PHYSICAL AND CHEMICAL CHARACTERIZATION OF ALKALINE PROTEASE FROM BACILLUS SUBTILIS VBC7 USING AGRO WASTE AS SUBSTRATE

Characterization of alkaline protease

Abstract

Alkaline proteases are the most important group of industrial enzymes that hydrolyse the peptide bond of proteins into small peptides. The industrial demand of alkaline protease predominantly from the microbial origin has been recently increased and enhances the research for alkaline protease with high stability at extreme industrial conditions. Thus, this study is aimed to characterize the alkaline protease from bacterial isolate, Bacillus subtilis VBC7 screened from dairy waste dumped soil. Extracellular alkaline protease production was carried out in alkaline broth by submerged fermentation. The production medium with 10% ground nut extract at pH 10 and 40 °C enhanced the alkaline protease activity (712 U/mL) than other wastes such as coconut pulp extract and sesame seed extract. This optimized media increased the bacterial growth rate and activity of alkaline protease compared to the unsupplemented basal medium. Further, alkaline protease was partially purified and assessed their molecular weight (~30 k Da) in 12 % SDS -PAGE. The enzyme activity was observed at pH 10 and 40 °C and stable over a wide range of pH (5-12), and temperatures (10-70 °C). In addition, enzyme activity was stimulated in the presence of Mg²⁺, Ca²⁺ and Mn²⁺ and was unpretentious after treating with surfactants (SDS, Triton X-100, Tween 80), organic solvents (ethanol, methanol, chloroform, acetone and hexane) and protease inhibitors (EDTA and β-mercaptoethanol). These compatible features could lead the enzyme as a potential candidate among the industrial sectors.