巨噬细胞应激的铜绿假单胞菌磷酸化蛋白质组分析  

Phosphoproteomic analysis of Pseudomonas aeruginosa strains in response to stress induced by mac- rophages

作　　者：潘建义[1] 王长一[1] 叶智鸧[1] 陈冉[1] 赵辅昆[1] 

机构地区：[1]浙江理工大学生命科学学院蛋白质组学与分子酶学研究所,杭州310018

出　　处：《中华微生物学和免疫学杂志》2014年第2期91-95,共5页Chinese Journal of Microbiology and Immunology

基　　金：国家自然科学基金（31200110）;浙江省自然科学基金项目（Y2090396）;浙江省公益性技术研究项目（2011C23067）

摘　　要：目的鉴定巨噬细胞应激的铜绿假单胞菌（Pseudomonas aeruginosa）磷酸化蛋白质组，分析磷酸化修饰在该菌应答应激中的作用。方法采用强阳离子交换层析和固相金属离子亲和层析（SCX-IMAC）富集应激细菌的磷酸化肽段，并利用纳升级液相色谱-质谱联用技术（1lanOLC-MS／MS）进行鉴定和分析其磷酸化蛋白质组。结果在应激细菌中鉴定到14个磷酸化肽段，对应于12个磷酸化蛋白质，其中膜蛋白占50％，提示膜蛋白的磷酸化修饰在这一过程中具有关键作用。蛋白质功能分析显示这些磷酸化蛋白质主要涉及应激应答、铁摄取转运、厌氧呼吸、过氧化氢的应激应答、磷酸化介导的信号转导等生物过程。结论铜绿假单胞菌的蛋白质磷酸化修饰对其转导应激信号及应答巨噬细胞内的缺铁、缺氧和氧化应激等不利生存环境具有关键作用，该结果为深入理解该菌的毒力及致病机制提供新的思路。Objective To investigate the role of protein phosphorylation in Pseudomonas aerugino- sa （P. aeruginosa） strains in response to stress triggered by mouse macrophages. Methods The strong cat- ion exchange-immobilized metal affinity chromatography （SCX-IMAC） was performed to enrich phosphopep- tides. The nanoscale liquid chromatography coupled to tandem mass spectrometry （nano LC-MS/MS） was carried out to identify and analyze phosphoproteome. Results Fourteen phosphopeptides from twelve pro- teins were identified within thirty-one phosphorylation sites on serine, threonine and tyrosine residues. Fifty percent of these phosphorylated proteins were membrane proteins, indicating that their phosphorylation modi- fication was more critical for bacteria in response to the stress. In terms of biological process of Gene Ontolo- gy, these identified proteins were involved in stress response, iron transport, anaerobic respiration, response to hydrogen peroxide and signal transduction by phosphorylation, etc. Conclusion These phosphorylated proteins in P. aeruginosa strains are necessary for signal transduction and their response to harsh environ- ment within the macrophages, such as iron limitation, hypoxia and oxidative stress. This study provides evi- dence for farther investigation on virulence and pathogenesis of P. aeruginosa.

关 键 词：铜绿假单胞菌 巨噬细胞 应激 蛋白质磷酸化 磷酸化蛋白质组 致病机制 

分 类 号：R378[医药卫生—病原生物学]