State Key Laboratory of Medicinal Chemical Biology and Department of Chemistry, Nankai University, Tianjin 300071, China.
J Proteome Res. 2013 Feb 1;12(2):844-51. doi: 10.1021/pr300912q. Epub 2013 Jan 17.
Protein lysine acetylation plays a key role in regulating chromatin dynamics, gene expression and metabolic pathways in eukaryotes, and, thus, contributes to diverse cellular processes like transcription, cell cycle regulation, and apoptosis. Although recent evidence suggests that acetylated proteins impact broadly cellular functions in prokaryotes, the substrates and localization of this modification remain widely unknown due to the limitations of analytical methods. Comprehensive identification of protein acetylation is a major bottleneck due to its dynamic property and pretty low abundance. A complete atlas of acetylome will significantly advance our understanding of this modification functions in prokaryotes. To achieve this goal, we have developed an intergraded approach to identifying lysine acetylation. Combining immunoaffinity enrichment with high sensitive mass spectrometry, we identified 349 acetylated proteins and addressed 1070 acetylation sites in Escherichia coli. To our knowledge, the acetylated proteins and acetylated sites were increased to 3 times and 8 times, respectively, compared to that in previous report. To further characterize this modification, we classified acetylated proteins into several groups according to cell components, molecular functions and biological process. Additionally, interaction networks and high confident domains architectures of acetylated proteins were investigated with the aid of bioinformatics tools. Finally, the acetylated metabolic enzymes were analyzed on the basis of acetylated proteins identified by proteomic survey in E. coli. Our study has demonstrated that the combined approach is powerful for identification and characterization of protein lysine acetylation on a large scale. These results not only greatly expand the number of acetylated proteins, but also provide a series of important information including localization, networks and characterization of acetylome.
蛋白质赖氨酸乙酰化在真核生物中调节染色质动力学、基因表达和代谢途径方面发挥着关键作用，因此有助于转录、细胞周期调控和细胞凋亡等多种细胞过程。尽管最近的证据表明，乙酰化蛋白质会影响原核生物的广泛细胞功能，但由于分析方法的限制，这种修饰的底物和定位仍然知之甚少。由于其动态特性和相当低的丰度，全面鉴定蛋白质乙酰化是一个主要的瓶颈。乙酰组图谱的完整图谱将极大地促进我们对原核生物中这种修饰功能的理解。为了实现这一目标，我们开发了一种综合的方法来鉴定赖氨酸乙酰化。我们将免疫亲和富集与高灵敏度质谱相结合，鉴定了 349 种乙酰化蛋白质，并确定了大肠杆菌中的 1070 个乙酰化位点。据我们所知，与之前的报道相比，乙酰化蛋白质和乙酰化位点分别增加了 3 倍和 8 倍。为了进一步表征这种修饰，我们根据细胞成分、分子功能和生物过程将乙酰化蛋白质分为几类。此外，还借助生物信息学工具研究了乙酰化蛋白质的相互作用网络和高置信结构域结构。最后，根据大肠杆菌蛋白质组学研究中鉴定的乙酰化蛋白质，分析了乙酰化代谢酶。我们的研究表明，这种联合方法对于大规模鉴定和表征蛋白质赖氨酸乙酰化非常有效。这些结果不仅大大增加了乙酰化蛋白质的数量，还提供了一系列重要信息，包括乙酰组的定位、网络和特征。
