Thermostable alkaline serine protease from Bacillus haloduranes C-125: gene expression and evaluation for its industrial usefulness

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Research Paper 01/08/2019
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Thermostable alkaline serine protease from Bacillus haloduranes C-125: gene expression and evaluation for its industrial usefulness

Tanveer Majeed, Shifa Ul Haq, Javed Iqbal Wattoo, Romana Tabassum
Int. J. Biosci.15( 2), 95-105, August 2019.
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Proteases are the group of hydrolytic enzymes which hydrolyzes peptide bonds in proteins showing potential applications in industrial and pharmaceutical sector. Therefore proteases production from novel strain with efficient characteristics of biotechnological interest is significant. A putative protease encoding gene (Apr) from alkaliphilic bacterium Bacillus haloduranes C-125 was successfully cloned and expressed in Bacillus subtilis expression system. Crude recombinant enzyme with specific activity of 41.5 U mg−1 was partially purified using ammonium sulfate precipitation, dialysis and ultrafiltration. The partially purified fraction was 2.8 folds having the specific activity of 116 U mg−1 of protein. The molecular weight of mature peptide was estimated as 28 kDa on SDS-PAGE. Zymographic analysis also showed the clear band on the same position. The partially purified enzyme was investigated for activity inhibition in the presence of various protease inhibitors. Serine protease nature of this enzyme was confirmed based on inhibition by PMSF. Biochemical characterization of partially purified enzyme demonstrated that it was highly active in alkaline conditions (pH10-13) and at high optimum temperature (60°C). The protease activity was studied in the presence of various oxidants, surfactants (sodium dodecyl sulphate (SDS), Triton X-100, Tween 20, H2O2, sodium perborate) and various metal ions. Enzyme showed high stability in presence of all additives which suggested the potential application of Bacillus haloduranes protease in industries specially the detergent market.


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