Efficient Production, Purification and Characterization of Therapeutically Significant L-Asparaginase from Bacillus licheniformis ASN51
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Genomics Research Lab, Department of Biological Sciences, International Islamic University Islamabad, Pakistan
Microbiology and Biotechnology Research Lab, Department of Biotechnology, Fatima Jinnah Women University, Rawalpindi, Pakistan
Department of Medical Laboratories, College of Applied Medical Sciences, Qassim University, Buraydah 51452, Saudi Arabia
Department of Biology, College of Science, Princess Nourah bint Abdulrahman University, P.O. Box 84428, Riyadh 11671, Saudi Arabia
Submission date: 2023-02-18
Final revision date: 2023-04-08
Acceptance date: 2023-05-15
Online publication date: 2023-07-12
Publication date: 2023-08-11
Corresponding author
Shaheen Shahzad   

Genomics Research Lab, Department of Biological Sciences, International Islamic University, Islamabad, Pakistan
Pol. J. Environ. Stud. 2023;32(5):4267–4280
L-asparaginase (L-ASNase) is a versatile anticancer and acrylamide reducing enzyme used in medical and food industries. Exploration of new sources of L-asparaginase is in demand due to immunogenic issues, short half-life and narrow range of pH and temperature stability of available formulations. The present study describes extracellular Type II L-ASNase production from soil Bacillus licheniformis ASN51 strain. Optimized asparaginase production with a specific activity of 499 U mg-1 was obtained at pH 8 and 37ºC in M9 medium containing 0.2% glucose and 1% L-asparagine. L-ASNase was precipitated using acetone and purified through Sephadex G-100 column yielding 61.2 and 34% recovery, respectively. The purified asparaginase has a molecular mass of 38 kDa and specific activity of 8,333 U mg-1. L-ASNase showed stability at a wide range of pH (4-9) and temperature (10-50ºC) while retained 100% of its activity for 24 h at 37ºC, pH 7. Enzyme kinetics revealed a Vmax of 7750 U and Km of 0.04 mM. Purified L-ASNase from B. licheniformis ASN51 showed antioxidant activity (IC50 value 53.1 μg mL-1) against 2, 2-diphenyl-1-picrylhydrazyl (DPPH) radical and anticancer activity (53.3%) against HepG2 cell line. To the best of knowledge, among reported strains of B. licheniformis, ASN51 is the highest asparaginase producing strain. Purified L-ASNase showed long-term stability at physiological pH and temperature indicating its suitability for therapeutic applications.