Cloning And Expression Of Enterocin A And Enterocin B From Enterococcus Faecalis BZ2 In Koumiss

Enterococcus belongs to lactic acid bacteria,whch is the human and animal intestinal and some important food microbial flora,can produce antibacterial peptide Bacteriocin-Enterocin.Enterococcus can not only inhibit the associated bacteria but also effect on Gram-positive pathogens and food-borne pathogenic microorganisms..Therefore it is expected to be used in animal feed additives and natural food preservative.In order to improve the production of bacteriocin,the prokaryotic expression of the bacteriocin gene that producted from Enterococcus faecalis BZ2 isolated from koumiss and its antibacterial properties were carried out,and it will provide theoretical basis for the further study of antibacterial properties and the development of new microecological preparations.The enteric expression vector of class IIa enterococcin Enterocin A and IIc enterococcin Enterocin B produced by enterococcus BZ2(Enterococcus faecalis BZ2)in koumiss using pEASY-Blunt expression vector was constructed and the expression of ent A and ent B recombinant expression in Escherichia coli and protein purification,and its antibacterial activity was identified.The total DNA was extracted from Enterococcus faecalis BZ2(E.faecalis BZ2)cells,and the entA and entB genes were amplified by PCR.The target fragment was inserted into the prokaryotic expression vector pEasy-blunt-E1.After sequencing analysis,And the gene homology with GeneBank accession number EFU87997 and AB292463 reached 100%.In order to promote the production of ent A and ent B and to facilitate the purification,two recombinant prokaryotic expression plasmids were transformed into E.coli(BL21(DE3))system.Tricine-SDS-PAGE analysis showed that the recombinant protein His-ent A And His-ent B are both highly Appeared with intracellular expression.His-ent A and His-ent B were purified from the supernatant of cell lysate by nickel column affinity chromatography The results showed that His-ent A and His-ent B had inhibitory effects on Listeria monocytogenes.