枯草杆菌素Subtillicin L的纯化鉴定及抑菌性质

Purification,identification and antimicrobial characteristics of Subtillicin L produced by Bacillus subtillis DH8043

该文对枯草芽孢杆菌DH8043产生的枯草杆菌素(Subtillicin L)进行了系统分离纯化、结构基因分析预测以及抑菌性质对比研究。将(NH 4)2SO 4沉淀的粗提物经过阴离子交换和层析柱分离纯化,纯化的Subtillicin L通过三甲基甘氨酸-十二烷基硫酸钠-聚丙烯酰胺凝胶电泳进行分子质量分析,得到分子质量约为12.3 kDa的唯一条带。结合全基因组序列的分析及比对,预测Subtillicin L与已知的amylocyclisin氨基酸序列相似度为99.11%,主要区别在于第88个氨基酸上的变化。最后通过比较Subtillicin L和乳酸链球菌素(Nisin)对蜡样芽孢杆菌的抑菌效果,高浓度条件下(4倍的最小半抑制浓度)的Subtillicin L对于蜡样芽胞杆菌属于完全抑制,而相同浓度条件下的Nisin对蜡样芽胞杆菌属于阻止生长。Subtillicin L的纯化和鉴定为抑制蜡样芽孢杆菌提供除Nisin外更多的抑菌选择。

The subtillicin L produced by Bacillus subtillis DH8043 was systematically purified.Meanwhile,the predicted structural gene and antibacterial properties of Subtillicin L were analyzed in the study.The results showed that one single band with molecular weight at approximately 12.3 kDa was found via tricine sodium dodecyl sulfate poly-acrylamide gel electrophoresis(Tricine-SDS-PAGE)after the purification by anion exchange column and Sephadex-G50 chromatography column from the crude extract precipitated by ammonium sulfate.To identify the similarity of Subtillicin L with other bacteriocins,the analysis and blast of the complete genome sequence were used.The predicted similarity between the amino acid sequence of Subtillicin L and the known amylocyclisin was 99.11%.The main difference between them was at the 88 th amino acid by alignment analysis.Finally,antibacterial activity between Subtillicin L and Nisin against Bacillus cereus LMGT2805 was studied.The results showed Subtillicin L can completely inhibit B.cereus LMGT2805 at high concentration,4 times of minimum inhibition concentration(MIC 50),while nisin only prevent the growth of B.cereus LMGT2805 at the same concentration.The antimicrobial results provided more options to inhibit B.cereus besides Nisin.