ISOLATION, CHARACTERIZATION, PRODUCTION AND PURIFICATION OF FIBRINOLYTIC ENZYME NATTOKINASE FROM BACILLUS SUBTILISAbstract
The objective of the present study was characterization, production, and purification of Nattokinase from Bacillus subtilis. A fibrinolytic enzyme-producing bacterium was isolated and identified as Bacillus subtilis from fermented soybean and soil. Bacillus subtilis showed a very strong protease activity by degrading casein skimmed milk agar. Nattokinase was produced on basal medium and then purified from supernatant. The crude enzyme was purified by ammonium sulphate precipitation, followed by dialysis and DEAE cellulose ion-exchange chromatography. Total protein was estimated by Lowry’s method (0.58 mg/ml for FS2 and 0.49 mg/ml for S1) and enzyme-specific activity was determined by tyrosine standard (0.52 µmole/mg/ml for S1 and 0.59 µmole/mg/ml for FS2). The molecular weight of purified enzyme was determined by SDS-PAGE (20-24 kDa for FS2 and 41-44 kDa for S1). Finally, the fibrinolytic activity was determined by application of extracted enzyme on pre-existing human blood clots (FS2 showed greater activity compared to S1).
D. Sharma *, S. K. Shekhar, A. Kumar and J. Godheja
Department of Biochemistry, King George’s Medical University, Lucknow, Uttar Pradesh, India.
04 June 2019
28 November 2019
30 November 2019
01 April 2020