Purification, Characterization and Anticancer Applications of Extracellular L-asparaginase Produced by the Novel Endophytic Aspergillus aflatoxiformans AUMC 16562 | ||
| Journal of Applied Molecular Biology | ||
| Volume 3, Issue 2 - Serial Number 2974, July 2025, Pages 139-166 PDF (1.6 M) | ||
| Document Type: Original Article | ||
| DOI: 10.21608/jamb.2025.367474.1044 | ||
| Authors | ||
| Abdullah Abobakr Saleh* 1; Hamdy Mohamed El-Aref2; Azza M. Ezzeldin3; Rania M. Ewida4; Osama A.M. Al-Bedak5 | ||
| 1Molecular Biology Research & Studies Institute, Assiut University, Department of Clinical Pathology and Hematological Malignancies, South Egypt Cancer Institute, Assiut University, Assiut 71511, Egypt. | ||
| 2Molecular Biology Research & Studies Institute, Assiut University, Department of Genetics, Faculty of Agriculture, Assiut University, Assiut 71511, Egypt | ||
| 3School of Applied Health Sciences, Badr University Assiut, Department of Clinical Pathology, Faculty of Medicine, Assiut University, Assiut 71511, Egypt. | ||
| 4Department of Food Hygiene, Safety and Technology, Faculty of Veterinary Medicine, New Valley University, El-Kharga 72511, Egypt | ||
| 5Assiut University Mycological Centre, Assiut University, Assiut 71511, ERU Science & Innovation Center of Excellence, Egyptian Russian University, Badr city 11829, Cairo, Egypt. | ||
| Abstract | ||
| L-asparaginase (L-ASNase) is a multifunctional enzyme utilized for its anticancer properties. The current study delineated the optimization, purification, and application of extracellular L-ASNase produced by the endophytic Aspergillus aflatoxiformans AUMC 16562. The generation of L-ASNase was optimized in M9 medium, resulting in L-ASNase high yield with a maximum activity of 62.3 U/mL at pH 8 and 27 ºC. L-ASNase was purified using ethanol precipitation, DEAE-cellulose anion exchange chromatography, and Sephacryl S-200 HR gel filtration. In the present study, the purified L-asparaginase exhibited a molecular weight of approximately 40.5 kDa, as determined by SDS-PAGE analysis. The enzyme reached a maximum specific activity of 3668.9 U/mg under optimal conditions at pH 8.0 and 30ºC. The purification process resulted in a 12.42-fold increase in purity, with a final recovery yield of 39.82%. These findings reflect the outcomes of the current research. The purified enzyme exhibited anticancer efficacy against the human hepatocellular carcinoma cell line (HePG-2) with an IC50 of 65.2 μg/mL, the Colon cell line (HCT-116) with an IC50 of 48.2 μg/mL, and the Prostate cell line (PC-3) with an IC50 of 68.9 μg/mL. Concerning the in-vivo assay of the purified enzyme, the biochemical profiles of pure L-ASNase exhibited no effect on glucose, other electrolytes, the liver, or the kidneys. Conversely, the total bilirubin level and the activities of aspartate aminotransferase (AST) and alanine transaminase (ALT) were marginally raised. The results indicated that L-ASNase had a little impact on liver function, with liver impairment presumably signified by AST and ALT indicators. All hematological values remained within normal limits during the investigation. The present work identified a robust L-ASNase from A. aflatoxiformans AUMC 16562, which demonstrated effective cytotoxic performance against various carcinogenic cell lines under different environmental conditions, highlighting its potential utility in multiple commercial and therapeutic applications. | ||
| Keywords | ||
| Asparaginase; Anti-cancer; Cytotoxicity; Fungi; In-vivo assay; HePG-2; PC-3; HCT-116 | ||
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