El Tor型霍乱弧菌分泌性蛋白初步分析

摘要：【目的】利用高通量蛋白质组分析技术，初步探讨El Tor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El Tor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El Tor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。

Analysis of exoproteins of El Tor Vibrio cholerae by 2DE and MALDI-TOF-MS/MS

Abstract: Objective] Our aim is to analyze the exoproteins of the epidemic strain and nonepidemic strain of El Tor Vibrio cholerae. Methods] We separated the exoproteins of two El Tor strains, one epidemic strain N16961 and one nonepidemic strain 92-3, by two dimension electrophoresis. All protein spots were identified by MALDI-TOF (Matrix Assisted Laser Desorption Ionization-Time of Flight) mass spectrometry and database referencing. Results] We identified 49 exoproteins from 206 protein spots in epidemic strain and 42 exoproteins from 236 protein spots in nonepidemic strain. Sixty-eight exoproteins were totally identified in two strains, and the proteins with signal peptides were about 55.88% in total. We classified the exoproteins into 10 subgroups according to their functions. Among them were metabolic enzymes, protein maintenance and folding, protein synthesis and signal transduction proteins, which accounted for 36.76% of all the proteins identified. We identified 11 hypothetical proteins and 2 unknown function proteins with signal peptides at the first time, accounting for 19.12%. Transporter, flagellum components, degradation enzymes, outer membrane proteins and toxin accounted for 14.71%, 11.76%, 10.29%, 5.88%,1.47% separately. Conclusion] We obtained the exoprotein profiles of epidemic strain and nonepidemic strain of El Tor Vibrio cholerae. The reasons of why the flagellum components without signal peptides that are associated closely with virulence could be released out of the bacteria, and why with high hemagglutinin/protease secretion in nonepidemic strains should be analyzed in the future.