A unique phosphorylation-dependent mechanism for the activation of Ca2+/calmodulin-dependent protein kinase type IV/GR
A unique phosphorylation-dependent mechanism for the activation of Ca2+/calmodulin-dependent protein kinase type IV/GR
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DOI：
10.1074/jbc.271.35.21542
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发表时间：
1996-08-30
影响因子：
4.8
通讯作者：
Means, AR
中科院分区：
文献类型：
--
作者：
Chatila, T;Anderson, KA;Means, AR
钙/钙调蛋白依赖的蛋白激酶IV/Gr(CaMKIV/Gr)的活性在体外和淋巴细胞中都受到三个残基的磷酸化和抗原受体介导的信号的严格调控。一个残基Thr-200对于CaMKIV/Gr激酶增强钙/钙调蛋白依赖的基础活性是必不可少的。此事件需要全长CaMKIV/Gr中的钙/钙调蛋白，但当使用CaMKIV/Gr的截短版本作为底物(Delta CaMKIV/Gr(1-317)(Delta 1-317))时，此事件不依赖于钙/钙调蛋白。另外两个残基Ser(12)和Ser(13)明显被钙/钙调蛋白结合的CaMKIV/Gr自动磷酸化。Ser(12)-Ser(13)和Thr(312)(位于CaMKIIα的Thr(286)的同源位置上的残基)的磷酸化都不会影响CaMKIV/Gr的钙/钙调素非依赖性活性的发展或任何其他性质。同样，去除NH2末端的20个氨基酸对CaMKIV/Gr的激活或功能没有影响。然而，在Delta 1-317中，Ser(12)和Ser(13)残基都突变为Ala，完全丧失活性，而单独的替换没有影响。这些结果表明，NH2末端的Ser簇介导了一种新型的空间结构抑制，并表明CaMKIV/Gr的激活需要三个事件：1)钙/钙调蛋白结合；2)钙/钙调蛋白结合酶在苏氨酸(200)上的磷酸化；以及3)Ser(12)-Ser(13)的自动磷酸化。这三个步骤的要求在多功能钙/钙调蛋白依赖的蛋白激酶中是唯一的。
The activity of the Ca2+/calmodulin-dependent protein kinase IV/Gr (CaMKIV/Gr) is shown to be strictly regulated by phosphorylation of three residues both in vitro and in response to antigen receptor-mediated signaling in lymphocytes. One residue, Thr-200, is indispensable for enhancement of Ca2+/calmodulin-dependent basal activity by CaMKIV/Gr kinase. This event requires Ca2+/calmodulin in the full-length CaMKIV/Gr but is Ca2+/calmodulin-independent when a truncated version of CaMKIV/Gr is used as a substrate (Delta CaMKIV/Gr(1-317) (Delta 1-317)). The other two residues, Ser(12) and Ser(13), are apparently autophosphorylated by the Ca2+/calmodulin-bound CaMKIV/Gr. Phosphorylation of neither Ser(12)-Ser(13) nor Thr(312) (the residue in a homologous position to Thr(286) of CaMKII alpha influences the development of Ca2+/calmodulin-independent activity or any other property of CaMKIV/Gr examined. Similarly, removal of the NH2-terminal 20 amino acids has no effect on the activation or function of CaMKIV/Gr. However, mutation of both Ser(12) and Ser(13) residues to Ala in Delta 1-317 completely abrogates activity, while individual substitutions have no effect. These results indicate that the NH2-terminal Ser cluster mediates a novel type of intrasteric inhibition and suggest that three events are required for CaMKIV/Gr activation: 1) Ca2+/calmodulin binding; 2) phosphorylation of the Ca2+/calmodulin-bound enzyme on Thr(200) by a Ca2+/calmodulin-dependent protein kinase kinase; and 3) autophosphorylation of Ser(12)-Ser(13). This three-step requirement is unique among the multifunctional Ca2+/calmodulin-dependent kinases.