多黏类芽孢杆菌抗菌肽的分离纯化及特性研究

doi:10.16431/j.cnki.1671-7236.2021.08.016

Abstract

Abstract: The purpose of the study was to sepatate and purify an antimicrobial peptides from the metabolites of Paenibacillus polymyxa BLCC1-0402, and to provide a reference for the preparation of antimicrobial peptides and the detection of their products.Centrifugation, membrane filtration with different molecular weight, ultrafiltration and Superdex peptide 10/300GL gel filtration chromatography were used to separate and purify the fermentation supernatant of Bacillus polymyxa.Bacteriostatic test was carried out on the collected liquid in different periods to compare and evaluate the effect of step chromatography.The standard strain of Escherichia coli O78 was used as the indicator, and the antibacterial activity was detected by the agar diffusion method.Tricine-SDS-PAGE was used to detect the molecular weight.The results showed that, the 3-5 ku component protein samples obtained by 5 and 3 ku roll membrane ultrafiltration had strong antibacterial activity.The 3-5 ku fraction was purified by gel filtration chromatography.The purified antibacterial peptide A3 had the strongest antibacterial activity.The Tricine-SDS-PAGE small molecule peptide electrophoresis showed that the antibacterial peptide A3 had reached the electrophoresis purity and the molecular weight was 4 ku.The antibacterial activity test showed that the antimicrobial peptide had antibacterial effect on the standard strain of Escherichia coli O78.At the same time, antimicrobial peptide A3 showed good heat resistance, and its antibacterial activity could be maintained at about 96% when treated at 90-100 ℃ for 15 min.It had a good acid-base stability, and its antibacterial activity remains above 90% at pH 2.0-9.0.After pepsin treatment, the antibacterial activity of antimicrobial peptide A3 decreased by 20%, and trypsin treatment, the antibacterial activity of antimicrobial peptide A3 decreased by 18%.Protease K had almost no effect on the antibacterial activity of antimicrobial peptide A3.The results showed that the isolated antimicrobial peptide A3 was a new antimicrobial peptide with antibacterial activity against Escherichia coli O78, which had a certain development potential, and provided a certain reference for the further study of antimicrobial peptide structure analysis and physicochemical properties analysis.

Key words: Paenibacillus polymyxa; antibacterial peptide; extraction; separation and purification; stability

CLC Number:

S816.7

YU Jiamin, ZHAO Qian, ZHANG Zhiyan, XU Haiyan, GU Wei. Separation,Purification and Characteristics Analysis of Antibacterial Peptides from Paenibacillus polymyxa[J]. China Animal Husbandry and Veterinary Medicine, 2021, 48(8): 2830-2837.

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Separation,Purification and Characteristics Analysis of Antibacterial Peptides from Paenibacillus polymyxa

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