Isolation, Purification and Characterization of an Extracellular Protease from a Locally Isolated Bacillus sphaericus SI-1
Journal Title: Bioresearch Communications - Year 2015, Vol 1, Issue 2
Abstract
An extracellular protease was purified from the culture supernatant of Bacillus Sphaericus strain SI1 isolated from local soil sample. The enzyme purification was primarily performed by ammonium sulfate fractionation, centrifugation and dialysis followed by centricon filtration. In each step of processing, enzyme activity was assayed by azocasein digestion method. Partially purified enzyme was further purified by sephadex G-75 column chromatography. Fraction having protease activity was visualized on SDS-PAGE. The molecular weight of the protease was 41.70 KDa, while optimum pH and temperature were 8.4 and 380C respectively. The isolated protease is positively modulated by Ca++ and Mg++ ions but significantly inhibited by EDTA, Cu++, Fe++ and Zn++ ions. The protease enzyme was slowly activated by sodium azide and inhibited by DTT and 2marcaptoethanol. The enzyme activity could be completely eliminated by phenyl methyl sulfonyl fluoride (PMSF). Therefore, all the characteristics suggest the isolated protease to be a serine protease.
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