Research On Genomics,structure And Antibacterial Effect Of Bacteriocin Produced By Lactobacillus Plantarum

Plantaricin as a biological preservative can replace chemical preservatives to improve food quality and safety because of its non-toxic and high efficiency.Pseudomonas fluorescens(G-,psychrophile)is a major contaminant in dairy products,which often causes heavy losses in dairy products transported even in cold temperature.Most of other bacteriocins cannot inhibit the growth of Pseudomonas fluorescens.However,Plantaricin had a significant inhibitory effect on Pseudomonas fluorescens in previous study.So this paper mainly studied the whole genome sequence of producer strain,the structure of bacteriocin as well as the antibacterial effect of bacteriocin in order to understand the characteristics of bacteriocin and its producer strain.This paper also can provide theoretical support for its application in food preservation.The specific contents and results are as follows :According to the method of whole genome sequencing(WGS),the results showed that Lactobacillus plantarum Q7(Gen Bank: CP019712)contained one complete chromosome(2952198 bp)and four complete plasmids,which were p LP-Q701(12458 bp),p LP-Q702(2968 bp),p LP-Q703(2411 bp)and p LP-Q704(11899 bp).Compared with other 8 typical Lactobacillus plantarum which also can produce bacteriocins in egg NOG,L.plantarum Q7 had a strong genetic adaptability to the transport and metabolism,and it also had the ability to adapt to external stimuli signal.According to blast the CDS,the gene cluster of bacteriocin was found in the p LP-Q704,including the bacteriocin ABC transport protein gene(pLP-Q704_orf00007),the bacteriocin helper protein(p LP-Q704_orf00008),the bacteriocin immunity protein(p LP-Q704_orf00009)and the bacteriocin encoding gene(p LP-Q704_orf000010).When we translated the bacteriocin encoding gene into a protein,it revealed that the precursor peptide contained 62 amino acids,which had a typical N-terminal double-glycine leader consensus and a conserved sequence(-YGNGV-)at their N-terminus.The amino acid sequence of the mature bacteriocin was KYYGNGVTCGKHSCSVDWGKATTCIINNGAMAWATGGHQGNHKC.According to the Prot Param program and Predict Protein program,we analysed the primary structure and secondary structure of bacteriocin.The results showed that the number of amino acids was 44 and the molecular weight was 4628.19.The predicted secondary structure contained helix(6.28%),strand(20.45%)and loop(72.73%).According to the method of circular dichroism(CD),the secondary structure of the plantaricin Q7 in the solution was ?-helix(35.5%),?-sheet(5.8%)and random curl(58.7%),which is different with the predicted secondary structure.According to MALDI-TOF MS,there were no results comparing with bacteria and Lactobacillus plantarum protein databases.According to the method of colorimetry and fluorescent probe,the results showed that Plantaricin Q7 leaded to the leakage of inorganic phosphorus ions in the Pseudomonas fluorescens,released the ATP,dissipated both the membrane potential(??)and the p H gradient(?p H),increased the permeability of the cell membrane,and even formed pores on the surface of the cell membrane.Eventually cell membranes were dissolved and caused the death of Pseudomonas fluorescens.The changes of morphological integrity and the broken cell membranes of Pseudomonas fluorescens were clearly observed by scanning electron microscopy(SEM)and transmission electron microscopy(TEM),leading to the death of Pseudomonas fluorescens.