耐铅屎肠球菌JT1与非耐铅屎肠球菌M差异蛋白解析研究

Identification of differentially expressed proteins of Pb-tolerance Enterococcus faecium JT1 and Pb-intolerance Enterococcus faecium M

采用蛋白质组学技术研究具有耐铅性的屎肠球菌JT1菌株,以期获得其与非耐铅屎肠球菌M菌株的差异蛋白质序列。首先利用二维凝胶电泳建立耐铅乳酸菌的蛋白质图谱,对比上述2种菌株的蛋白质图谱,找出显著表达的差异蛋白点,再进行质谱鉴定,继而对相关的差异蛋白进行蛋白质的生物信息学分析。研究结果表明:得到21个差异点对应的19种差异蛋白主要分为6类功能蛋白,分别是细胞胁迫响应蛋白、糖类代谢蛋白、小分子代谢蛋白、核酸代谢蛋白、细胞增殖蛋白、蛋白质代谢蛋白;其中,半胱氨酸合成酶A是与小分子代谢相关的蛋白,在有氧生长中能增加半胱氨酸和胱硫醚-β-合成酶的酶活,半胱氨酸则与Pb^(2+)等金属离子可形成不溶性的硫醇盐,进而使菌体能够抵抗氧化胁迫,对Pb^(2+)造成的活性氧增加引起的细胞结构损伤有着良好的缓解作用,从而提高菌株耐铅性;此外半胱氨酸合成酶还与谷胱甘肽的合成相关,谷胱甘肽可以提升菌体抗酸胁迫和抗毒胁迫的能力,增加JT1菌株对Pb^(2+)的抗渗透力,以及作为直接作用细胞内金属离子的螯合剂。

Using proteomics technology, this study revealed the differentially expressed proteins of Pb- tolerance Enterococcus faecium JT1 and Pb-intolerance Enterococcus faecium M. We established two- dimensional gel electrophoresis profiles of proteins in lactobacillus and compared the protein profiles of the two strains above. Furthermore, mass spectrometry was used to identify differentially expressed proteins and related bioinformatics analysis. Our study revealed 19 differentially expressed proteins, which divided into six categories： cell stress response protein, carbohydrate metabolism protein, small molecule metabolism protein, nucleic acid metabolism protein, cell proliferation and protein metabolism protein. Cysteine synthase A, involving in small molecule metabolism, could increase the activities of homocysteine and cystathionine beta synthase. Cysteine and Pb2＋ could effectively alleviated reactive oxygen induced cell structure damage by forming insoluble thiolate and increasing bacterial oxidation resistance, thus improving Pb-tolerance of Enterococcus faecium. In addition, cysteine synthase was also associated with glutathione synthesis, which could enhance anti-acid and anti-toxic abilities of strains, increasing the anti-permeability of JT1 strain to Pb2＋, as well as acting as an intracellular metal ions chelating agent.