三种植物源增殖性底物对嗜酸乳杆菌发酵及蛋白表达的影响

Fermentation and Influence of Protein Expression of Three Plant-Based Proliferative Substrates for Lactobacillus acidophilus CICC6005

为探究植物源增殖性底物对嗜酸乳杆菌(Lactobacillus acidophilus CICC6005)生物量的影响,并在此基础上重点解析发酵菌体蛋白质表达水平上的变化,本研究通过采用正交优化方法,在TJA培养基中添加各种增殖性底物确定其生物量的水平,利用SDS-PAGE凝胶电泳技术对胞内胞外蛋白质进行纯化鉴定。结果显示,三种植物源增殖性底物最优浓度分别为番茄汁5%、土豆汁3%、苹果汁2%,发酵液活菌数达2.31×10^(10) CFU/mL;由R值分析得出,影响L.acidophilus CICC6005菌株生长的各因素的主次顺序为:番茄汁、土豆汁和苹果汁。发酵液中表达全蛋白质的图谱显示,L.acidophilus CICC6005菌株在该营养条件下表达蛋白的数量有明显的增加,并根据标准曲线得出各峰的蛋白分子量大小,即胞内蛋白的分子量为59、17 ku;胞外蛋白的分子量为66、35 ku,其表达量分别为2.144、0.744、2.394、4.644 mg/mL。综上,在TJA基础培养基上,适宜浓度的植物源增殖性底物对L.acidophilus CICC6005菌株的增殖以及蛋白质的表达水平均起到了一定的促进作用,同时纯化鉴定出该菌表达的两种主要蛋白质,对进一步其益生功能具有科学和实践意义。

In order to explore the effects of botanical proliferative substrates on the biomass of Lactobacillus acidophilus CICC6005,and to analyze the changes of protein expression level in fermented cells,orthogonal optimization was applied to determine the biomass of TJA medium by adding various proliferative substrates in this study.The intracellular and extracellular proteins were purified and identified by SDS-PAGE gel electrophoresis.The results showed that,the optimum concentration of proliferative substrates from three plant sources was 5%tomato juice,3%potato juice,2%apple juice,and the number of viable bacteria in fermentation broth was 2.31×1010 CFU/mL.The order of factors affecting the growth of L.acidophilus CICC6005 strain was tomato juice,potato juice and apple juice.The maps of all proteins expressed in fermentation broth showed that L.acidophilus CICC6005 strain had a significant increase in the number of proteins expressed under the nutritional conditions.According to the standard curve,the molecular weight of each peak was 59 and 17 ku,and the molecular weight of extracellular protein was 66 and 35 ku,with the expression quantities of 2.144,0.744,2.394 and 4.644 mg/mL,respectively.The maps of expression of all protein in the fermentation broth showed that the number of protein expressed in L.acidophilus CICC6005 strain increased obviously under the nutritional conditions.According to the standard curve,the molecular weight of each peak was 59 and 17 ku,and the molecular weight of extracellular protein was 66 and 35 ku,the expression quantities were 2.144,0.744,2.394 and 4.644 mg/mL respectively.In conclusion,on the TJA basic medium,the suitable concentration of plant source proliferative substrates played a certain role in promoting the proliferation of L.acidophilus CICC6005 strain and protein expression level.At the same time,two major proteins expressed by the strain were purified and identified,which was of scientific and practical significance for further analysis and identification of its probiotic function.