白酒发酵环境对乳杆菌细菌素群体感应合成调控行为的影响及其机制

项目名称： 白酒发酵环境对乳杆菌细菌素群体感应合成调控行为的影响及其机制

项目编号： No.31671832

项目类型： 面上项目

立项/批准年度： 2017

项目学科： 农业科学

项目作者： 刘国荣

作者单位： 北京工商大学

项目金额： 25万元

中文摘要： 产细菌素弯曲乳杆菌 RX-6自然存在于衡水老白干酒醅中，实验室单独培养试验发现该菌代谢产细菌素表现强烈群体密度感应和环境条件依赖效应。本项目拟基于老白干酒醅发酵过程，分析乳杆菌细菌素curvacin RX-6分泌表达及活性变化规律，确定影响其合成的主要环境胁迫条件参数；通过基因定位和染色体步移技术，明确细菌素合成相关群体感应系统信号肽及双组份信号转导蛋白编码基因；基于不产细菌素培养物模型构建，并结合实时荧光定量PCR、高效液相色谱-电喷雾离子化串联质谱及傅里叶红外光谱技术，研究主要胁迫条件对信号肽分泌表达、分子结构及其诱导活性影响。通过添加组氨酸蛋白激酶抑制剂，分析双组份信号转导蛋白失活前后信号肽诱导效应强度变化，阐释胁迫条件对信号肽与双组份相互作用的影响机制。研究结果对于从分子水平全面揭示乳酸菌细菌素在实际发酵环境中的合成调控行为具有重要科学意义。

中文关键词： 细菌素；群体感应；合成；调控；酒醅发酵环境

英文摘要： Bacteriocin producing Lactobacillus curvatus RX-6 is naturally existed in fermented grains of Hengshui Laobaigan liquor. Previous laboratory studies showed that the synthesis of curvacin RX-6 is regulated by quorum sensing system and is an environmental conditions-sensitive process. In this program, the influence mechanism of liquor fermented grains fermentation environment on quorum sensing synthesis and regulation behavior of curvacin RX-6 is studied. The main fermentation-related environmental stress conditions affecting bacteriocin synthesis are investigated by analyzing the variation of curvacin RX-6 synthesis and activity in different fermentation stages of Laobaigan liquor fermented grains fermentation process. The coding genes of autoinducer signal peptide and two-component signal transduction protein of quorum sensing system is determined by gene targeting and chromosome walking technology. Then the effect of main stress conditions on secretion expression, molecular strucuture and induced activity of the signal peptide are studied based on no-bacteriocin producing cultures model construction, combined with real time fluorescent quantitative PCR technique (RT-qPCR), high performance liquid chromatography-electrospray tandem mass spectrometry (HPLC-ESI-MS/MS) and fourier transform infrared spectrometer (FTIR). To explain the effect of the main stress conditions on the interaction between the signal peptide and two-component, the two-component are inactivated and its effect on the inducer activity of signal peptide is analyzed. The research results are of great scientific significance to comprehensively reveal the synthesis and regulation mechanism of bacteriocins produced by lactic acid bacteria in the actual fermentation environment from the molecular level.

英文关键词： bacteriocin;quorum sensing;synthesis;regulation;fermented grains fermentation environment