Dettori Rosalia, Sonzogni Silvina, Meyer Lucas, Lopez-Garcia Laura A, Morrice Nick A, Zeuzem Stefan, Engel Matthias, Piiper Albrecht, Neimanis Sonja, Frödin Morten, Biondi Ricardo M
Department of Internal Medicine I, Research Group PhosphoSites, Johann Wolfgang Goethe University Hospital, D-60590 Frankfurt, Germany.
J Biol Chem. 2009 Oct 30;284(44):30318-27. doi: 10.1074/jbc.M109.051151. Epub 2009 Sep 1.
The members of the AGC kinase family frequently exhibit three conserved phosphorylation sites: the activation loop, the hydrophobic motif (HM), and the zipper (Z)/turn-motif (TM) phosphorylation site. 3-Phosphoinositide-dependent protein kinase 1 (PDK1) phosphorylates the activation loop of numerous AGC kinases, including the protein kinase C-related protein kinases (PRKs). Here we studied the docking interaction between PDK1 and PRK2 and analyzed the mechanisms that regulate this interaction. In vivo labeling of recombinant PRK2 by (32)P(i) revealed phosphorylation at two sites, the activation loop and the Z/TM in the C-terminal extension. We provide evidence that phosphorylation of the Z/TM site of PRK2 inhibits its interaction with PDK1. Our studies further provide a mechanistic model to explain different steps in the docking interaction and regulation. Interestingly, we found that the mechanism that negatively regulates the docking interaction of PRK2 to the upstream kinase PDK1 is directly linked to the activation mechanism of PRK2 itself. Finally, our results indicate that the mechanisms underlying the regulation of the interaction between PRK2 and PDK1 are specific for PRK2 and do not apply for other AGC kinases.
AGC激酶家族成员通常有三个保守的磷酸化位点：激活环、疏水基序（HM）和拉链（Z）/转角基序（TM）磷酸化位点。3-磷酸肌醇依赖性蛋白激酶1（PDK1）可磷酸化众多AGC激酶的激活环，包括蛋白激酶C相关蛋白激酶（PRKs）。在此，我们研究了PDK1与PRK2之间的对接相互作用，并分析了调节这种相互作用的机制。用（32）P（i）对重组PRK2进行体内标记，结果显示在两个位点发生了磷酸化，即激活环和C端延伸区的Z/TM。我们提供的证据表明，PRK2的Z/TM位点磷酸化会抑制其与PDK1的相互作用。我们的研究进一步提供了一个机制模型，以解释对接相互作用和调节的不同步骤。有趣的是，我们发现负向调节PRK2与上游激酶PDK1对接相互作用的机制与PRK2自身的激活机制直接相关。最后，我们的结果表明，PRK2与PDK1相互作用调节的潜在机制对PRK2具有特异性，并不适用于其他AGC激酶。
