Purification, characterization and application of novel alkaline protease from new Bacillus cereus UV-15 mutant

  • Sreedevi Basavaraju Department of Microbiology, S. V. University, Tirupati, A.P, INDIA
  • Chandrasekhar Kathera 1Department of Microbiology, S. V. University, Tirupati, A.P, INDIA 2Jiangsu Key Laboratory for Molecular and Medical Biotechnology, College of Life Sciences, Nanjing Normal University, Nanjing, China
  • Pramoda Kumari Jasti Department of Microbiology, S. V. University, Tirupati, A.P, INDIA


The alkaline protease produced by Bacillus cereus UV-15 mutant was purified by precipitation with ammonium sulphate and gel filtration through sephadex G-100. The enzyme has shown to have a molecular weight of 29kDa by SDS polyacrylamide gel electrophoresis. The extracted protease enzyme was purified by 16.64 fold through ammonium sulphate precipitation and chromatography separation in Sephadex G-100. The purified protease had a specific activity of 2915 (U/mg). The zymogram also revealed a clear hydrolytic zone due to proteolytic activity, which coincided with the band obtained with SDS–PAGE. The enzyme was remained active and stable at pH 8-11, with an optimum at pH 10.0. The protease was stable in the temperature ranging from 40°C to 60°C, but gradually decreased at temperature 70°C. The optimum temperature for protease activity was determined at 60°C. The enzyme showed stability towards non-ionic and anionic surfactants, and oxidizing agents. At 1% concentration of Tween-20 and Tween-80, the enzyme retained 78% and 94% relative activity respectively. Alkaline protease retained 95% activity toward 0.5% concentration of the anionic detergent SDS. The enzyme showed compatibility at 50°C with commercial detergents such as Ariel, Surf excel, Rin, wheel, Tide and Nirma. In the presence of Ariel and Rin the enzyme retained about 72 and 75% of the original activity respectively. The supplementation of the enzyme in detergents could improve the cleansing performance towards the blood stains and suggested to be used as a detergent additive. The enzyme also removed goat hide hairs completely after 15 hr of incubation. These characteristics may make the enzyme suitable for several industrial applications, especially in leather industries.

Author Biography

Sreedevi Basavaraju, Department of Microbiology, S. V. University, Tirupati, A.P, INDIA
department of microbiology, NAAC A++


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