Biochemical and Biological Evaluation of an L-Asparaginase from Isolated Escherichia coli MF-107 as an Anti-Tumor Enzyme on MCF7 Cell Line

Authors
Masoumeh Shahnazari 1
Razieh Bigdeli 1
Aziz Dashbolaghi 1
Reza Ahangari Cohan 2
Alireza Shoari 1
Hossein Hosseini 1
Davoud Nouri Inanlou 3
Vahid Asgary 1
1 Research and Development Laboratory, Javid Biotechnology Institute, Tehran, Iran
2 Department of Nanobiotechnology, New Technologies Research Group, Pasteur Institute of Iran, Tehran, Iran
3 Molecular and Cellular Biology Laboratory, Department of Quality Control, Pasteur Institute of Iran, Tehran, Iran
10.52547/ibj.3494
Abstract
Background: One of the most widely used anticancer agents is microbial L-ASNase. Herein, we assessed the biochemical and biological properties of an isolated L-ASNase from a Gram-negative bacteria strain, Escherichia coli MF-107.

Methods: Using garden asparagus, we obtained several bacterial isolates. These strains were further screened for L-ASNase activity. A promising bacterial isolate was selected for L-ASNase production and subsequent purification. The molecular weight of purified L-ASNase was determined. The MTT assay was applied to assess the cytotoxic effect of the purified enzyme. Also, for caspase activity determination and the apoptotic effect of purified enzyme on in cells, we conducted a real-time PCR method.

Results: The molecular weight of the enzyme was approximately 37 kDa. In the pH range of 7.5 to 8, the enzyme had considerable stability. At 35 °C, the purified L-ASNase optimum activity was recorded. The cytotoxic effect of the enzyme on treated cells was dose-dependent with an IC50 value of 5.7 IU/ml. The Bax gene expression considerably raised by 5.75-fold (p < 0.001) upon L-ASNase treatment. On the other hand, the anti-apoptotic Bcl-2 gene expression showed a 2.63-fold increase compared to the control (p < 0.05). It was detected that the mRNA levels of caspase-3 and p53 were considerably upregulated (5.93 and 1.85-fold, respectively). We did not find any alternation in the caspase-8 activity of the treated cells compared to untreated cells.

Conclusion: In this research, the proliferation of the breast cancer cells remarkably inhibited via the cytotoxic effect of isolated L-ASNase from microbial sources.
Keywords
Apoptosis
Breast cancer
Escherichia coli
MCF7 Cell Line