Production of Milk Clotting Aspartic Protease from Bacterial Species Isolated from Dumping Site of Mehmood Booti, Lahore

Abstract

Food and dairy industries play a very important role in the economy of every country.  Aspartic proteases are important enzyme of dairy industry and is used in cheese making. Previously main sources of protease enzyme were plants, animal or fungi, but due to increased demand globally they are now mostly isolated from bacteria. Objectives: To isolate the milk clotting bacteria from the soil collected from dumping site of Mehmood Booti and produce aspartic protease from them. Methods: Soil sample was collected from Mahmood Booti dumping site near ring road, Lahore. After serial dilutions, sample was inoculated on nutrient agar plates. After 24 hours at 37°C temperature, opaque, round and cream-colored colonies were observed which were sub cultured in LB agar. From there colonies were grown on selective medium made of K2HPO4, (NH4)2 HPO4, casein, MgCl2, yeast extract and agar. After incubation, a colony with clear zone was selected and grown in LB broth for enzyme production. After incubation, broth was centrifuged and supernatant was isolated. While performing protease assay, 3 mL of 5% TCA was added in the mixture. Results: The mixture remained clear which depicted the hydrolysis of casein by protease. While the test tube containing water as blank showed precipitation of casein after the addition of TCA because in this enzyme was not present. Conclusions: This shows that the isolated bacteria had the ability to produce protease which was evident from the protease activity assay and that such bacteria are abundant in dumping site