Production, Purification and Characterization of Alkaline and Thermostable Protease by Shewanella putrefaciens-EGKSA21 Isolated from El-Khorma Governorate KSA

Proteases catalyzes the hydrolysis of protein. Among the various proteases, bacterial proteases are the most significant when compared with animal and fungal proteases. The aim of the present study was to production of alkaline thermostable bacterial protease for application in biodetergent technology. Screening studies were carried out for twenty one thermophilic bacterial isolates with respect to their ability to produce both protease and lipase when grown on mineral salts medium supplemented with gelatin as only source and carbon and energy at 50 C and pH 9. The most potent thermophilic bacterial isolate for production of two enzymes was identified as Shewanella putrefaciens-EGKSA21. The optimum incubation temperature and pH for maximum alkaline-thermostable protease production were 50 degrees C and 9 under fermentation conditions. Optimum substrate concentrations for protease production were 3 % gelatine. The best carbon sources that induce protease production by Shewanella putrefaciens-EGKSA21 was D (+) arabinose and D-xylose. Potassium nitrate, sodium nitrate and ammonium chloride were the optimal nitrogen sources for alkaline-thermostable protease production by Shewanella putrefaciens-EGKSA21. Maximum protease production was observed at the end of 48hrs. The overall steps protocol resulted in raising the purification fold to 411.9 times. Optimum incubation temperature and thermal-stability were 50 and 50-55 degrees C for the purified protease. The activities of the purified enzymes increased gradually with the increase of time up to 48 h incubation of the reaction mixture. The activities of the purified enzyme increased gradually by the increase of enzyme concentrations. The effect of different metalic ions on the purified enzyme activities recorded that Sodium azide (50 ppm), Lead acetate (50 ppm) and EDTA (50ppm) exhibited maximal activities while cadmium chloride and magnesium chloride inhibited the purified enzyme activities. The purified enzymes exhibited good stability towards organic solvents. The crude and purified protease of produced by Shewanella putrefaciens-EGKSA21 bacterial strain with a potential to be a candidate for the application in the detergent industry. [Bahobil, A. S. Production, Purification and Characterization of Alkaline and Thermostable Protease by Shewanella putrefaciens-EGKSA21 Isolated from El-Khorma Governorate KSA. Life Science Journal. 2011; 8(2): 580-590] (ISSN: 1097-8135). http://www.lifesciencesite.com.