Purification and biotechnological applications of L-asparaginase from newly isolated Bacillus halotolerans OHEM18 as antitumor and antioxidant agent

Abstract

In the present study, a new bacterial strain, Bacillus halotolerans OHEM18 was significantly found to produce extracellular L-asparaginase. L-asparaginase was purified using ammonium sulfate precipitation and QFF column to 3.84-fold with specific activity of 215.33 U/mg and its molecular mass was assessed to be 41.5 kDa. Maximum enzyme activity was determined at pH 8.2 and 40 °C and with retaining 70% of its activity after incubation for 1 h at 50 °C. Km and Vmax values were determined to be 0.0047 M and 92.74, respectively. Cytotoxicity test indicated a significant safety of L-asparaginase on Vero cells with selectivity against leukemia, breast cancer and hepatoma cells. NFS-60 cells was the most sensitive tumor cells to L-asparaginase with IC₅₀ of 10.29 µg/ml and selectivity index of 30.61. This selectivity was recognized to be an apoptosis-dependent mechanism proven via cell cycle arrest in sub-G1 phase and fragmentation of genomic DNA. L-asparaginase showed antioxidant activity against both DPPH and ABTS radicals with IC₅₀ values of 64.07 and 177.1 mg/ml, respectively. These competitive advantage of bacterial L-asparaginase over than other sources is that it might be produced in large amounts through production in large-scale biofermenters, which decreases costs, besides having a sustainable bacterial source. Our findings established that the potent cytotoxic effect of L-asparaginase isolated from B. halotolerans OHEM18 may be a promise candidate for further medicinal applications as an antioxidant and antitumor drug.

Communicated by Ramaswamy H. Sarma

Keywords:

• L-asparaginase
• purification
• characterization
• anticancer
• apoptosis

Disclosure statement

No potential conflict of interest was reported by the authors.

Ethical approval

This manuscript does not contain any studies with human participants or animals performed by any of the authors.

Author contributions

Esmail M. El-Fakharany: Conceptualization, designed the study, performed the experiments, formal analysis, data curation, collected literature data, wrote and edited the manuscript. Hanaa Orabi: performed the experiments, formal analysis, collected literature data and wrote the manuscript. Eman Abdelkhalek: collected literature data and contributed to the manuscript editing. Nagwa M. Sidkey: designed the study, data curation, collected literature data and edited the manuscript.