Non-membrane mechanisms of antimicrobial peptide P7 against Escherichia coli

Abstract:

[Objective] The molecular mechanism of antimicrobial peptide P7 against Escherichia coli was studied.[Methods] The binding mode between P7 and DNA was analyzed through fluorescence spectroscopy of P7 binding with E. coli genome DNA. The effects of P7 on E. coli cell cycle were determined through flow cytometry. Magnetic beads coupled with peptide were ussed to enrich peptide DNA-binding fragments, and PCR methods were used to analyze specific DNA to which P7 bound with. The influence of P7 on the gene expression levels of DNA replication and SOS damage and repair was analyzed through quantitative real-time PCR. The effects of P7 on E. coli DNA and RNA synthesis were analyzed according to the fluorescence spectra of nuclear stains.[Results] P7 intercalated into the base pairs of E. coli genomic DNA and then formed peptide-DNA complexes. As a result, the fluorescence intensity of the EB-DNA complex decreased. P7 could significantly increase the number of E. coli cells in phase S. The effect of P7 on normal E. coli cell cycle could significantly inhibit the DNA replication of E. coli. The binding of P7 with rnhA down-regulated the gene expression level by 2.24 times. The gene expression levels of ssb, dnaG, ligB, and rnhA that participated in E. coli DNA replication significantly decreased, and the gene expression levels of recA and recN in DNA damage and repair were significantly up-regulated under the effect of P7. P7 reduced E. coli DNA and RNA synthesis.[Conclusion] P7 also bound with rnhA. This binding resulted in DNA damage and inhibition of DNA replication of E. coli. P7 down-regulated the gene expression level of DNA replication, and the gene expression levels of DNA damage and repair were significantly up regulated. P7 reduced DNA and RNA synthesis of E. coli.