School of Chemical Engineering & Technology, Tianjin University, Tianjin 300072, P.R. China.
J Proteome Res. 2012 Nov 2;11(11):5286-300. doi: 10.1021/pr300504w. Epub 2012 Oct 23.
Recent progress in metabolic engineering has led to autotrophic production of ethanol in various cyanobacterial hosts. However, cyanobacteria are known to be sensitive to ethanol, which restricts further efforts to increase ethanol production levels in these renewable host systems. To understand the mechanisms of ethanol tolerance so that engineering more robust cyanobacterial hosts can be possible, in this study, the responses of model cyanobacterial Synechocystis sp. PCC 6803 to ethanol were determined using a quantitative proteomics approach with iTRAQ LC-MS/MS technologies. The resulting high-quality proteomic data set consisted of 24,887 unique peptides corresponding to 1509 identified proteins, a coverage of approximately 42% of the predicted proteins in the Synechocystis genome. Using a cutoff of 1.5-fold change and a p-value less than 0.05, 135 and 293 unique proteins with differential abundance levels were identified between control and ethanol-treated samples at 24 and 48 h, respectively. Functional analysis showed that the Synechocystis cells employed a combination of induced common stress response, modifications of cell membrane and envelope, and induction of multiple transporters and cell mobility-related proteins as protection mechanisms against ethanol toxicity. Interestingly, our proteomic analysis revealed that proteins related to multiple aspects of photosynthesis were up-regulated in the ethanol-treated Synechocystis cells, consistent with increased chlorophyll a concentration in the cells upon ethanol exposure. The study provided the first comprehensive view of the complicated molecular mechanisms against ethanol stress and also provided a list of potential gene targets for further engineering ethanol tolerance in Synechocystis PCC 6803.
近年来，代谢工程的进展使得各种蓝藻宿主能够进行自养生产乙醇。然而，众所周知，蓝藻对乙醇敏感，这限制了在这些可再生宿主系统中进一步提高乙醇产量的努力。为了了解乙醇耐受的机制，以便有可能对更稳健的蓝藻宿主进行工程改造，在这项研究中，我们使用 iTRAQ LC-MS/MS 技术的定量蛋白质组学方法来确定模式蓝藻集胞藻 PCC 6803 对乙醇的反应。由此产生的高质量蛋白质组数据集由 24887 个独特肽段组成，对应于 1509 个鉴定的蛋白质，涵盖了集胞藻基因组中预测蛋白质的大约 42%。使用 1.5 倍变化的截止值和小于 0.05 的 p 值，在 24 和 48 小时时，分别在对照和乙醇处理的样品之间鉴定出 135 和 293 个具有差异丰度水平的独特蛋白质。功能分析表明，集胞藻细胞采用了诱导共同应激反应、细胞膜和包膜的修饰以及多种转运蛋白和细胞迁移相关蛋白的诱导相结合的保护机制，以抵御乙醇毒性。有趣的是，我们的蛋白质组学分析表明，与光合作用的多个方面相关的蛋白质在乙醇处理的集胞藻细胞中上调，这与细胞在乙醇暴露时叶绿素 a 浓度的增加一致。该研究提供了对乙醇应激复杂分子机制的首次全面了解，并为进一步在集胞藻 PCC 6803 中工程改造乙醇耐受提供了潜在的基因靶标列表。
