Protein acetylation dynamics in response to carbon overflow in Escherichia coli.
Protein acetylation dynamics in response to carbon overflow in Escherichia coli.
复制标题
DOI：
10.1111/mmi.13161
复制
发表时间：
2015-12
影响因子：
3.6
通讯作者：
Wolfe AJ
中科院分区：
文献类型：
--
作者：
Schilling B;Christensen D;Davis R;Sahu AK;Hu LI;Walker-Peddakotla A;Sorensen DJ;Zemaitaitis B;Gibson BW;Wolfe AJ
在大肠杆菌中，蛋白质在赖氨酸处的乙酰化在很大程度上取决于非酶促乙酰磷酸依赖性机制。为了评估这种翻译后修饰的功能意义，我们首先在含有葡萄糖的缓冲胰蛋白胨肉汤中培养野生型细胞，并通过免疫化学监测乙酰化随时间的变化。大多数乙酰化发生在稳定期，并且在进入稳定期后开始减少葡萄糖消耗和乙酸盐排泄。转录rprA，一个固定相调节器，表现出类似的行为。为了确定具有显著乙酰化变化的位点和底物，我们使用无标记定量蛋白质组学来监测蛋白质乙酰化的变化。在生长过程中，确定的网站和乙酰化的程度增加了相当大的变化之间的赖氨酸从相同的蛋白质。由于葡萄糖调节的赖氨酸乙酰化在中心代谢途径中占主导地位，并与乙酰磷酸调节的乙酰化位点重叠，我们删除了主要的碳调节剂CRP，并观察到乙酰化的急剧损失，可以通过删除降解乙酰磷酸的酶来恢复。我们认为，乙酰磷酸依赖的乙酰化是对碳通量的一种反应，可以调节中枢代谢。
In Escherichia coli, acetylation of proteins at lysines depends largely on a non-enzymatic acetyl-phosphate-dependent mechanism. To assess the functional significance of this post-translational modification, we first grew wild-type cells in buffered tryptone broth with glucose, and monitored acetylation over time by immunochemistry. Most acetylation occurred in stationary phase and paralleled glucose consumption and acetate excretion, which began upon entry into stationary phase. Transcription of rprA, a stationary phase regulator, exhibited similar behavior. To identify sites and substrates with significant acetylation changes, we used label-free, quantitative proteomics to monitor changes in protein acetylation. During growth, both the number of identified sites and the extent of acetylation increased with considerable variation among lysines from the same protein. Since glucose-regulated lysine acetylation was predominant in central metabolic pathways and overlapped with acetyl-phosphate-regulated acetylation sites, we deleted the major carbon regulator CRP and observed a dramatic loss of acetylation that could be restored by deleting the enzyme that degrades acetyl phosphate. We propose that acetyl-phosphate-dependent acetylation is a response to carbon flux that could regulate central metabolism.