利用ΔSRP抑制子挖掘蛋白转运新途径的基础研究

Protein synthesis, folding, targeting and degradation are hot felids in cell biology. The signal recognition particle (SRP) and protein chaperons are universal and conserved cellular machineries for protein targeting to the membrane. In this study, we are not restricted to the research of conserved SRP, Sec and Tat protein targeting pathway, but will focus on discovering the new protein targeting pathway. Temperature sensitive vector and mutation methods will be used to screen ΔSRP suppressor containing new protein targeting pathway. Genome sequencing and quantitative proteomics approaches will be used to identify gene mutations and proteingram differences. Phenotype restoration in vivo and protein interaction in vitro experiments will unveil the new factors and mechanism in the new targeting pathway. Bioinformatics tools will be used to discover the new targeting pathway in other bacteria in nature. This work will provide natural compensatory protein targeting pathways and help us in depth understand the protein targeting pathway. The new targeting pathway discovering will provide a new strategy for protein express system in bacteria and will also provide a basis for drug design to pathogenic bacteria.

蛋白质的合成、折叠、转运和降解是细胞生物学研究的重点。信号识别因子（Signal Recognition Particle, SRP）及各种分子伴侣介导的蛋白转运过程是广泛存在且高度保守的机制之一。本课题不局限于对保守的SRP、Sec、Tat蛋白转运途径的研究，而是着重于挖掘蛋白转运新途径。通过温度敏感型载体结合诱变方法筛选具有蛋白转运新途径的ΔSRP抑制子，基因组与定量蛋白质组学技术解析ΔSRP抑制子的突变基因及蛋白谱变化，体内回补与体外蛋白互作实验验证新转运途径因子及其作用机理，在此基础上结合生物信息学方法挖掘自然界中天然利用这种蛋白转运新途径的微生物。本工作揭示了SRP蛋白转运途径在物种进化中存在着补偿途径，丰富完善了对蛋白转运途径的认识和理解。同时，蛋白新转运途径的挖掘与发现，为微生物蛋白表达策略提供了新思路，也为医学上针对病原微生物研制新药提供新靶点打下理论基础。

蛋白质的合成、折叠、转运和降解是细胞生物学研究的重点。信号识别颗粒（Signal Recognition Particle， SRP）及各种分子伴侣介导的蛋白转运过程是广泛存在且高度保守的机制之一。本课题不局限于对保守的SRP、Sec、Tat蛋白转运途径的研究，而是着重于挖掘蛋白转运新途径。通过温度敏感型载体结合诱变方法筛选具有蛋白转运新途径的ΔSRP抑制子，基因组与定量蛋白质组学技术解析ΔSRP抑制子的突变基因及蛋白谱变化，体内回补与核糖体组装、翻译系统保真性等实验验证新转运途径因子及其作用机理，在此基础上挖掘大肠杆菌中的蛋白转运新途径。本工作首次获得了缺失ffh基因的大肠杆菌抑制子菌株，在此基础上分析了抑制子菌株的基因组蛋白质组及蛋白翻译合成系统，证实了核糖体和转录起始因子是抑制子菌株中代偿机制的核心突变位点，揭示了SRP蛋白转运途径在物种进化中存在着补偿途径，丰富完善了对蛋白转运途径的认识和理解。同时，微生物蛋白新转运途径的挖掘与发现，为微生物蛋白表达策略提供了新思路。