Autophosphorylation of FGFR1 kinase is mediated by a sequential and precisely ordered reaction
Autophosphorylation of FGFR1 kinase is mediated by a sequential and precisely ordered reaction
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DOI：
10.1016/j.molcel.2006.01.022
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发表时间：
2006-03-03
期刊：
影响因子：
16
通讯作者：
Anderson, KS
中科院分区：
文献类型：
--
作者：
Furdui, CM;Lew, ED;Anderson, KS
由细胞外信号诱导的细胞蛋白酪氨酸磷酸化是控制多种细胞过程的关键介质。在这里，我们描述了一种集成的实验方法，包括快速猝灭方法和ESI-LC-MS/MS以及时间分辨ESI-MS，以证明fgf受体1 （FGFR1）的催化酪氨酸激酶结构域的酪氨酸自磷酸化是由一个顺序和精确有序的反应介导的。我们还证明，激活环中Y653的自磷酸化后，两种FGFR底物的催化速率提高了50- 100倍，而激活环中第二个位点（Y654）的自磷酸化导致底物磷酸化速率提高了500- 1000倍。我们提出FGFR1是由严格有序和受调控的自磷酸化介导的两步机制激活的，这表明不同的磷酸化状态可能为受体信号传导提供了时间和空间分辨率。
Tyrosine phosphorylation of cellular proteins induced by extracellular cues serves as a critical mediator in the control of a great variety of cellular processes. Here, we describe an integrated experimental approach including rapid quench methodology and ESI-LC-MS/MS as well as time-resolved ESI-MS to demonstrate that tyrosine autophosphorylation of the catalytic tyrosine kinase domain of FGF-receptor-1 (FGFR1) is mediated by a sequential and precisely ordered reaction. We also demonstrate that the rate of catalysis of two FGFR substrates is enhanced by 50- to 100-fold after autophosphorylation of Y653 in the activation loop, whereas autophosphorylation of the second site in the activation loop (Y654) results in 500- to 1000-fold increase in the rate of substrate phosphorylation. We propose that FGFR1 is activated by a two-step mechanism mediated by strictly ordered and regulated autophosphorylation, suggesting that distinct phosphorylation states may provide both temporal and spatial resolution to receptor signaling.