Structure and regulation of non-receptor tyrosine kinases

非受体酪氨酸激酶(NRTK)的功能受到严格调控，整合了细胞信号中的开关 转导。这里我们关注三个NRTK家族的代表，它们具有不同的多域 结构：JAK家族、粘着斑激酶(FAK)和TEC家族成员ITK(IL-2可诱导 酪氨酸激酶)。我们的长期目标是从结构上阐明NRTK的自我调节机制 水平，以了解这些调控机制是如何在癌症中被破坏的，并使用结构性见解来 促进新型抑制剂的发现。与我们的合作者一起，我们正在结合结构生物学的工具， 生物化学、细胞生物学和化学以统一的方式推进这些目标。我们提出了三个具体的 目标。首先，我们将发现Jak激酶如何识别它们的同源细胞因子受体，以及它们是如何 受其构成域之间的相互作用所调节。我们已经结晶了一段N末端片段 JAK2，并制备了这部分JAK2与促红细胞生成素胞质尾部的复合体 用于结构分析的受体。我们对JAK调节的研究建立在我们最近确定的 接头/伪激酶调节模块，并通过阐明基本上全长的Jak2的结构， 我们将解释这个模块是如何控制相邻的激酶域的活性的，以及这种控制是如何进行的 在骨髓增殖性肿瘤中被V617F突变破坏。第二，我们将从结构上决定如何 粘着斑激酶(FAK)被PI(4，5)P2激活。这一目标建立在我们对 FAK处于自抑制状态，我们发现它与PI(4，5)P2结合并被激活。第三，我们将 阐明ITK的SH3-SH2-激酶片段的结构，以了解其调控机制，为进一步研究ITK的功能奠定基础 抑制剂的发现。基于我们的结构洞察力，我们正在开发专门针对JAK3的不可逆转抑制剂 这可能对治疗自身免疫性和炎症性疾病有用。

Non-receptor tyrosine kinases (NRTKs) function as tightly regulated, integrating switches in cellular signal transduction. Here we focus on representatives of three NRTK families, which have distinct multi-domain architectures: Jak-family kinases, Focal Adhesion Kinase (FAK), and the Tec-family member ITK (Il-2 inducible tyrosine kinase). Our longterm goals are to elucidate autoregulatory mechanisms of NRTKs at a structural level, to understand how these regulatory mechanisms are disrupted in cancer, and to use structural insights to facilitate discovery of novel inhibitors. With our collaborators, we are combining the tools of structural biology, biochemistry, cell biology, and chemistry in a unified way to advance these goals. We propose three Specific Aims. First, we will discover how Jak kinases recognize their cognate cytokine receptors and how they are regulated by interactions among their constituent domains. We have crystallized an Nterminal fragment of Jak2, and have also prepared a complex of this portion of Jak2 with the cytoplasmic tail of the erythropoietin receptor for structural analysis. Our studies of Jak regulation build on our recently determined structure of a linker/pseudokinase regulatory module, and through elucidation of the structure of essentially full-length Jak2, we will explain how the this module controls the activity of the adjacent kinase domain, and how this control is disrupted by the V617F mutation in myeloproliferative neoplasms. Second, we will determine structurally how focal adhesion kinase (FAK) is activated by PI(4,5)P2. This aim builds on our determination of the structure of FAK in its autoinhibited state, and our discovery that it binds and is activated by the PI(4,5)P2. Third, we will elucidate the structure an SH3-SH2-kinase fragment of Itk in order to understand its regulation and to facilitate inhibitor discovery. Based on our structural insights, we are developing irreversible inhibitors specific for Jak3 that may be useful in treatment of autoimmune and inflammatory disorders.