Molecular Characterization and Optimization of Alkaline Protease Production by Bacillus cereus LS23B

Abstract

The study aimed to determine the conditions leading to maximum protease production using submerged fermentation and detecting the presence of protease genes in the bacteria. This is necessary to meet the increasing demand for protease enzymes in the industrial market. The ability of the bacterial isolate to produce protease enzymes was evaluated through primary screening. After that, morphological characterization, biochemical tests, and 16S rRNA analysis were done to identify the bacterial strain. To confirm the presence of the gene-encoding enzyme, the protease (npr) gene primer was amplified using a polymerase chain reaction. The pH, incubation duration, temperature, carbon, and nitrogen sources were studied. Others include metal ions, substrate concentration, and agitation speed. The bacterial strain has 100% similarity to Bacillus cereus A9, while the protease encoding gene was confirmed with positive bands of 951bp. The enzyme was optimally produced at 40 oC with a pH of 9 after 72 h incubation. Starch, gelatin, 1 % substrate concentration, 1mM K+, and agitation speed of 160 rpm fully supported protease production. The presence of the npr gene in the isolate was confirmed. Also, the optimization study reveals that Bacillus cereus LS23B can be used in large-scale protease production, which may be used in different biotechnological applications.