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Dynamics of protein phosphorylation on Ser/Thr/Tyr in Bacillus subtilis

机译：枯草芽孢杆菌中Ser / Thr / Tyr蛋白磷酸化的动力学

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摘要

The Ser/Thr/Tyr phosphoproteome of Bacillus subtilis was analyzed by a 2-D gel-based approach combining Pro-Q Diamond staining and [P-33]-labeling. In exponentially growing B. subtilis cells 27 proteins could be identified after staining with Pro-Q Diamond and/or [(33)p]-labeling and one additional protein was labeled solely by [(33)p] resulting in a total of 28 potentially phosphorylated proteins. These proteins are mainly involved in enzymatic reactions of basic carbon metabolism and the regulation of the alternative sigma factor sigma(B). We also found significant changes of the phosphoproteome including increased phosphorylation and dephosphorylation rates of some proteins as well as the detection of four newly phosphorylated proteins in response to stress or starvation. For nine proteins, phosphorylation sites at serine or threonine residues were determined by MS. These include the known phosphorylation sites of Crh, PtsH, and RsbV. Additionally, we were able to identify novel phosphorylation sites of AroA, Pyk, and YbbT. Interestingly, the phosphorylation of RsbRA, B, C, and D, four proteins of a multicomponent protein complex involved in environmental stress signaling, was found during exponential growth. For RsbRA, B, and D, phosphorylation of one of the conserved threonine residues in their C-termini were verified by MS (T171, T186, T181, respectively).
机译：枯草芽孢杆菌的Ser / Thr / Tyr磷酸化蛋白质组通过结合Pro-Q Diamond染色和[P-33]标记的基于2D凝胶的方法进行分析。在以指数方式生长的枯草芽孢杆菌细胞中，用Pro-Q Diamond和/或[（33）p]标记染色后可鉴定出27种蛋白质，另外一种蛋白质仅通过[（33）p]标记，总共28种潜在的磷酸化蛋白。这些蛋白质主要参与基本碳代谢的酶促反应和替代sigma因子sigma（B）的调节。我们还发现了磷酸化蛋白质组的显着变化，包括某些蛋白质的磷酸化和去磷酸化速率提高，以及响应压力或饥饿而检测到的四个新磷酸化蛋白质。对于9种蛋白质，通过MS确定丝氨酸或苏氨酸残基的磷酸化位点。这些包括Crh，PtsH和RsbV的已知磷酸化位点。此外，我们能够鉴定AroA，Pyk和YbbT的新型磷酸化位点。有趣的是，在指数生长过程中发现了RsbRA，B，C和D的磷酸化，这是参与环境胁迫信号传导的多组分蛋白质复合物中的四种蛋白质。对于RsbRA，B和D，通过MS（分别为T171，T186和T181）验证了其C末端中一个保守的苏氨酸残基的磷酸化。

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