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贝莱斯芽孢杆菌(Bacillus velezensis) MCCC 1A15695抑菌蛋白分离纯化及应用研究
付欢,李亚婷,肖贵谦,彭磊磊,夏金梅,邵宗泽,曹军伟,赖其良
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(上海海洋大学海洋科学学院,上海 201306;自然资源部第三海洋研究所、自然资源部海洋生物遗传资源重点实验室,福建 厦门 361005)
摘要:
对虾“玻璃苗”病的暴发,会导致对虾幼苗大批死亡,给水产养殖带来巨大损失。利用本实验室最近鉴定的“玻璃苗”致病菌副溶血弧菌(Vibrio parahaemolyticus)vp-HL,从海洋来源芽孢杆菌中筛选拮抗菌,获得一株能够高效抑制vp-HL菌株的贝莱斯芽孢杆菌(Bacillus velezensis)MCCC 1A15695。经分离鉴定与活性测定,其有效抑菌物质为蛋白类,该蛋白抑菌谱较广,对包括多种弧菌病原在内的17种病原微生物有抑制作用。经Q Seharose Fast Flow阴离子柱层析、聚丙烯酰胺凝胶电泳(SDS-PAGE)及液相色谱质谱联用技术(LC-MS)分离纯化鉴定,确定活性蛋白组分中的有效抑菌物质为枯草杆菌素(subtilisin)、未知蛋白(hypothetical protein)及α-淀粉酶(Alpha-amylase)。蛋白类物质具有高生物安全性,本研究为后续益生菌的开发利用提供了理论基础。
关键词:  海洋生物学  贝莱斯芽孢杆菌  抗菌蛋白  抑菌谱  弧菌  玻璃苗
DOI:10.3969/J.ISSN.2095-4972.20230531002
基金项目:科技部2021年科技基础资源调查专项(2021FY100903)
Isolation and purification of bacteriostatic protein from Bacillus velezensisMCCC 1A15695 and its application
FU Huan,LI Yating,XIAO Guiqian,PENG Leilei,XIA Jinmei,SHAO Zongze,CAO Junwei,LAI Qiliang
(College of Marine Sciences, Shanghai Ocean University, Shanghai 201306, China;Key Laboratory of Marine Genetic Resources, Third Institute of Oceanography, MNR, Xiamen 361005, China)
Abstract:
The outbreak of shrimp “glass post-larval disease” has led to a large number of diseases of shrimp larvae, bringing huge losses to aquaculture. Using the recently identified Vibrio parahaemolyticus vp-HL of the “glass post-larval disease” pathogen Vibrio parahaemolyticus, the antagonistic antibacterial screening of Bacillus of marine origin was carried out to obtain a strain of Bacillus velezensis MCCC 1A15695 that can effectively inhibit the strains MCCC 1A15695. After isolation and activity determination, the effective bacteriostatic substances were proteins, with a wide inhibitory spectrum, and had inhibitory effects on 17 pathogenic microorganisms, including a variety of Vibrio pathogens. After Q Seharose Fast Flow anion column chromatography, polyacrylamide gel electrophoresis (SDS-PAGE) and liquid chromatography-mass spectrometry (LC-MS), the effective bacteriostatic substances in the active protein components were subtilisin, hypothetical protein and Alpha-amylase. In view of the high biosafety of proteins, the results provide a theoretical basis for the subsequent development and utilization of probiotics.
Key words:  marine biology  Bacillus velezensis  antimicrobial protein  antibacterial spectrum  Vibrio  glass post larval disease

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