Structure and substrate specificity of the Pim-1 kinase
Structure and substrate specificity of the Pim-1 kinase
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DOI：
10.1074/jbc.m510711200
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发表时间：
2005-12-16
影响因子：
4.8
通讯作者：
Turk, BE
中科院分区：
文献类型：
--
作者：
Bullock, AN;Debreczeni, J;Turk, BE
Pim激酶是属于CAMK（钙调蛋白依赖性蛋白激酶相关）组的三种脊椎动物蛋白丝氨酸/苏氨酸激酶（Pim-1、-2和-3）的家族。Pim激酶作为造血细胞中细胞因子信号传导途径的重要介体出现，并且它们有助于某些白血病和实体瘤的进展。许多细胞质和核蛋白被Pim激酶磷酸化，并可在正常生理和疾病中作为其效应物。最近的Pim-1晶体学研究已经确定了独特的结构特征，但还没有提供深入了解激酶如何识别其靶底物。在这里，我们进行了肽库筛选，以彻底确定活性位点介导的Pim-1和Pim-3磷酸化的序列特异性。我们已经确定了Pim-1自磷酸化的主要位点，并令人惊讶地发现，它映射到一个新的网站，偏离其共识磷酸化基序。我们已经解析了与ATP类似物AMP-PNP或双吲哚马来酰亚胺激酶抑制剂2-[1-（3-二甲基氨基丙基）-1H-吲哚-3-基]-3-（1H-吲哚-3-基）马来酰亚胺盐酸盐复合的高亲和力肽底物结合的Pim-1的晶体结构。这些结构揭示了一个意想不到的模式识别的磷酸化位点上游的碱性残基，不同于其他激酶具有类似的底物特异性。这些结构为Pim激酶对肽底物具有异常高的亲和力提供了理论基础，并提出了底物与CAMK组成员结合的一般模式。
The Pim kinases are a family of three vertebrate protein serine/threonine kinases (Pim-1, -2, and -3) belonging to the CAMK (calmodulin-dependent protein kinase-related) group. Pim kinases are emerging as important mediators of cytokine signaling pathways in hematopoietic cells, and they contribute to the progression of certain leukemias and solid tumors. A number of cytoplasmic and nuclear proteins are phosphorylated by Pim kinases and may act as their effectors in normal physiology and in disease. Recent crystallographic studies of Pim-1 have identified unique structural features but have not provided insight into how the kinase recognizes its target substrates. Here, we have conducted peptide library screens to exhaustively determine the sequence specificity of active site-mediated phosphorylation by Pim-1 and Pim-3. We have identified the major site of Pim-1 autophosphorylation and find surprisingly that it maps to a novel site that diverges from its consensus phosphorylation motif. We have solved the crystal structure of Pim-1 bound to a high affinity peptide substrate in complexes with either the ATP analog AMP-PNP or the bisindolylmaleimide kinase inhibitor 2-[1-(3-dimethylaminopropyl)-1H-indol-3- yl]-3-(1H-indol3-yl) maleimide HCl. These structures reveal an unanticipated mode of recognition for basic residues upstream of the phosphorylation site, distinct from that of other kinases with similar substrate specificity. The structures provide a rationale for the unusually high affinity of Pim kinases for peptide substrates and suggest a general mode for substrate binding to members of the CAMK group.