Obtaining of Recombinant Camel Chymosin and Testing Its Milk-Clotting Activity on Cow's, Goat's, Ewes', Camel's and Mare's Milk

Simple Summary In the present work, we have described the procedure for obtaining camel milk-clotting enzyme (chymosin) by cultivating the genetically engineered yeast strain-producer in a bioreactor. This recombinant enzyme has high specific activity and versatility and has been used to produce cheese from cow's, goat's, ewes', camel's and mare's milk. The proposed biotechnological scheme for the production of chymosin seems to be effective and promising for other types of enzymes and for use in biotechnological enterprises. The enzyme will be of interest in the cheese-making industry as a potential milk-clotting enzyme. In the cheese-making industry, commonly chymosin is used as the main milk-clotting enzyme. Bactrian camel (Camelus bactrianus) chymosin (BacChym) has a milk-clotting activity higher than that of calf chymosin for cow's, goat's, ewes', mare's and camel's milk. A procedure for obtaining milk-clotting reagent based on recombinant camel chymosin is proposed here. Submerged fermentation by a recombinant yeast (Pichia pastoris GS115/pGAPZ alpha A/ProchymCB) was implemented in a 50 L bioreactor, and the recombinant camel chymosin was prepared successfully. The activity of BacChym in yeast culture was 174.5 U/mL. The chymosin was concentrated 5.6-fold by cross-flow ultrafiltration and was purified by ion exchange chromatography. The activity of the purified BacChym was 4700 U/mL. By sublimation-drying with casein peptone, the BacChym powder was obtained with an activity of 36,000 U/g. By means of this chymosin, cheese was prepared from cow's, goat's, ewes', camel's and mare's milk with a yield of 18%, 17.3%, 15.9%, 10.4% and 3%, respectively. Thus, the proposed procedure for obtaining a milk-clotting reagent based on BacChym via submerged fermentation by a recombinant yeast has some prospects for biotechnological applications. BacChym could be a prospective milk-clotting enzyme for different types of milk and their mixtures.