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Abstract:  
Background. One of the most significant anticancer agents that are utilized extensively worldwide is microbial L-asparaginase. Via conducting this study we assessed the biochemical and biomedical properties of an isolated L-asparaginase from a gram-negative bacterial strain on the breast cancer cell (MCF7). Material and methods. In this research, using garden asparagus, we obtained many bacterial isolates. We further screened for the output of L-asparaginase utilizing a qualitative conventional plate assay method with bromothymol blue indicator. Then, the quantitative enzymatic assay performed on the strains revealed an encouraging outcome for L-asparaginase production, and a promising isolate was selected for L-asparaginase purification. With the sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) method the purified L-asparaginase molecular weight was determined.  The 3- (4, 5 - dimethyl - 2 - thiazolyl) - 2, 5- diphenyltetrazolium bromide (MTT) assay was applied for the study of the cytotoxic effect of the purified enzyme. Also, for caspase activity determination and the apoptotic effect in cancer cells, we conducted a real-time PCR method. Results. The molecular weight of the enzyme, which measured by SDS-PAGE analysis, was approximately 37 kDa. pH evaluation showed that in the pH range of 7.5 to 8, the enzyme had considerable stability. At 35°C, the purified L‐Asparaginase optimum activity was recorded, and with temperature increasing the enzyme activity gradually diminishing. The cytotoxic effect of the enzyme was dose-dependent with an IC50 value of 5.7 IU/ml. The Bax gene expression was considerably raised by 5.75-fold (P< 0.001). On the other hand, the anti-apoptotic Bcl-2 gene expression showed a 2.63-fold increase compared to the control (P< 0.05). As well as, it was detected that the mRNA levels of caspase-3 and p53 were considerably upregulated (5.93 and 1.85-fold respectively). L-ASNase considerably enhanced the activity of caspase-3 and -9, a major apoptotic enzyme, compared to untreated controls (P< 0.001). Note that in comparison with untreated cells, we did not find any alternation in the caspase-8 activity of the treated cells. Conclusions. The outcomes of the current research exhibited that the potential of asparagus as a powerful provenance of bacterial strains for the development of the L-asparaginase enzyme. In this research, the proliferation of the breast cancer cells remarkably inhibited via cytotoxic effect of isolated L-asparaginase from a bacteria strain. Also, cells death via the p53 dependent apoptosis mitochondrial pathway prompted via the enzyme.
     
Type of Study: Full Length | Subject: Pharmaceutical Biotechnology

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