食品与发酵工业 >

2022 , Vol. 48 >Issue 17: 71 - 78

DOI: https://doi.org/10.13995/j.cnki.11-1802/ts.030314

研究报告

凝结芽孢杆菌CGMCC 9951新型抗菌肽的挖掘、表达及活性测定

  • 张婧 ,
  • 吴影 ,
  • 古绍彬 ,
  • 马金亮 ,
  • 田萍萍 ,
  • 赵丽娜 ,
  • 李欣 ,
  • 张杰
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  • 1(河南科技大学 食品与生物工程学院,河南 洛阳,471023)
    2(河南省食品微生物工程技术研究中心,河南 洛阳,471023)
第一作者:硕士研究生(古绍彬教授为通信作者,E-mail:shaobingu@haust.edu.cn)

收稿日期: 2021-12-03

  修回日期: 2021-12-29

  网络出版日期: 2022-10-01

基金资助

河南省科技攻关项目(202102110296,212102110326)

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Genome mining, expression and activity determination of a novel antimicrobial peptide from Bacillus coagulans CGMCC 9951

  • ZHANG Jing ,
  • WU Ying ,
  • GU Shaobin ,
  • MA Jinliang ,
  • TIAN Pingping ,
  • ZHAO Lina ,
  • LI Xin ,
  • ZHANG Jie
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  • 1(Henan University of Science and Technology, Luoyang 471023, China)
    2(Henan Engineering Research Center of Food Microbiology, Luoyang 471023, China)

Received date: 2021-12-03

  Revised date: 2021-12-29

  Online published: 2022-10-01

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摘要

抗菌肽是一类能够抑制食源性致病菌生长的天然生物防腐剂。为获得凝结芽孢杆菌(Bacillus coagulans)CGMCC 9951抗菌肽,利用全基因组测序和生物信息学分析,对抗菌肽合成基因簇进行挖掘,并进行表达和活性验证。结果表明,应用antiSMASH软件共挖掘出2个抗菌肽,与NCBI数据库比对后,发现Region 3与Amylocyclicin相似度较高,后续选择Region 3进行表达。通过Expasy等软件对Region 3抗菌肽结构进行分析,发现成熟Region 3抗菌肽由64个氨基酸残基组成,分子质量为6 371.57 Da,呈带正电的疏水性蛋白;Region 3同源比对结果为环状抗菌肽,三级结构主要为α螺旋结构。为研究该肽的抗菌活性,成功在大肠杆菌中构建了含谷胱甘肽巯基转移酶标签的融合蛋白,并进行质谱验证。以单核细胞增生李斯特氏菌为指示菌,抑菌圈直径为(15.2±1.4) mm,证实表达的融合蛋白具有抗菌活性。融合蛋白的成功表达为凝结芽孢杆菌CGMCC 9951抗菌肽的进一步研究和应用奠定基础。

关键词： 凝结芽孢杆菌CGMCC 9951; 全基因组测序; 环肽; 表达; 抗菌活性

本文引用格式

张婧 , 吴影 , 古绍彬 , 马金亮 , 田萍萍 , 赵丽娜 , 李欣 , 张杰 . 凝结芽孢杆菌CGMCC 9951新型抗菌肽的挖掘、表达及活性测定[J]. 食品与发酵工业, 2022 , 48(17) : 71 -78 . DOI: 10.13995/j.cnki.11-1802/ts.030314

Abstract

Antimicrobial peptides are natural biological preservatives that inhibit the growth of food-borne pathogens. In order to obtain antimicrobial peptide from Bacillus coagulans CGMCC 9951, whole genome sequencing and bioinformatics analysis were used to mine the antimicrobial peptide synthesis gene cluster, and their expression and activity were verified. In order to study the antibacterial activity of the peptide, the fusion protein containing glutathione S-transferase (GST) tag was successfully constructed in Escherichia coli and verified by mass spectrometry. The results showed that two antimicrobial peptides were excavated by antiSMASH software. After comparison with National Center for Biotechnology Information (NCBI) database, Region 3 was found to be greatly similar with Amylocyclicin, and was selected for expression. It was found that mature Region 3 was composed of 64 amino acid residues with a molecular weight of 6 371.57 Da, which was positively hydrophobic protein. Homology of Region 3 showed cyclic antimicrobial peptide, and the tertiary structure was mainly α helix structure. Using Listeria monocytogenes as an indicator bacterium, the inhibition zone diameter was (15.2±1.4) mm, which confirmed that the expressed fusion protein had antibacterial activity. The successful expression of the fusion protein lays a foundation for the further study of B. coagulans CGMCC 9951 antimicrobial peptide.

Key words： Bacillus coagulans CGMCC 9951; whole genome sequencing; cyclic peptide; expression; antimicrobial activity

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