Aboulnaga, E. (2019). Cloning and Expression of Camel Pro-Chymosin Encoding Gene in E. coli and Characterization of the Obtained Active Enzyme. Journal of Food and Dairy Sciences, 10(3), 71-78. doi: 10.21608/jfds.2019.36156
E. A. Aboulnaga. "Cloning and Expression of Camel Pro-Chymosin Encoding Gene in E. coli and Characterization of the Obtained Active Enzyme". Journal of Food and Dairy Sciences, 10, 3, 2019, 71-78. doi: 10.21608/jfds.2019.36156
Aboulnaga, E. (2019). 'Cloning and Expression of Camel Pro-Chymosin Encoding Gene in E. coli and Characterization of the Obtained Active Enzyme', Journal of Food and Dairy Sciences, 10(3), pp. 71-78. doi: 10.21608/jfds.2019.36156
Aboulnaga, E. Cloning and Expression of Camel Pro-Chymosin Encoding Gene in E. coli and Characterization of the Obtained Active Enzyme. Journal of Food and Dairy Sciences, 2019; 10(3): 71-78. doi: 10.21608/jfds.2019.36156
Cloning and Expression of Camel Pro-Chymosin Encoding Gene in E. coli and Characterization of the Obtained Active Enzyme
Chymosin is considered the main enzyme in milk industry since it is used for commercial production of cheese. Conventional animal chymosin production losses large numbers of unweaned calves which affect on the animal wealth. In order to reduce the manufacturing cost, recombinant production of chymosin is the good choice. In the current study, camel pro-chymosin gene (1101 bp) was in vitro synthesized and cloned into pASG-vector. The obtained plasmid pASG_pro.chym was transformed into the bacteria E. coli BL21(DE3). The bacterial system expressed pro-chymosin under control with tet-promoter and it was able to produce approximately 260 mg/L of recombinant enzyme under lab scale. The SDS-PAGE showed that the zymogen protein (367 amino acids residues long) has a molecular weight of 40.6 kDa, while the active form (323 amino acids residues long) has a molecular weight of 35.6 kDa. The recombinant pro-chymosin is presented in inclusion bodies and it is solubilized in 4-8 M urea. After solubilization and renaturation, recombinant pro-chymosin was subjected to a low pH and it was converted into mature active chymosin. The optimum milk clotting conditions were a pH of 5.75, temperature of 50-55 °C, and 15 mM of CaCl2. It can be concluded that the obtained recombinant chymosin from E. coli is suitable for commercial cheese production.
View on SCiNiTO
Top