枯草芽孢杆菌细菌素的分离、表达及稳定性分析

doi:10.11843/j.issn.0366-6964.2024.01.030

Abstract

Abstract: The aim of the present study was to isolate Bacillus subtilis and its bacteriocins, and to analyse the antibacterial activity and stability of the recombinant bacteriocin. The Bacillus spp. producing bacteriocins screened from alpaca feces were isolated by Oxford Cup diffusion method and identified by 16S rRNA. The amino acid sequence of bacteriocins were obtained by (NH₄)₂SO₄ precipitation, chloroform extraction, SDS-PAGE and mass spectrometry analysis. The target bacteriocins were expressed by E. coli expression system, and the antibacterial activity and stability of the recombinant bacteriocins were identified by Oxford Cup diffusion. The results showed that the isolated strain was Bacillus subtilis, and was named as Bacillus subtilis SXAU18, the bacteriocins it produced could inhibit the growth of S.aureus, S. epidermidis, Micrococcus luteus and L. monocytogene. Analysis of the purified bacteriocins showed that they were probably a 10-20 ku DarA protein and two unknown proteins, which belonged to the bacteriocins category. The unknown proteins were named as BLIS SXAU181 and 182, respectively. Recombinant BLIS SXAU181 and 182 were mainly expressed in the soluble form in E. coli expression system, with a single band after purification. Recombinant BLIS SXAU182 had good antibacterial activity, and exhibited high resistance to high temperature, high acidity and alkalini, artificial gastri and intestinal fruid, but recombinant BLIS SXAU181 had no antibacterial activity. In conclusion, a bacteriocin with inhibitory activity against Gram-positive bacteria was isolated from Bacillus subtilis SXAU18 in this study, and the recombinant BLIS SXAU182 showed good antibacterial activity and stability.

Key words: Bacillus subtilis, bacteriocins, isolation, expression, stability

CLC Number:

S829.9

YU Xiuju, ZHANG Min'ai, HU Yanjiao, ZHU Zhiwei, WANG Haidong, YANG Lihua, FAN Kuohai. Isolation, Expression and Stability Analysis of Bacteriocins from Bacillus subtilis[J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(1): 323-333.

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Isolation, Expression and Stability Analysis of Bacteriocins from Bacillus subtilis

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