Abstract:In order to produce bacteriocin in large quantity through prokaryotic expression and investigate its physicochemical properties, this study amplified and recycled the Pediococcus acidilactici R-4 bacteriocin pedA gene. The gene was subsequently ligated into the pMD19-T vector and transferred into E. coli DH5α receptor cells for cloning. The cloned pedA gene was extracted and connected to the expression vector pET-32a(+) to form a recombinant plasmid pET-32a-pedA, which was then transferred into E. coli BL21(DE3) receptor cells. Following induction with isopropyl-β-D- thiogalactoside(IPTG), Pediococcus acidilactici R-4 bacteriocin PA-1 was expressed within E. coli BL21 (DE3) cells. The expressed protein was purified by a Ni-NTA column and its physicochemical properties were determined using Staphylococcus aureus as an indicator. The results demonstrated successful expression and purification of the 26 000 of Pediococcus acidilactici R-4 bacteriocin PA-1 in E. coli BL21 (DE3) cells. The purified Pediococcus acidilactici R-4 bacteriocin PA-1 exhibited antibacterial activity within the ranges of 14.7~15.6 mm, 14.0~16.5 mm, 15.1~15.8 mm, and 14.9 mm, respectively, under the treatment of 40~121 ℃ for 20 min, pH 2 to 12, 0~10 h ultraviolet irradiation, and 2 h catalase. However, the antibacterial activity was lost after treatment with pepsin and trypsin for 2 hours. This indicates that Pediococcus acidilactici R-4 bacteriocin PA-1 has good stability to high temperature, strong acid and alkali, UV radiation, and catalase. However, it could be deactivated by pepsin and trypsin.
