Characterization and immobilization of partially purified alkaline protease extracted from rhizospheric soil bacterium, Bacillus megaterium strain EN-2 and Bacillus subtilis strain EN-3

摘要

In this study, extracellular alkaline protease producing bacterial isolates EN-2 and EN-3 from the agricultural soil of C.R.C. Pantnagar were identified as Bacillus megaterium strains EN-2 and Bacillus subtilis strain EN-3 on the basis of 16S rDNA gene sequencing. During kinetic characterization, optimum pH for EN-2 and EN-3 protease activity was 10 and 9, respectively. While optimum temperature, for maximum protease activity in both isolates, was 50°C. The crude extracellular alkaline protease from isolates EN-2 and EN-3 were partially purified using ammonium sulphate fractionation and dialysis to 1.50 and 1.42 fold with 53.77% and 42% recovery respectively. The observed values of Vmax and Km for protease from isolate EN-2 were found to be 11.57 U/ml and 17.442 mg/ml, while for EN-3 protease these were 42 U/ml and 10.62 mg/ml, respectively. The partially purified enzyme from both bacterial strains was then immobilized in sodium alginate beads with maximum immobilization efficiency at 3% (w/w) and some change in their kinetic properties. The immobilized alkaline protease from EN-2 and EN-3 showed their maximum protease activity at pH 9 and 10, and temperature 60 and 50°C, respectively. Due to these properties, isolated extracellular alkaline proteases from the two strains are ideal choice for application in detergent formulation, leather and food industries.