Purification, Characterization and Anticancer Activity of L-asparaginase Produced by Marine Aspergillus terreus
Purification, Characterization and Anticancer Activity of L-asparaginase Produced by Marine Aspergillus terreus
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DOI：
10.22207/jpam.12.4.19
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发表时间：
2018-12-01
影响因子：
0.8
通讯作者：
Beltagy, Ehab A.
中科院分区：
文献类型：
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作者：
Hassan, Sahar W. M.;Farag, Aida M.;Beltagy, Ehab A.
L-天冬酰胺酶（L-asparaginase，E.C.3.5.1.1）是一种将L-天冬酰胺水解为天冬氨酸和氨的酶，在医药和食品工业中有重要的应用。它是由海洋Aspergillus terreus产生的，用65%硫酸铵沉淀，然后用Sephadex G-100凝胶过滤和DEAE-纤维素离子交换层析纯化，得到11.96倍的纯化。纯化的L-天冬酰胺酶的分子量约为85 kDa，由十二烷基硫酸钠聚丙烯酰胺凝胶电泳测定。L-天冬酰胺酶对L-天冬酰胺有很高的亲和力，Km为31.5mM，Vmax为500 U/ml。纯化的酶的最适pH和温度分别为5.8和40 ℃。L-天冬酰胺酶在pH 4至5.8范围内稳定，在高达70 ℃时稳定。研究了激活剂和抑制剂的作用，前提是CdCl 2、Pb Cl 2-和Hg Cl 2-强烈抑制酶活性，而Na Cl高度增强酶活性。对HCT-116、Hep-G2和MCF-7细胞株的IC_（50）为3.79-12.6 μ g/ml。
L-asparaginase (E.C.3.5.1.1) is an enzyme responsible for hydrolysis of L-asparagine into aspartic acid and ammonia, and has its significant applications in the therapeutics and food technology. It was produced by the marine Aspergillus terreus and precipitated by 65% ammonium sulphate, followed by purification using gel filtration on Sephadex G-100 and DEAE-cellulose ion exchange chromatography, which yielded 11.96 fold purification. The molecular weight of the purified L-asparaginase was approximately 85 kDa, determined by a sodium dodecyl sulphate polyacrylamide gel electrophoresis. L-asparaginase showed high affinity for L-asparagine with a Km of 31.5 mM and Vmax of 500 U/ml. The optimum pH and temperature of the purified enzyme were 5.8 and 40 degrees C, respectively. The L-asparaginase enzyme was stable from pH 4 to 5.8 and stable up to 70 degrees C. The effect of activators and inhibitors was studied providing that CdCl2, Pb Cl-2, and Hg Cl-2 strongly inhibited the enzyme activity, while Na Cl highly enhanced activity. Anticancer activity of the purified L-asparaginase was detected against HCT-116, Hep-G2 and MCF-7 cell lines with IC50 ranged from 3.79-12.6 mu g/ml.