Identification of a Novel Extracellular Alkaline Protease-Producing <i>Bacillus cereus</i> G2 Isolated From Petroleum Sludge and Optimization of Enzyme Production

Background: Alkaline proteases are of significant utility in the field of biotechnology, with extensive applications in spanning numerous sectors, including those in the medical and health, pharmaceuticals, food, detergents, and leather industries. Methods: The bacterial strain G2, isolated from petroleum sludge at the Homs Refinery in Syria, was evaluated for its ability to produce alkaline protease using a skim milk agar medium. Protease production was confirmed through a protease assay. The molecular identification of the selected isolate was conducted through sequencing of the 16S ribosomal deoxyribonucleic acid (rDNA). Culture conditions, including variables such as inoculum size, incubation duration, temperature, pH, and various carbon and nitrogen sources, were optimized to achieve protease production. Results: Strain G2 exhibited alkaline proteolytic activity, thereby establishing its status as an effective protease producer. A 16S rDNA analysis of the intended strain revealed that the organism was identified as Bacillus cereus. The highest level of enzyme production was observed when the isolate, at an inoculum size of 3%, was cultured in the production medium for 48 hours at a pH rate of 10 and 45° C, using 1% glucose and 1% yeast extract as carbon and nitrogen sources, respectively. The alkaline protease activity was ultimately found to be 4.44-fold higher (533.3 U/mL) compared to the initial optimization of enzyme production (120 U/mL). Conclusion: This constitutes the inaugural account of the alkaliphilic B. cereus G2 strain, which was isolated from sludge petroleum and is capable of secreting alkaline protease. The findings regarding the optimal cultural conditions for the production of thermophilic alkaline protease provide a crucial foundation for future research into the potential biotechnological applications of this strain in alkaline protease production.