金属抗菌肽SIF4基于胞内生物大分子靶点的大肠杆菌非膜损伤抑菌机理

doi:10.7506/spkx1002-6630-20221121-230

食品科学 ›› 2023, Vol. 44 ›› Issue (21): 62-68.doi: 10.7506/spkx1002-6630-20221121-230

• 基础研究 • 上一篇

金属抗菌肽SIF4基于胞内生物大分子靶点的大肠杆菌非膜损伤抑菌机理

李玉珍, 肖怀秋, 刘淼, 王琳, 曾梦琪, 赵谋明

（1.湖南化工职业技术学院制药与生物工程学院，湖南株洲 412000；2.华南理工大学食品科学与工程学院，广东广州 510000）

发布日期:2023-12-13
基金资助:
湖南省自然科学基金科教联合基金项目（2022JJ60046）

Metal-Binding Antimicrobial Peptide SIF4 Kills Escherichia coli by Targeting Cytoplasmic Biomacromolecules without Cytoplasmic Membrane Damage: A Mechanistic Study

LI Yuzhen, XIAO Huaiqiu, LIU Miao, WANG Lin, ZENG Mengqi, ZHAO Mouming

(1. School of Pharmaceutical and Bioengineering, Hunan Chemical Vocational Technology College, Zhuzhou 412000, China; 2. School of Food Science and Engineering, South China University of Technology, Guangzhou 510000, China)

Published:2023-12-13

摘要/Abstract

摘要： 为阐明金属抗菌肽SIF4对食源性大肠杆菌基于胞内核酸和蛋白质靶点的非膜损伤抑菌机理，本实验对SIF4处理下胞内核酸生物合成情况进行研究，并对SIF4与溴化乙锭（ethidium bromide，EB）竞争性结合基因组DNA荧光光谱、与基因组DNA互作紫外光谱以及SIF4与基因组DNA的结合方式进行分析，同时对胞内蛋白质生物合成影响进行系统研究。结果表明，SIF4可通过沟槽嵌入与大肠杆菌基因组DNA结合，对核酸生物合成抑制存在剂量效应关系；EB竞争性结合DNA荧光光谱分析结果表明，SIF4可通过嵌插结合和静电吸附竞争结合EB与基因组DNA的结合位点；与基因组DNA互作紫外光谱分析结果表明，SIF4与基因组DNA结合可改变其分子构象，但不断裂基因组DNA双链结构；圆二色光谱分析结果表明，与SIF4结合后，基因组DNA碱基堆积力被削弱，双螺旋结构变得松散，基因组DNA结构由B构型向C型转变；胞内蛋白质生物合成影响分析结果表明，SIF4可显著影响胞内蛋白质生物合成，其抑制效应与SIF4处理时间和处理剂量存在正相关关系。综上，SIF4可通过与基因组DNA进行静电吸附或嵌插结合进入到DNA沟槽，影响DNA复制、RNA转录生物量以及蛋白质翻译，从而实现大肠杆菌非膜损伤抑制。本研究结果可为阐明基于核酸与蛋白质靶点的非膜损伤抑菌机理和食源性大肠杆菌生物防控提供支持。

关键词: 金属抗菌肽；大肠杆菌；胞内生物大分子；非细胞质膜损伤；抑菌机理

Abstract: To explore how metal-binding antimicrobial peptide SIF4 kills foodborne Escherichia coli by targeting nucleic acid and protein in the cytoplasmic membrane without cytoplasmic membrane damage, the effect of SIF4 on intracellular nucleic acid biosynthesis was investigated, and fluorescence spectral analysis of the competition between SIF4 and ethidium bromide (EB) for binding to genomic DNA, ultraviolet (UV) spectral analysis of the interaction between SIF4 and genomic DNA, and the binding mode between SIF4 and genomic DNA were studied. Besides, the effect of SIF4 on intracellular protein biosynthesis was systematically evaluated. Results demonstrated that SIF4 could bind to E. coli genomic DNA through groove insertion and inhibit dose-dependently nucleic acid biosynthesis. Fluorescence spectral analysis showed that SIF4 could compete with EB for binding to genomic DNA through intercalation binding and electrostatic adsorption. UV spectroscopy showed that combination with SIF4 changed the molecular conformation of genomic DNA, but did not break its double strand structure. Circular dichroism (CD) spectroscopy showed that the base stacking force of genomic DNA was weakened, the double helix structure became loose, and the genomic DNA structure was changed from B to C configuration after combination with SIF4. In addition, SIF4 could significantly affect intracellular protein biosynthesis, and its inhibition effect was positively correlated with the treatment time and dose of SIF4. It is believed that SIF4 can enter the DNA groove through electrostatic adsorption or intercalation with genomic DNA, affecting DNA replication, RNA transcriptional biomass and protein translation to produce antimicrobial effect against Escherichia coli without cytoplasmic membrane damage. These results can provide support for the biocontrol of foodborne E. coli.

Key words: metal antimicrobial peptide; Escherichia coli; intracellular biomacromolecules; non-cytoplasmic membrane damage; antimicrobial mechanism

中图分类号:

李玉珍, 肖怀秋, 刘淼, 王琳, 曾梦琪, 赵谋明. 金属抗菌肽SIF4基于胞内生物大分子靶点的大肠杆菌非膜损伤抑菌机理[J]. 食品科学, 2023, 44(21): 62-68.

LI Yuzhen, XIAO Huaiqiu, LIU Miao, WANG Lin, ZENG Mengqi, ZHAO Mouming. Metal-Binding Antimicrobial Peptide SIF4 Kills Escherichia coli by Targeting Cytoplasmic Biomacromolecules without Cytoplasmic Membrane Damage: A Mechanistic Study[J]. FOOD SCIENCE, 2023, 44(21): 62-68.

金属抗菌肽SIF4基于胞内生物大分子靶点的大肠杆菌非膜损伤抑菌机理

Metal-Binding Antimicrobial Peptide SIF4 Kills Escherichia coli by Targeting Cytoplasmic Biomacromolecules without Cytoplasmic Membrane Damage: A Mechanistic Study

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