植物乳杆菌抑菌蛋白的分离鉴定及抑菌特性研究

doi:10.16431/j.cnki.1671-7236.2022.08.042

Abstract

Abstract: 【Objective】 This study was aimed to screen broader-spectrum and effective antibacterial proteins so as to lay the foundation for analyzing antibacterial mechanism of Lactobacillus plantarum thoroughly and exploring new antibacterial agents.【Method】 Lactobacillus plantarum GX20200417-1 was chosen to be the object of this study.Oxford cup method was selected to determine the antimicrobial activity.Proteins were extracted from Lactobacillus plantarum GX20200417-1 by the means of centrifugation at low temperature and ultrafiltration and their antibacterial activities were measured.The extracellular proteins were treated with different temperatures,pH and protease to study the physicochemical properties of antibacterial proteins.Antimicrobial components from Lactobacillus plantarum metabolites were further analyzed by LC-MS/MS.【Result】 Lactobacillus plantarum GX20200417-1 had strong antibacterial effect on Salmonella,Staphylococcus aureus and Escherichia coli.The antimicrobial activity of fermentation was tested at different time which was the strongest at 24 h with the antimicrobial diameter.The proteins extracted from fermentation had the similar antimicrobial activity with the fermentation.Lactobacillus plantarum antibacterial protein exhibited a high stability towards the temperatures. Compared with control group, there was no significant change in antibacterial activity at 20-80 ℃ (P<0.05),the best antibacterial activity at pH 6.0-8.0 and sensitive to protease.By GC-MS/MS analysis,five kinds of proteins were obtained which were not only corrected with bacteriostasis,but also had much more credibility.They were pediocin PA-1,lysin,polyketide synthase,accessory protein and LysM peptidoglycan-binding domain-containing protein and the molecular weight were 5.348,7.348,8.348,6.348 and 19.662 ku respectively.Pediocin PA-1 and lysin achieve bacteriostatic effect mainly by destroying bacterial cell wall and cell membrane.LysM peptidoglycan-binding domain-containing protein could recognize peptidoglycans including N-acetylglucosamine (GlcNAc) residue and upregulate the expression of antibacterial peptides.Accessory protein participated in the synthesis of bacteriocin,polyketide synthase participated in the synthesis of antibiotics,both of which played an indirect bacteriostatic role by participating in the synthesis of antimicrobial substances.【Conclusion】 In this study,five antimicrobial proteins of Lactobacillus plantarum GX20200417-1 were successfully identified,which laid a foundation for protein separation and purification.Its speculated that Lactobacillus plantarum GX20200417-1 could play an antibacterial role by the coordination of antibacterial proteins in metabolites.

Key words: Lactobacillus plantarum; antibacterial proteins; ultrafiltration; LC-MS/MS; antibacterial mechanism

CLC Number:

S816.73

LI Xiaoning, LI Jun, YIN Yangyan, LI Changting, MA Chunxia, TAO Li, GONG Yu, ZHONG Shuhong, BAI Huili, PENG Hao, LIAO Yuying. Isolation,Identification and Antibacterial Property of Antibacterial Proteins from Lactobacillus plantarum[J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(8): 3263-3272.

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Isolation,Identification and Antibacterial Property of Antibacterial Proteins from Lactobacillus plantarum

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