Antibacterial Activity And Action Mode Of Bacteriocin CHQS Produced By Enterococcus Faecalis

Bacteriocin CHQS is the antibacterial peptide produced by Enterococcus faecalis.Previous research revealed that CHQS had the specific N-terminal sequence of class IIa and broad-spectrum bacteriostasis,especially,had significantly strong antibacterial efficacy towards Listeria and showed potential value as a natural food preservative.This thesis aims to further investigate antibacterial activity of the bacteriocin and elucidate its action mode with Listeria ivanovii DSMZ 20750~T as indicator bacteria.Ammonium sulfate precipitation,cation-exchange chromatography and gel filtration chromatography were used to extract the bacteriocin from E.faecalis culture medium.The concentration of purified bacteriocin CHQS was 291.13?g/m L,and its antibacterial potency against L.ivanovii DSMZ 20750~T was 20,971,520 AU/m L.The minimum inhibitory concentration and minimum bactericide concentration of the bacteriocin were 26.74 ng/m L and 107.30 ng/m L,respectively,demonstrating that CHQS had significantly strong inhibitory activity towards L.ivanovii DSMZ 20750~T.The results of time-inhibition assay showed that the bacteriocin could effectively inhibit the growth of bacteria by increasing the lag time and cytolysis.Through glucose energy-enhancement experiment,it was found that L.ivanovii DSMZ 20750~T was more sensitive to the bacteriocin without glucose addition.Based on the experimental results of antibacterial action mode of cell membrane,it was found that the bacteriocin inhibits sensitive bacteria by rendering cell membrane permeable and destructing cell membrane integrity,which occurs simultaneously with the dissipation of the proton motive force.During this process,bacteriocin CHQS induced the leakage of intracellular substances,such as K~+,LDH,ATP,nucleic acid,protein,and eventually led to cell death.Results of gel retardation analysis showed that the cellular targets of CHQS maybe not included bacterial DNA,CHQS could neither bind to nor disconnect bacterial DNA.Through the observation of scanning electron microscopy and transmission electron microscopy,it was found that the cell membrane of bacteriocin-treated indicator bacteria showed extensive pore formation,the cytoplasmic contents were released to the external environment and even existed cellular lysis.This thesis deepened the understanding of antibacterial activity of bacteriocin CHQS produced by E.faecalis,clarified its action mode and provided a theoretical basis for the development and utilization of bacteriocin CHQS as a food preservative.