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| 绿僵菌几丁质酶的分离纯化及性质 | |
| 引用本文： | 杨革,陈洪章,李佐虎. 绿僵菌几丁质酶的分离纯化及性质[J]. 化工学报, 2005, 56(4): 672-676 |
| 作者姓名： | 杨革 陈洪章 李佐虎 |
| 作者单位： | 中国科学院过程工程研究所生化工程国家重点实验室，北京 100080 |
| 基金项目： | 国家重点基础研究发展计划项目 ( 973 计划)(2004CB719700)，中国科学院知识创新工程重要方向项目(KSCXZ SW 301).~~ |
| 摘 要： | 从自然罹病死亡的金龟子体内分离到一株金龟子绿僵菌（Metarhizium anisopliae）,它在几丁质的诱导下能产生较高活性的几丁质酶.发酵液经硫酸铵盐析、DEAE纤维素柱层析、Phenyl SepharoseTM 6 Fast Flow疏水柱层析等方法，得到电泳纯的几丁质酶.用SDS-PAGE测得该酶相对分子质量为61.5 kD，而经质谱分析为57.14 kD.最适反应温度为55 ℃，最适反应pH值为6.0，酶的等电点pI为4.02，其N末端序列为VIGPAAPL，用硫酸-酚法测得其含糖量为56.2%.水解几丁质的Km为14.5 μmol•L-1.该酶在45 ℃，pH值3.0～9.5较为稳定.Zn2+、Ca2+、Ba2+和Mn2+离子对几丁质酶活性有明显的促进作用，而Hg2+、Co2+和Fe2+离子完全抑制几丁质酶的活性.此酶还可被EDTA所抑制，表明金属离子为其活性所必需.PMSF试剂对几丁质酶的活力影响比较大，丝氨酸可能是酶活力的必需基团. |
| 关 键 词： | 金龟子绿僵菌 几丁质酶 分离纯化 酶学性质 |
| 文章编号： | 0438-1157(2005)04-0672-05 |
| 收稿时间： | 2004-01-15 |
| 修稿时间： | 2004-3-24 |
Purification and properties of chitinase from Metarhizium anisopliae |
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| YANG Ge,CHEN Hongzhang,LI Zuohu. Purification and properties of chitinase from Metarhizium anisopliae[J]. Journal of Chemical Industry and Engineering(China), 2005, 56(4): 672-676 | |
| Authors: | YANG Ge CHEN Hongzhang LI Zuohu |
| Abstract: | A fungium-producing chitinase was isolated from the dead body of Anisopliae. A chitinase was isolated from the culture of Metarhizium anisopliae and purified to electrophoretic homogeneity by the steps of ammonium sulfate precipitation,DEAE-cellulose and hydrophobic interaction column chromatography. Its molecular mass was estimated to be about 61.5 kD by SDS-PAGE,and 57.14 kD by mass spectroscopy.The isoelectric temperature and pH of the enzyme activity were 55 ℃ and 6.0 respectively.The isoelectric point was 4.02.Its N-terminal sequence was VIGPAAPL. The carbohydrate content was 56.2% by the phenol-sulfuric acid method. Michaelis constant of the enzyme was 14.5 μmol•L-1.The enzyme activity was stable under 45 ℃ and in the pH range of 3.0—9.5. The activity was enhanced by Zn2+、Ca2+、Ba2+ and Mn2+,and was strongly inhibited by Hg2+、Co2+ and Fe2+.EDTA also inhibited the activity.Ser was the possible essential residue for enzyme activity. |
| Keywords: | Metarhizium anisopliae chitinase isolation and purification enzyme properities |
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