Isolation and Identification of an Alkaliplilic Thermo-tolerant Protease producing Bacillus sp. from Dehloran Hot Spring: Production Optimization and Investigation of the Activity and Stability of the Enzyme

Thermophilic proteases can be used in various industries, including detergents, pharmaceuticals, food, etc. These enzymes are produced by thermophilic microorganisms, includingbacteria. Sampling was carried out from Dehloran hot springs in Ilam province in the west of Iran to find new protease producing bacteria. Then, the effect of pH, temperature and finally the effect of different heating time on the production of protease enzyme were evaluated. Then, Polymerase Chain Reaction (PCR) was performed to detect the strain. Protease was purified through anion exchange using DEAE-sepharose column and its molecular weight was estimated using SDS-PAGE technique. Then, activity and stability of the enzyme were investigated in temperature and pH range. Among isolated strains, bacteria Bacillus sp. DEM07 (registered in the World Gene Bank with access number KY392988) with the highest diameter of the protease clear zone, was selected to produce thermo tolerant alkaline protease. The maximum production of the alkaline protease enzyme was observed at 50 ° C, pH 7 and 48 hours after culture. The protease enzyme was purified by anionic chromatography and its molecular weight was estimated to be about 27.5 kDa after purification. The enzyme was active and stable at the temperature range of 30 to 55 ° C and the optimum temperature of the enzyme activity was observed at 50 ° C. The pH range for activity and its stability was from 4 to 11, and the optimum activity of the enzyme was observed at pH 10.
Discussion and In this study, the protease enzyme purified from Bacillus sp. DEM07 is a thermo tolerant alkalophilic protease. On the other hand, by creating the optimal conditions for achieving high production of thermo-tolerant alkalophilic protease, this enzyme has a high potential for use in various industries.