A型魏氏梭菌α毒素氨基端PLC结构分析与生物学活性鉴定

doi:10.3969/j.issn.1004-1524.2017.02.06

浙江农业学报 ›› 2017, Vol. 29 ›› Issue (2): 213-219.DOI: 10.3969/j.issn.1004-1524.2017.02.06

A型魏氏梭菌α毒素氨基端PLC结构分析与生物学活性鉴定

许崇利¹, 许崇波^{2, *}, 宫语晨³

1.吉林化工学院生物与食品工程学院,吉林吉林132022;
2.韶关学院英东生命科学院,粤北生猪生产及疫病防控协同创新发展中心,广东韶关512005;
3.吉林农业大学动物科学技术学院,吉林长春 130118

收稿日期:2016-09-26 出版日期:2017-02-15 发布日期:2017-03-06
通讯作者: 许崇波,E-mail:xcb902@163.com
作者简介:许崇利(1974—),男,黑龙江鸡东人,博士,教授,研究方向为微生物分子生物学。E-mail:xcl902@163.com
基金资助:
吉林省教育厅“十三五”科学研究规划项目(2016138)

Structural analysis and identification of biological activity of alpha-toxin amino-terminal PLC from Clostridium welchii type A

XU Chongli¹, XU Chongbo^{2, *}, GONG Yuchen³

1. College of Biology and Food Engineering, Jilin Institute of Chemical Technology, Jilin 132022, China;
2. North Guangdong Collaborative Innovation and Development Center for Swine Farming and Disease Control, Yingdong College of Life Sciences, Shaoguan University, Shaoguan 512005, China;
3. College of Animal Science, Jilin Agricultural University, Changchun 130118, China

Received:2016-09-26 Online:2017-02-15 Published:2017-03-06

摘要/Abstract

摘要： 利用PCR扩增技术克隆A型魏氏梭菌α毒素氨基端的PLC1-250基因,构建含PLC1-250基因表达质粒的BL21(DE3)(pN-PLC1-250)重组菌株,序列分析和酶切鉴定证实构建的pN-PLC1-250重组质粒含有目的基因且基因序列与阅读框架均正确。SDS-PAGE分析表明,PLC1-250蛋白表达量占菌体总蛋白相对含量的18.76%。利用SOPMA法预测PLC1-250蛋白分子的二级结构,且同源模建了其3D结构,结果表明,PLC1-250蛋白分子的二级结构主要为α螺旋和无规则卷曲,三级结构与α毒素相类似。此外,还对其生物学活性进行了鉴定,可为进一步探索α毒素作用的分子机制,以及其分子结构与生物学功能的关系奠定基础。

关键词: A型魏氏梭菌, α, 毒素PLC基因, 生物学活性, 圆二色光谱

Abstract: Amino-terminal PLC1-250 gene of Clostridium welchii α-toxin was amplified by PCR. The recombinant strain BL21 (DE3)(pN-PLC1-250) containing PLC1-250 gene was constructed. It was shown that the recombinant plasmid pN-PLC1-250 contained PLC1-250 gene with correct sequence and ORF by identification of endonuclease-digesting and sequence analysis. SDS-PAGE analysis showed that the expression level of PLC1-250 proteins were about 18.76% of total cellular proteins. The secondary structure and three-dimensional structure of PLC1-250 proteins were predicted by SOPMA method on EXPASY website. The results showed that the secondary structure of PLC1-250 protein was composed of alpha helices and random coils. The three-dimensional structure of PLC1-250 protein was similar with amino-terminal domain of α-toxin protein. The study of biological activity laid foundation for further research of the molecular mechanism of α-toxin and the relation of its molecular structure and biological function.

Key words: Clostridium welchii type A, alpha-toxin PLC-gene, biological activity, circular dichroism spectra

中图分类号:

S852
Q78

许崇利, 许崇波, 宫语晨. A型魏氏梭菌α毒素氨基端PLC结构分析与生物学活性鉴定[J]. 浙江农业学报, 2017, 29(2): 213-219.

XU Chongli, XU Chongbo, GONG Yuchen. Structural analysis and identification of biological activity of alpha-toxin amino-terminal PLC from Clostridium welchii type A[J]. , 2017, 29(2): 213-219.

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A型魏氏梭菌α毒素氨基端PLC结构分析与生物学活性鉴定

Structural analysis and identification of biological activity of alpha-toxin amino-terminal PLC from Clostridium welchii type A

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