This study aimed to systematically elaborate on the inhibitory mechanism of metal-binding antimicrobial peptide SIF₄ on the respiratory and energy metabolism of Escherichia coli. By analyzing changes in the metabolic activity, individual and synergistic respiratory inhibition rate, cytoplasmic membrane ion channel ATPase and intracellular ATP level of E. coli after being treated with SIF₄, the effects of SIF₄ on the cell metabolic activity, respiratory metabolic pathway, cytoplasmic membrane ion channel ATPase and intracellular ATP biosynthesis were studied. Results showed that the metabolic activity of E. coli decreased significantly with increasing dose of SIF₄ (P < 0.05), and decreased by 70.41% in the 2 × minimal inhibitory concentration (MIC) group compared with the control group. SIF₄ had good inhibitory effect on the respiration of Escherichia coli, with an inhibition rate of (19.387 ± 0.168)% and (25.222 ± 0.326)% at MIC and 2 × MIC, respectively. The synergistic respiratory inhibition rate of SIF₄ combined with iodoacetic acid was the lowest ((19.982 ± 0.133)%), indicating that SIF₄ could exhibit high antimicrobial activity mainly by inhibiting the glycolysis pathway of E. coli. The activities of cytoplasmic membrane ion channel Na⁺K⁺-ATPase and Ca²⁺Mg²⁺-ATPase decreased after treatment with SIF₄, and this effect was positively correlated with SIF₄ dose and treatment time, but weaker than that of the positive control Triton X-100. As SIF₄ dose and treatment time increased, the intracellular ATP concentration decreased significantly, and after 12 h, the intracellular ATP concentration in the 2 × MIC group was significantly lower than that in the control group but higher than that in the positive control group (P < 0.05). All results confirmed that SIF₄ could exhibit high antimicrobial activity against E. coli by interfering with respiratory metabolism, weakening cytoplasmic membrane ion channel ATPase activity and inhibiting the biosynthesis of intracellular ATP, which can provide theoretical support for the biocontrol of foodborne E. coli.

Due to their high contents of protein, fat, and carbohydrate, foods are extremely susceptible to be contaminated by pathogenic or spoilage microorganisms during production, processing, storage and transportation, which can lead to the production of bio-toxins or food spoilage and loss. Many studies indicate that antimicrobial peptides (AMPs) have good antibacterial activity against pathogenic or spoilage microorganisms. Some AMPs possess bioactivities such as antioxidant and free radical scavenging effects. The efficacy of AMPs as a food preservative can be enhanced by the synergistic action of their antibacterial and antioxidant activity. Hence, AMPs, as novel biological antimicrobial preservatives, have a promising prospect in the field of food preservation. First, this review systematically elucidates the antibacterial mechanism of AMPs from the perspectives of cytoplasmic membrane damage and non-cytoplasmic membrane damage, and then provides a detailed overview of the application of AMPs in the preservation of various types of food. Finally, the problems existing in this field are discussed and possible solutions for these problems are proposed, with the aim of providing theoretical and practical guidance for the application of AMPs in food preservation.

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.

To systematically elucidate how metal-binding antimicrobial peptide SIF₄ exerts its antimicrobial activity by targeting DNA topoisomerase without destroying the cytoplasmic membrane, Escherichia coli was used as a model strain to investigate the binding mode of SIF₄ with its genomic DNA and the impact of SIF₄ on DNA topoisomerase Ⅰ and Ⅱ activities and intracellular nucleic acid biosynthesis. Results showed that SIF₄ could bind to genomic DNA in a manner similar to ethidium bromide (EB) intercalation with strong inhibitory effect on topoisomerase Ⅰ but weak effect on topoisomerase Ⅱ, and catalyzed RNA transcription to exert antimicrobial activity by interfering with the unwinding of negative DNA supercoils and RNA polymerase binding. It was also found that the biosynthesis of intracellular DNA and RNA was inhibited to different degrees after 12 h treatment with SIF₄, which exhibited a good dose-effect relationship. There was no significant difference in the amounts of intracellular DNA and RNA between the 1/2 minimum inhibitory concentration (MIC) group and the control group (P > 0.05), but there was a significant difference between the MIC and 2 MIC groups and the control group (P < 0.05). Our results may provide theoretical support for the application of SIF₄ in the biocontrol of foodborne E. coli.