Immobilization of bacillus licheniformis protease enzyme for the production of cow's milk not cause to protein allergy

The most common allergy among infants is the cow's milk protein allergy and occurs in 2-5 % of all infants. With the consumption of the protein-free cow's milk, this widespread problem can be greatly reduced. In this thesis study, immobilization of Bacillus licheniformis protease was performed by covalent binding on the commercial immobilization matrix Eupergit CM. By optimizing the immobilization conditions, 100 % immobilization and 107,7 % activity yields were achieved. The optimum pH ranges (7-8) and optimum temperature (70 °C) of the enzyme has not changed after immobilization. Kinetic constants of the free and immobilized enzyme were also determined using the Lineweaver-Burk plot. The Km values for casein as substrates for free and immobilized enzyme were 0,0387 and 0,0269 g / L, while the Vmax values were 2,95 and 3,38 g L-Tyrosine / L.min respectively. The immobilized enzyme has not lost its initial activity during repeated twenty uses and twenty days of storage. Using the immobilized enzyme, semi-skimmed cow's milk proteins were hydrolyzed for two hours at the optimum temperature in an orbital shaked and heated incubator at 200 rpm shaking speed. Protein hydrolysis has been shown to be complete within 120 minutes. It has also been shown that the pH of the hydrolyzed semi-skimmed cow's milk is clearer than the unprocessed milk due to the hydrolysis of proteins, while the pH drops from 6,89 to 6,53. As a result, immobilized Bacillus licheniformis protease has been suggested to be used in the industrial production of cow's milk which does not cause protein allergy.