副溶血性弧菌脂蛋白定位系统转运蛋白结构与功能的生物信息学分析

Bioinformatics analysis for structure and function of localization of lipoprotein system transporters in Vibrio parahaemolyticus

【目的】副溶血性弧菌是一种重要的人畜共患病原菌,脂蛋白定位系统(Localization of lipoprotein system,Lol)负责该菌脂蛋白的转运与定位,与其致病力及耐药性密切相关,对Lol系统转运蛋白进行系统的生物信息学分析,有助于推动副溶血性弧菌致病与耐药机理的进一步研究。【方法】本文通过生物信息学分析技术,结合ExPASy在线工具、SignalP 4.0 Server、TMHMM-2.0、STRING、SWISS-MODEL等软件,分析了副溶血性弧菌Lol系统转运蛋白LolA-E及LolCD2E的基本性质、蛋白互作关系及三级结构。【结果】LolA和LolB为酸性亲水蛋白,含信号肽位点,无跨膜区域。LolC和LolE为碱性疏水膜蛋白,LolCD2E为中性疏水膜蛋白,LolC-E及LolCD2E均无显著的信号肽位点。蛋白相互作用网络显示,LolA–E五个蛋白的编码基因均共表达,负责脂蛋白的合成与转运,并与BamA、Pal、MacB、CmeC等外膜蛋白具有密切的互作关系。三级结构同源建模发现,副溶血性弧菌与大肠杆菌拥有相似的LolA和LolB结构,LolC-E含有MacB蛋白的同源结构,赋予了该系统消耗ATP运输脂蛋白的重要功能。此外,本研究还首次发现了副溶血性弧菌LolC和LolE中存在一段保守的Hook结构,是LolCD2E复合物与LolA结合并转运脂蛋白的关键区域。【结论】本研究为副溶血性弧菌Lol系统转运蛋白的表达纯化、结构与功能的研究提供了重要的数据基础,为后续抗菌药物的研发提供了新型作用靶点。

[Objective] Vibrio parahaemolyticus is an important zoonotic pathogen. Its localization of lipoprotein(Lol) system is responsible for the transport and localization of lipoproteins in this bacterium, which is closely related to its pathogenicity and drug resistance. Systematic bioinformatics analysis of the Lol system transporters is helpful to promote further research on pathogenesis and resistance mechanism of V. parahaemolyticus. [Methods] The basic properties, protein interactions and tertiary structures of Lol system transporters LolA-E and LolCD2E of V. parahaemolyticus were investigated by bioinformatics analysis, combining with ExPASy online tools, SignalP 4.0 Server, TMHMM-2.0, STRING, SWISS-MODEL and other softwares. [Results] LolA and LolB were acidic hydrophilic proteins with signal peptide sites and no transmembrane region, while LolC and LolE were alkaline hydrophobic membrane proteins. LolCD2E was an neutral hydrophobic membrane protein, and LolC-E and LolCD2E had no significant signal peptide sites. Subsequent studies should add a signal peptide sequence in the recombinant expression vector of LolCD2E protein, and combine with detergent treatment to ensure the protein expression and purification. Protein interaction network showed that the coding genes of five proteins LolA-E can co-express, and be responsible for the synthesis and transport of lipoprotein. LolA-E had close interaction with other outer membrane proteins such as BamA, Pal, MacB and CmeC. Tertiary structural homology modeling showed that V. parahaemolyticus and Escherichia coli had the similar structures in LolA and LolB proteins. And LolC-E contained the homologous structure of MacB protein, which gave Lol system an important function for consuming ATP to transport lipoproteins. Furthermore, this study found a conserved Hook structure in LolC and LolE of V. parahaemolyticus for the first tim, which is a key region for LolCD2E complex to bind with LolA and transport lipoprotein. [Conclusion] This study provides an important data basis for the study of expression purification, structure and function of Lol system transporter in V. parahaemolyticus, and provides new targets for the subsequent research and development of new antibacterials.