来源于青霉菌聚半乳糖醛酸酶的筛选及其在大肠杆菌中的异源表达

doi:10.3969/j.issn.2095-2341.2011.06.08

生物技术进展 ›› 2011, Vol. 1 ›› Issue (6): 426-431.DOI: 10.3969/j.issn.2095-2341.2011.06.08

来源于青霉菌聚半乳糖醛酸酶的筛选及其在大肠杆菌中的异源表达

曹威1,2,周利伟1,张伟2,张宇宏1*

1.中国农业科学院生物技术研究所, 北京 100081;
2.河北农业大学食品科技学院, 河北保定 071001

收稿日期:2011-08-25 出版日期:2011-12-25 发布日期:2011-10-15
通讯作者: 张宇宏,博士,助理研究员,主要从事酶工程研究。E-mail:zhangyuhong@caas.net.cn
作者简介:曹威,硕士研究生,从事微生物酶基因工程的研究。
基金资助:
中国农业科学院生物技术研究所基本科研业务费专项资金(2008-08)资助。

Screen and Expression of the Penicillium sp. FJ2 Polygalacturonase gene in Escherichia coli

CAO Wei, ZHOU Li-wei, ZHANG Wei, ZHANG Yu-hong

1.Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, China;
2.College of Food Science and Technology, Agricultural University of Hebei, Hebei Baoding 071001, China

Received:2011-08-25 Online:2011-12-25 Published:2011-10-15

摘要/Abstract

摘要： 聚半乳糖醛酸酶属于果胶水解酶类,在食品、饲料、纺织和造纸等工业生产中应用广泛。本研究筛选获得一株能分解果胶的青霉菌Penicillium sp. FJ2,使用简并引物PCR和TAIL-PCR方法从该菌中克隆得到一个聚半乳糖醛酸酶基因pgp1。pgp1基因全长1 225 bp,包含2个内含子,其cDNA全长1 104 bp,编码367个氨基酸和一个终止密码子。将pgp1基因连接pET-30a(+)载体并转化大肠杆菌BL21(DE3),重组蛋白以包涵体形式获得表达。通过尿素溶解和梯度稀释方法对重组蛋白PGP1进行了重折叠复性试验,复性后的PGP1聚半乳糖醛酸酶活力达到12.9 U/mL,比活力为583 U/mg。

关键词: 聚半乳糖醛酸酶, 果胶酶, 青霉菌, 包涵体, 复性

Abstract: Polygalacturonases are a class of pectinase that can hydrolyze pectin, and they are widely used in food, feed, textile and paper industry. A Penicillium sp. FJ2, which produced polygalacturonase, has been screened in this research. In addition, a polygalacturonase gene pgp1 has been cloned using degenerated PCR and TAIL-PCR. Length of the polygalacturonase pgp1 gene and its cDNA are 1 225 bp and 1 104 bp, respectively. The cDNA of pgp1 encodes 367 amino acids and a termination codon. Pgp1 was cloned into pET-30a(+) vector and expressed in Escherichia coli BL21(DE3). The recombinant PGP1 was expressed as inclusion bodies without activity towards polygalacturonic acid. The renaturation of the PGP1 was through with urea resolution and gradient dilution. Polygalacturonase activity of the renatured PGP1 was detected as 12.9 U/mL, and with specific activity of 5.83 U/mg.

Key words: polygalacturonase, pectinase, Penicillium, inclusion body, renaturation

曹威,周利伟,张伟,张宇宏. 来源于青霉菌聚半乳糖醛酸酶的筛选及其在大肠杆菌中的异源表达[J]. 生物技术进展, 2011, 1(6): 426-431.

CAO Wei1,2, ZHOU Li-wei1, ZHANG Wei2, ZHANG Yu-hong1*. Screen and Expression of the Penicillium sp. FJ2 Polygalacturonase gene in Escherichia coli[J]. journal1, 2011, 1(6): 426-431.

[1]	李力群, 孙志康, 郝捷, 季嫱, 李选文, 吴晗, 吴娜, 郑超, 杨婧. 果胶酶生产及工业应用进展[J]. 生物技术进展, 2022, 12(4): 549-558.
[2]	吴斌,郭芹,王刚霞,陈维信,李雪萍. 一氧化氮处理对香蕉多聚半乳糖醛酸酶及MaPGs基因表达的影响[J]. 生物技术进展, 2014, 4(1): 30-35.
[3]	郑科,刘正初,段盛文,成莉凤,郑霞,冯湘沅. 果胶酶在麻类脱胶中的应用及其作用机理[J]. 生物技术进展, 2012, 2(6): 404-410.

来源于青霉菌聚半乳糖醛酸酶的筛选及其在大肠杆菌中的异源表达

Screen and Expression of the Penicillium sp. FJ2 Polygalacturonase gene in Escherichia coli

PDF

可视化

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 3

编辑推荐

Metrics

本文评价