| 南极磷虾体内胰蛋白酶的纯化及性质研究 |
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| 引用本文： | 田鑫,汪之和,施文正,李燕.南极磷虾体内胰蛋白酶的纯化及性质研究[J].上海海洋大学学报,2014,23(5):741-747. |
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| 作者姓名： | 田鑫 汪之和 施文正 李燕 |
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| 作者单位： | 上海海洋大学,上海海洋大学 |
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| 基金项目： | 国家高技术研究发展计划（863计划） |
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| 摘 要： | 以南极磷虾(Euphausia superb)为研究对象,通过硫酸铵分级沉淀、Phenyl-Sepharose疏水层析、DEAE-Sepharose FF离子交换层析等方法,从南极磷虾体内分离纯化出胰蛋白酶。其纯化倍数为5.44倍,比活力为38.3 U/mg,得率为26%。SDS-PAGE电泳结果显示,该酶的分子质量为28 ku。蛋白酶最适温度为37℃、最适pH为7.5,Mg2+、Ca2+、Mn2+对南极磷虾蛋白酶具有激活性,Zn2+、Cu2+、Fe3+具有酶活抑制性,其中Cu2+的抑制性最强。酶的动力学实验结果表明,以BApNA为底物测得Km为0.073 mmol/L,Vmax为1.44×10-2mmol/L·s,kcat为0.6S-1,kcat/Km为8.22×103,PMSF作为蛋白酶抑制剂,对南极磷虾蛋白酶作用机制为不可逆抑制。
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| 关 键 词： | 南极磷虾 蛋白酶 纯化 酶学性质 |
| 收稿时间： | 2014/3/14 0:00:00 |
| 修稿时间： | 5/3/2014 12:00:00 AM |
Purification and characterization of serine proteinase from Euphausia superba |
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| TIAN Xin,WANG Zhi-he,SHI Wen-zheng and LI Yan.Purification and characterization of serine proteinase from Euphausia superba[J].Journal of Shanghai Ocean University,2014,23(5):741-747. |
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| Authors: | TIAN Xin WANG Zhi-he SHI Wen-zheng and LI Yan |
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| Institution: | Shanghai ocean university,Shanghai ocean university |
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| Abstract: | A serine protease from Euphausia superba was purified by a series of procedures, including ammonium sulfate precipitation, column chromatographies on DEAE-Sepharose and Phenyl-Sepharose. The purification multiple of the protease was 5.44 times, and the yield of the protease was 26%, with specific activity of 38.3 U/mg. As shown in the result of SDS-PAGE electrophoresis, the molecular weight of this protease is 28 ku. The optimum temperature of protease was 37 ℃ and the most suitable pH was 7.5. Mg2＋ , Ca2＋ and Mn2＋ were activated to protease from Euphausia superba. However, Zn2＋, Cu2＋ and Fe3＋ were inhibited to the enzyme activity, and the inhibition ability of Cu2＋ was the strongest. The enzyme kinetics experiments were performed by using BApNA as substrate. The results showed that the Km value was 0. 073 mmol/L, Vmax value was 1.44 × 10^-2 mmol/L · s,kcat value was 0.6 S-1 and k cat/Km value was 8.22 × 10^3. PMSF was a protease inhibitor, and its mechanism of action was irreversible inhibition. |
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| Keywords: | Euphausia superba Protease Purification Enzymatic properties |
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