not require any physical separation of the casein and whey fractions, but rather makes use of the information contained in the whole spectrum to differentiate between these proteins. Proteins exhibit three characteristic absorption bands in the mid-infrared spectrum, designated as the amide I (1695-1600 cm-i), amide II (1560-1520 cm-i) and amide III (1300-1230 cm >) bands, and the positions of these bands are sensitive to protein secondary structure. From a structural viewpoint, caseins and whey proteins differ substantially, as the whey proteins are globular proteins whereas the caseins have little secondary structure. These structural differences make it possible to differentiate these proteins by FTIR spectroscopy. In addition to their different conformations, other differences between caseins and whey proteins, such as their differences in amino acid compositions and the presence of phosphate ester linkages in caseins but not whey proteins, are also reflected in their FTIR spectra. These spectroscopic differences are illustrated in Figure 15, which shows the so-called fingerprint region in the FTIR spectra of sodium caseinate and whey protein concentrate. Thus, FTIR spectroscopy can provide a means for quantitative determination of casein and whey proteins in the presence of each other. 

A segment perpendicular to a surface results in an effective steric barrier, while the number of contact points with the interface influences the strength of adsorption. For example, flexible caseins have numerous proline residues, so they have little ordered secondary structure and no intramolecular crosslink. As a result, caseins are able to adopt a number of different conformational states when being adsorbed at the oil-water interface. They are usually adsorbed at the interface in such a way that considerable portions of their structures protruding into the aqueous phase are available (Dickinson, 1992). On the other hand, serum milk proteins, such as p-lg and a-lactalbumin (a-la), bind relatively close to the interface and do not protrude... 

Raap, J., Kerling, K. E. T., Vreeman, H. E., and Visser, S. (1983). Peptide-substrates for chymosin (rennin)-conformational studies of x-casein and some x-casein-related oligopeptides by circular-dichroism and secondary structure prediction. Arch. Biochem. Biophys. 221, 117-124. 

The casein proteins tend to be a special case. Because these proteins appear not to contain much rigid secondary structure (P-helix or 13-pleated sheet) (33) and because they possess considerable numbers of hydrophobic residues (34), they adsorb well (35, 36). However, because of the lack of definition of their original native structures, it is impossible to determine whether conformational changes occur during adsorption, as neither spectroscopic changes nor DSC are capable of demonstrating conformational changes in these proteins. 

The caseins do not have much highly organised secondary structure such as a-helices or P-sheets which characterise many other proteins (Chapter 10.2). This may arise from the presence of relatively large numbers of proline groups and relatively few cystine groups which would otherwise form interchain S-S linkages. 

---