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Purification and Characterization of Microbial Protease Produced Extracellularly from Bacillus subtilis FBL-1

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Abstract

An ammonium sulfate precipitation of fermentation broth produced by Bacillus subtilis FBL-1 resulted in 2.9-fold increase of specific protease activity. An eluted protein fraction from the column chromatographies using DEAE-Cellulose and Sephadex G-75 had 94.2- and 94.9-fold higher specific protease activity, respectively. An SDS-PAGE revealed a band of purified protease at approximately 37.6 kDa. Although purified protease showed the highest activity at 45°C and pH 9.0, the activity remained stable in temperature range from 30 to 50°C and pH range from 7.0 to 9.0. Protease activity was activated by metal ions such as Ca2+, Mg2+, Mn2+, Fe2+, Ca2+ and K+, but 10 mM Fe3+ significantly inhibited enzyme activity (53%). Protease activity was inhibited by 2 mM EDTA as a metalloprotease inhibitor, but it showed good stability against surfactants and organic solvents. The preferred substrates for protease activity were found to be casein (100%) and soybean flour (71.6%).

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Authors and Affiliations

  1. Department of Food science and Technology, Yeungnam University, Gyeongsan, 38541, Korea

    Jin-Beom Si, Eun-Ju Jang & Young-Jung Wee

  2. Department of Food and Nutritional Sciences, University of Reading, Whiteknights, P.O. Box 226, Reading, RG6 6AP, UK

    Dimitris Charalampopoulos

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  1. Jin-Beom Si
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  2. Eun-Ju Jang
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  3. Dimitris Charalampopoulos
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  4. Young-Jung Wee
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Correspondence to Young-Jung Wee.

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Si, JB., Jang, EJ., Charalampopoulos, D. et al. Purification and Characterization of Microbial Protease Produced Extracellularly from Bacillus subtilis FBL-1. Biotechnol Bioproc E 23, 176–182 (2018). https://doi.org/10.1007/s12257-017-0495-3

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