Pasteur Institute of Iran
The goal of this research was to isolate and identify the thermostable alkaline protease producing bacteria among several native Iranian microorganisms. At the end of screening program, a Bacillus subtilis BP-36 strain producing thermophilic alkaline protease was isolated from a hot spring in Ardebil province. The target enzyme was purified using a one-step Aqueous two-phase systems (ATPS) protocol involving 22% (w/w) polyethylene glycol (PEG)-10,000, and 18% (w/w) citrate with a yield of 39.7%, specific activity of 2600 U/mg and purification factor of 4.8. It was shown to have a molecular weight of 40 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The purified thermophile enzyme was stable in alkaline pH range (9.0-11.0) with the optimum pH of 9.0. It was highly stable at 60 °C and retained 100% activity even after 90 minutes, suggesting that it belong to the family of thermophilous. Collectively, our obtained data revealed that the thermophilic protease produced by B. subtilis BP-36 has the potential application in industrial processes under high temperature.