| 深海高温淀粉普鲁兰酶异源表达及酶活分析 |
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| 引用本文： | 焦豫良,王淑军,吕明生,房耀维,刘姝.深海高温淀粉普鲁兰酶异源表达及酶活分析[J].海洋学报,2011,33(5):133-138. |
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| 作者姓名： | 焦豫良 王淑军 吕明生 房耀维 刘姝 |
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| 作者单位： | 淮海工学院 海洋学院,江苏 连云港 222005 |
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| 基金项目： | 国家自然科学基金项目(40746030);江苏省高校自然科学研究重大项目(09KJA170001)。 |
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| 摘 要： | 深海热液口厌氧古菌Thermococcus siculi HJ21中的高温淀粉普鲁兰酶进行分子进化树系分析,并在大肠杆菌中通过pMal-c2x载体表达并纯化其N端催化结构域.通过融合表达,在N端催化结构域的N端融合有麦芽糖结合蛋白MalE.对该融合蛋白的α-淀粉酶和普鲁兰酶活性进行了实验分析.融合蛋白的两种酶活的最适温...
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| 关 键 词： | 高温淀粉普鲁兰酶 异源表达 酶活分析 |
| 收稿时间： | 2010/12/30 0:00:00 |
Heterologous expression of a deep-sea thermostable amylopullulanase and enzymatic activity analysis of the fusion protein |
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| JIAO Yu-liang,WANG Shu-jun,LU Ming-sheng,FANG Yao-wei and LIU Shu.Heterologous expression of a deep-sea thermostable amylopullulanase and enzymatic activity analysis of the fusion protein[J].Acta Oceanologica Sinica (in Chinese),2011,33(5):133-138. |
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| Authors: | JIAO Yu-liang WANG Shu-jun LU Ming-sheng FANG Yao-wei and LIU Shu |
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| Institution: | College of Food Science and Technology, Huaihai Institute of Technology, Lianyungang 222005, China |
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| Abstract: | Molecular phylogenetic tree analysis on a thermostable amypopullulanase in an Archaeaon strain Thermococcus siculi HJ21 isolated from a deep-sea hydrothermal vents was performed based on experimental enzymatic analysis and amino acid squences of amylopullulanases deposited in GenBank. The N-terminal catalytic region of the amylopullulanase was heterologously expressed in E. coli through pMal-c2x expression system, resulting a fusion protein in which there is a maltose binding protein fused to the N-terminus of the N-terminal catalytic region of the amylopullulanase. Alpha-amylase and pullulanase activities of the fusion protein were experimentally analyzed. The optimal temperatures of the two activities were both at 100 ℃. The optimal pHs of amylase and pullulanase activities were at 5 and 6 respectively. The specific activities of the amylase and pullulanase activities were 6.5 and 11.5 U/mg respectively. The results showed that α-amylase activity was lower than pullulanase activity and the C-terminal region of the thermostable amylopullulanase was non-necessary for the enzymatic activities. The thermostable amylopullulanase fusion protein obtained in this study could be further used for combination with thermostable amylases in the sugar industry. |
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| Keywords: | thermostable amylopullulanase heterologous expression enzymatic analysis |
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