A Study of the Structure of Bovine Casein Micelles and the Secondary Phase of Milk Coagulation Using Immobilized Enzymes
Mehaia, Mohamed Abd El-Fattah
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https://hdl.handle.net/2142/70074
Description
Title
A Study of the Structure of Bovine Casein Micelles and the Secondary Phase of Milk Coagulation Using Immobilized Enzymes
Author(s)
Mehaia, Mohamed Abd El-Fattah
Issue Date
1983
Department of Study
Food Science
Discipline
Food Science
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
Agriculture, Food Science and Technology
Abstract
The location of (kappa)-casein in casein micelles, and specifically the distribution of glyco- and carbohydrate-poor (kappa)-casein, has been studied by treating isolated casein micelles with soluble and immobilized proteases, (chymosin and pepsin) and soluble and immobilized (neuraminidase). Analysis of the reaction products gave us an indication of the surface structure of the micelle.
With both soluble and immobilized chymosin, the initial rates of release, as well as the maximum amounts released, of the glycomacropeptides is much greater than that of the non-glycomacropeptides. Immobilized chymosin released at least 88% of the glycopeptides and 78% of the non-glycopeptides released by soluble chymosin. Sialic acid data corroborated the NPN data. The results obtained with soluble and immobilized pepsin are very similar to that obtained with chymosin. With both chymosin and pepsin, the rate of, and the maximum increase in sialic acid in the 2% TCA filtrate was almost identical to that obtained with the 12% TCA filtrate. Immobilized neuraminidase released about 77% of micellar sialic acid released by soluble neuraminidase. However, even when 99% of the total sialic acid had been released by soluble neuramendase, there was no sign of coagulation. All our data points to the fact that most of the (kappa)-casein is on the surface, and further that the surface is highly glycosylated. Thus, our data favor casein micelle models that propose a surface location for (kappa)-casein.
The mechanism of the secondary phase of milk coagulation was also studied using immobilized pepsin. Skimmilk, after passing through a column of immobilized pepsin, was subjected to changes in calcium concentration, temperature and pH. A 40-fold decrease in clotting time occurred when the calcium concentration was increased from 30 to 50 mM at pH 6.6, but only a 2.8-fold decrease was observed at pH 5.6. Calcium appears to affect only the secondary phase and not the primary phase. The 30-fold decrease in clotting time when pH of milk is lowered from 6.6 to 5.6 is due most exclusively to the pH effect on the secondary phase. The secondary phase exhibited a high activation energy of 43.84 Kcal/mole compared to the primary phase which had an activation energy of 12.96 Kcal/mole. Thus it appears the physical coagulation of casein micelles is due primarily to a charge-neutralization process, together with a conformation-change type of reaction occurring in the secondary phase. The net result is a reduction of repulsion between micelles and the formation of a coagulum.
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