School of Medicine and Institute of Biomedical Technology, University of Tampere, 33014 Tampere, Finland.
Biochem Soc Trans. 2013 Aug;41(4):1002-7. doi: 10.1042/BST20130005.
JAK (Janus kinase) 2 plays a critical role in signal transduction through several cytokine receptors. JAKs contain a typical tyrosine kinase domain preceded by a pseudokinase [JH2 (JAK homology 2)] domain which has been considered to be catalytically inactive. Identification of activating mutations in the JH2 domain of JAK2 as the major cause for polycythaemia vera and other MPNs (myeloproliferative neoplasms) demonstrate the critical regulatory function for this domain, but the underlying mechanisms have remained elusive. We have performed biochemical and functional analysis on the JH2 domain of JAK2. The results indicate that JH2 functions as an active protein kinase and phosphorylates two residues in JAK2 (Ser523 and Tyr570) that have been shown previously to be negative regulatory sites for JAK2 activity. The crystal structure of the JAK2 JH2 domain provides an explanation for the functional findings and shows that JH2 adopts a prototypical kinase fold, but binds MgATP through a non-canonical mode. The structure of the most prevalent pathogenic JH2 mutation V617F shows a high level of similarity to wild-type JH2. The most notable structural deviation is observed in the N-lobe αC-helix. The structural and biochemical data together with MD (molecular dynamics) simulations show that the V617F mutation rigidifies the αC-helix, which results in hyperactivation of the JH1 domain through an as yet unidentified mechanism. These results provide structural and functional insights into the normal and pathogenic function of the JH2 domain of JAK2.
JAK（Janus kinase）2 通过几种细胞因子受体在信号转导中发挥关键作用。JAK 包含一个典型的酪氨酸激酶结构域，前面是一个假激酶 [JH2（JAK 同源 2）] 结构域，该结构域被认为是无催化活性的。在 JAK2 的 JH2 结构域中鉴定出激活突变是导致真性红细胞增多症和其他 MPN（骨髓增殖性肿瘤）的主要原因，这证明了该结构域的关键调节功能，但潜在的机制仍不清楚。我们对 JAK2 的 JH2 结构域进行了生化和功能分析。结果表明，JH2 作为一种活性蛋白激酶发挥作用，并磷酸化 JAK2 中的两个残基（Ser523 和 Tyr570），这两个残基先前被证明是 JAK2 活性的负调节位点。JAK2 JH2 结构域的晶体结构为功能发现提供了一个解释，并表明 JH2 采用了典型的激酶折叠，但通过非典型模式结合 MgATP。最常见的致病性 JH2 突变 V617F 的结构显示出与野生型 JH2 高度相似。最显著的结构偏差出现在 N- lobe αC-helix 中。结构和生化数据以及 MD（分子动力学）模拟表明，V617F 突变使 αC-helix 僵化，通过尚未确定的机制导致 JH1 结构域的过度激活。这些结果为 JAK2 的 JH2 结构域的正常和病理功能提供了结构和功能上的见解。
