The Casein is a protein of the milk of the phosphoprotein type which is separated from milk by acidification and form a white mass. Phosphoproteins are a group of proteins that are chemically bound to a substance that contains phosphoric acid , therefore its molecule contains a phosphorous element. Casein represents about 77 to 82 percent of the proteins present in milk and 2.7 percent in the composition of liquid milk.
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- 1 Characteristics of casein
- 2 Structure of casein micelles
- 3 Structure of casein molecules
- 1 Casein αs1
- 2 Casein αs2
- 3 Casein β
- 4 Casein κ
- 5 Casein γ
- 4 Stability of casein micelles
- 5 Uses and applications of casein
- 6 Sources
Casein is a white-yellowish solid, tasteless and odorless, insoluble in water and different organic solvents. May have a slight lactic odor. It disperses well in an alkaline medium, such as an aqueous solution of sodium hydroxide : NaOH, forming sodium caseinates.
Casein is obtained by coagulating skim milk with dilute hydrochloric acid, thus mimicking spontaneous acidification. The clots are decanted, washed with water, dried and finally ground. When it coagulates with renin, it is called paracasein, and when it coagulates through lowering the pH, it is called acid casein. When it is not coagulated it is called caseinogen. In cow’s milk, caseins represent around 80% of total protein, that is, from 25 to 28 grams per liter of milk. In human milk the presence of whey proteins is much higher, so that caseins are only on the order of half of total proteins, between 5 and 8 grams per liter.
Structure of casein micelles
Casein micelles are particles of a size between 50 and 500 nanometers, formed by the association of casein molecules together with calcium phosphate in colloidal form. The “mineral” component represents about 7% of the weight of casein. The internal structure of casein micelles. According to the most accepted model, micelles are formed by the aggregation of other smaller particles, the so-called “sub-micelles”, linked together through phosphate bridges and by hydrophobic interactions. The individual casein molecules are bound within the sub-micelles primarily through hydrophobic bonds.
the “submycels” are probably less defined structures than previously assumed, and it seems likely that the interior of the micelles has a less organized structure, with associations of casein molecules produced by their interaction with colloidal calcium phosphate particles nano-sized. The other proposed casein micelle model implies the existence of calcium phosphate nanoclusters distributed within a more or less homogeneous particle formed by the association of individual caseins.
The outer surface of the micelle is also changeable, especially sensitive to modifications of the medium that will alter the hydration of the molecules.
Phosphoserine, which is an essential component in maintaining the structure of any casein micelle model, can form bridges with calcium ions.
Calcium phosphate is found in caseins as “colloidal phosphate”, in non-crystalline form. The amorphous or colloidal phosphate particles within the caseins have a size of the order of 2.5 nanometers, and are formed by a calcium phosphate nucleus covered by a protein layer with a thickness of around 1.5 nanometers, joined to the calcium phosphate by bridges through phosphoserines.
Structure of casein molecules
The individual casein molecules are generally characterized by their medium size (about 200 amino acids , with casein κ being somewhat smaller). It has few sections with an organized secondary structure, due to the presence of abundant proline residues, and having phosphate groups covalently attached to some of the serine residues, and very occasionally to threonine residues. The lack of organization of casein molecules has so far not been able to crystallize to carry out detailed studies of their secondary and tertiary structure.
A classic property, which serves as its operational definition, is that caseins precipitate at pH 4.6, which is their isoelectric point (at room temperature).
There are four caseins, known as αs1, αs2, (the s in the subfix indicates that they are “sensitive” to calcium, that is, they can precipitate when associated with it), β and κ. The so-called “γ caseins” are simply fragments of the casein β produced by proteolysis by plasmin. All caseins have genetic variants, produced by amino acid substitution and in some cases by deletion.
There are differences in the proportion that each type represents in the total caseins. Among the most common species, the greatest differences are found in the κ casein content, which represents 3% of the buffalo milk caseins , 13% of those of cow’s milk and 26% of those of milk human.
Casein αs1 is the majority in cow’s milk. The most common variant has 199 amino acidsin its sequence, with 8 or 9 phosphate groups. From a structural point of view, it is made up of three hydrophobic regions, with two of them located at the ends (amino acids 1-41, 90-113 and 132-199), and a very polar zone (between amino acids 42 and 80) , in which all but one phosphate group are found, which gives a very significant negative net charge to the pH of milk (around 6.6). Cow’s α1 casein contains 17 proline residues, distributed along the entire chain, which means that it has very few areas with an organized secondary structure. The association with other casein molecules occurs through hydrophobic interactions in which the area located between amino acids 136 and 196 is fundamentally involved.
