苏云金芽孢杆菌HZP7完整和截短几丁质酶在大肠杆菌中表达后的纯化及部分特性分析

Purification and partial characterization of intact and truncated chitinase from Bacillus thuringiensis HZP7 expressed in Escherichia coli.

作者信息

Sha Li, Shao Ensi, Guan Xiong, Huang Zhipeng

机构信息

Key Laboratory of Biopesticide and Chemical Biology, Ministry of Education, Fujian Agriculture and Forestry University, Fuzhou, 350002, People's Republic of China.

School of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou, 350002, People's Republic of China.

出版信息

Biotechnol Lett. 2016 Feb;38(2):279-84. doi: 10.1007/s10529-015-1970-6. Epub 2015 Oct 13.

DOI:10.1007/s10529-015-1970-6

PMID:26463368

Abstract

OBJECTIVE

To ascertain the effect of chitin-binding domain (ChBD) and fibronectin type III domain (FN3) on the characterization of the intact chitinase from Bacillus thuringiensis.

RESULTS

An intact chitinase gene (chi74) from B. thuringiensis HZP7 and its truncated genes (chi54, chi63 and chi66) were expressed in Escherichia coli BL21. The expression products were analyzed after purification. All chitinases were active from pH 4-7.5 and from 20 to 80 °C with identical optimal: pH 5.5 and 60 °C. The activity of colloid chitin degradation for Chi74 was the highest, followed by Chi66, Chi63 and Chi54. Ag(+) reduced the activity of Chi74, Chi54, Chi63 and Chi66, but Mg(2+) enhanced them. The effect of Ag(+) and Mg(2+) was more significant on the activity of Chi54 than on the activities of Chi63, Chi66 and Chi74.

CONCLUSION

ChBDChi74 and FN3Chi74 domains play a role in exerting enzymatic activity and can improve the stability of chitinase.

摘要

目的

确定几丁质结合结构域（ChBD）和Ⅲ型纤连蛋白结构域（FN3）对苏云金芽孢杆菌完整几丁质酶特性的影响。

结果

苏云金芽孢杆菌HZP7的完整几丁质酶基因（chi74）及其截短基因（chi54、chi63和chi66）在大肠杆菌BL21中表达。纯化后对表达产物进行分析。所有几丁质酶在pH 4至7.5以及20至80℃范围内均有活性，最适pH为5.5，最适温度为60℃。Chi74对胶体几丁质的降解活性最高，其次是Chi66、Chi63和Chi54。Ag(+)降低了Chi74、Chi54、Chi63和Chi66的活性，但Mg(2+)增强了它们的活性。Ag(+)和Mg(2+)对Chi54活性的影响比对Chi63、Chi66和Chi74活性的影响更显著。