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【目的】利用高通量蛋白质组分析技术,初步探讨ElTor型霍乱弧菌的分泌性蛋白组成,为进一步的功能研究打下基础。【方法】选取ElTor型霍乱弧菌流行株N16961和非流行株92-3,用双相电泳和激光辅助解析-飞行时间串联质谱(Matrix Assisted Laser Desorption Ionization-Time of Flight,MALDI-TOF)技术对培养上清的全部蛋白质组份进行鉴定和分析。【结果】从N16961株培养上清中可检测到206个蛋白点,并鉴定出49种蛋白;从92-3株培养上清中可检测到236个蛋白点,并鉴定出42种蛋白,两株菌共鉴定蛋白68种,其中经预测含有信号肽的蛋白占总数的55.88%(38/68)。按功能不同将所鉴定出的蛋白分为10类,其中代谢酶、蛋白折叠/伴侣蛋白、蛋白合成以及信号传导相关蛋白占全部培养上清蛋白数的36.76%,转运蛋白占14.71%,鞭毛蛋白占11.76%,降解酶占10.29%,外膜蛋白占5.88%,毒素占1.47%,在所鉴定出的蛋白中有11种具有信号肽的假想蛋白和2种功能未知的蛋白为首次被实验证实可出现在霍乱弧菌培养上清中,占19.12%。【结论】初步获得了ElTor型霍乱弧菌流行株和非流行株的分泌性蛋白谱及其组成特征,并提示与霍乱弧菌致病关系密切的鞭毛蛋白在胞外释放机制、红血球凝集素/蛋白酶在非流行株中的高表达特征等值得高度关注。
【Objective】 The purpose of this study was to explore the composition of secreted proteins of ElTor-type Vibrio cholerae by high-throughput proteomic analysis and lay the foundation for further functional studies. 【Method】 ElTor Vibrio cholerae strain N16961 and non-epidemic strain 92-3 were selected and analyzed by two-phase electrophoresis and matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) All the protein components of the culture supernatant were identified and analyzed. 【Result】 206 protein spots were detected in culture supernatant of N16961 strain and 49 proteins were identified. 236 protein spots were detected in 92-3 culture supernatant and 42 proteins were identified. Two Sixty-eight strains of bacteria were identified, of which 55.88% (38/68) were predicted to contain the signal peptide. According to different functions, the identified proteins were divided into 10 categories, of which metabolic enzymes, protein folding / chaperones, protein synthesis and signal transduction related proteins accounted for 36.76% of the total cultured supernatant protein, transport protein accounted for 14.71%, flagellin Accounting for 11.76%, 10.29% of degrading enzymes, 5.88% of outer membrane proteins and 1.47% of toxins. Among the identified proteins, 11 hypothetical proteins with signal peptide and two unknown proteins were first confirmed by experiments Vibrio cholerae can appear in the culture supernatant, accounting for 19.12%. 【Conclusion】 The secreted protein profiles and their components of the strains of epidemic and non-epidemic strains of ElTor-type Vibrio cholera were preliminarily obtained, and the mechanism of extracellular release of flagellin closely related to Vibrio cholerae pathogenesis was suggested. Erythrocyte agglutinin / The high expression of protease in non-epidemic strains is of great concern.