Lailaja V P, Sumithra T G, Sharma S R Krupesha, Mohammed M Ashif, Sivan Unnikrishnan, Anusree V N
Marine Biotechnology, Fish Nutrition and Health Division, ICAR-Central Marine Fisheries Research Institute (CMFRI), Ernakulam North P.O, Post Box No. 1603, Kochi, 682 018, India.
Cochin University of Science and Technology Kochi, Kochi, Kerala, 682022, India.
Sci Rep. 2025 Jul 23;15(1):26759. doi: 10.1038/s41598-025-08962-8.
L-asparaginase is a vital antineoplastic drug. The side effects of the current formulations intrigued researchers to explore novel candidates. The study evaluated a novel type-II L-asparaginase from Chryseomicrobium amylolyticum from marine crab, Scylla serrata. Statistical optimization enhanced production to 281.6 from 115.69 IU/ml. The purified enzyme had 173.69 IU/mg protein specific activity. The Km, Vmax, and Kcat were 6.364 µM, 909.09 µM/min, and 222.82 /sec. The enzyme didn't react with L-glutamine, D-glutamine, L-glutamic acid, and urea, a clinically desirable feature. In vitro trypsin and serum half-life were ~ 39 h and ~ 27 min. The study enlisted ions, protein-modifying agents, and surfactants influencing its activity. The detailed in silico analyses of the functional gene predicted its physicochemical and structural features. The low sequence identity with the commercial enzymes and the predicted nonallergenic and nontoxic features suggested its clinical suitability. Molecular docking revealed a higher binding affinity for L-asparagine than L-glutamine and urea. The IC50 against MCF7, MOLT4 and THP-1 cell lines were 0.043, 0.041, and 0.05 IU/ml, respectively. Phase contrast microscopy confirmed its cytotoxicity. Flow cytometry demonstrated 81.4% apoptotic cells in MOLT-4 cells at IC50, signifying potent anticancer activity. The enzyme did not affect normal human fibroblast cells and was non-haemolytic. The results underscore the therapeutic potential of C. amylolyticum type-II L-asparaginase and support further in vivo studies to advance L-asparaginase-based cancer therapies.
L-天冬酰胺酶是一种重要的抗肿瘤药物。当前制剂的副作用促使研究人员探索新型候选药物。该研究评估了一种从海洋螃蟹锯缘青蟹的解淀粉金色微球菌中提取的新型II型L-天冬酰胺酶。统计优化使产量从115.69 IU/ml提高到281.6 IU/ml。纯化后的酶比活性为173.69 IU/mg蛋白质。米氏常数（Km）、最大反应速度（Vmax）和催化常数（Kcat）分别为6.364 μM、909.09 μM/分钟和222.82 /秒。该酶不与L-谷氨酰胺、D-谷氨酰胺、L-谷氨酸和尿素发生反应，这是临床上所期望的特性。体外胰蛋白酶和血清半衰期分别约为39小时和约27分钟。该研究列出了影响其活性的离子、蛋白质修饰剂和表面活性剂。对功能基因进行的详细计算机分析预测了其物理化学和结构特征。与商业酶的低序列同一性以及预测的无过敏和无毒特征表明其具有临床适用性。分子对接显示对L-天冬酰胺的结合亲和力高于L-谷氨酰胺和尿素。对MCF7、MOLT4和THP-1细胞系的半数抑制浓度（IC50）分别为0.043、0.041和0.05 IU/ml。相差显微镜证实了其细胞毒性。流式细胞术显示在IC50时MOLT-4细胞中有81.4%的凋亡细胞，表明其具有强大的抗癌活性。该酶不影响正常人类成纤维细胞且无溶血作用。结果强调了解淀粉金色微球菌II型L-天冬酰胺酶的治疗潜力，并支持进一步开展体内研究以推进基于L-天冬酰胺酶的癌症治疗。