This casein is made up of 207 amino acids in the cow. Several genetic variants are known, and also several variants in the degree of phosphorylation. Maximum phosphorylation affects 12 serines and one threonine. This casein has a disulfide bridge between the cysteines that occupy positions 36 and 40 of the sequence, and is more hydrophilic than casein αs1, with three regions of net negative charge, one of them at the N-terminus. Hydrophobic amino acids with a net positive charge are located in the C-terminal region.
Casein β is the most hydrophobic casein, and it also has a particular structure, with a clear division into two zones. The one corresponding to the C-terminus is particularly hydrophobic, while the most hydrophilic amino acids, and all serine-bound phosphate groups, are concentrated at the N-terminus. The most common genetic variant in the cow is made up of 209 amino acids, with five phosphate groups.
Casein κ has a clearly different structure from that of other caseins. Firstly, it is somewhat smaller, it is made up of 169 amino acids in the cow . It is also very poorly phosphorylated, having only one phosphate group. This makes it interact with the calcium ion much less than the other caseins. However, it shares with casein β the property of having well-marked and separate predominantly hydrophilic and hydrophobic zones.
A peculiarity of this casein is the presence of a zone with a positive net charge between amino acids 20 and 115. This zone with a positive net charge allows the interaction of casein with polysaccharides, such as carrageenans, which are negatively charged. It also has two groups of cysteine in the chain.
Casein κ is the only casein that has part of the glycosylated molecules. The glucidic group is formed either by a trisaccharide or by a tetrasaccharide attached to a threonine residue, or in the threonine that occupies position 131, in 133, or in another one still closer to the carboxyl-terminal end of chain. Given the presence of N-acetyl neuraminic acid, this glucidic group provides a net charge to casein & kappa.
Casein κ is easily broken by proteolysis at the link between phenylalanine 105 and methionine 106, in a region rich in proline residues and probably easily accessible. When this proteolysis occurs, the N-terminal fragment 1-105 (for κ-casein), which is primarily hydrophobic, is bound to the other caseins in the micelle, while the C-terminal fragment 106-169 (casein macropptide) , highly hydrophilic, and in which the glycidic moiety is located in the glycosylated molecules, is free in solution. The breakdown of the casein κ produces the destabilization of the micelle, and (at temperatures above 20º C) its aggregation. This process is what occurs in the manufacture of the vast majority of cheeses.
Casein γ is a set of fragments of casein β formed by the action of plasmin, a proteinase present in milk. Under normal conditions, this “casein” represents about 3% of the total casein. The smallest β casein fragments formed in this proteolytic process remain in the whey, and are called the “protease-peptone fraction”.
Stability of casein micelles
Under normal conditions of pH and concentration of salts, the casein micelles are highly hydrated, having around 3.7 grams of water bound per gram of protein.
Casein micelles are fundamentally destabilized by two processes: by acidity, and by proteolysis of casein κ.
Acidity has two effects: First, as the pH decreases, the bonds between the phosphate groups and the calcium ion are broken , as the ionization of phosphates is reduced. Second, the repulsions between the micelles are reduced, as the pH approaches the isoelectric point of the caseins. At a pH of around 4.5 (and at a temperature above 20ºC) the caseins are added, forming a low mineralized curd.
In chymosin treatment , casein κ loses its hydrophilic region by proteolysis, directed towards the outside of the micelles. Reduced hydrophilicity facilitates aggregation. At low, cooling temperatures, the hydrophobic forces that hold the casein molecules together weaken, and even part of the casein leaves the micelle. The vast majority remain, but less strongly united. In particular, the forces acting on the hydrophobic region of β casein are weakened, causing it to further expose its hydrophilic region. This increases the hydration and bulkiness of the micelle. As a consequence, at refrigeration temperatures the aggregation of casein does not take place either by the action of acidity or by that of chymosin.
Uses and applications of casein
Casein is generally used in the manufacture of non-food products: glues and paints, protective covers, plastics.
Other technological uses are the clarification of wines or as an ingredient in preparations of molecular biology and microbiology (enriched media for microbial culture).
In the special diet, casein is used for the preparation of medical preparations and protein concentrates intended for the nutrition of athletes, especially after their training. Thus, it has been observed that the digestion of caseins is slower than that of soluble lactoproteins (also called seroproteins) and, therefore, more appropriate to repair the anabolism of amino acids during the period after a meal.
The so-called soluble caseins are mixtures of pure powder and / or potassium carbonate (maximum 25 p. 100) or potassium hydrogen carbonate.