Structural Studies of a T cell Specific Tyrosine Kinase

描述（由申请人提供）：我们在免疫蛋白激酶白介素-2酪氨酸激酶（Itk）中发现了一种新的脯氨酸驱动的构象开关。Itk Src同源性2 （SH2）结构域内的脯氨酸顺式/反式异构化介导Itk信号伙伴的构象特异性识别以及与普遍存在的肽基脯氨酸异构酶亲环蛋白A （CypA）的重要调节相互作用。该建议的重点是明确定义脯氨酸异构化在Itk调控机构中的作用。我们在结构和功能层面与Itk和CypA合作的丰富经验使我们处于剖析Itk调控机制的理想位置。脯氨酸异构化是一种最容易用核磁共振光谱研究的内在构象交换事件。因此，我们的方法是锚定在结构研究，提供必要的原子水平的信息，以充分检查这些细胞内信号蛋白在T细胞中的功能。我们的结构见解已经在小鼠模型系统中进行了测试，进一步证明Itk调节与单个脯氨酸残基有关，该残基是CypA的靶标。此外，Itk中的脯氨酸异构化事件可能代表了另一种形式的翻译后“修饰”，类似于蛋白质磷酸化。像磷酸化的氨基酸侧链一样，关键的脯氨酸残基可以控制配体的结合，并作为肽基-脯氨酸异构酶等调节伙伴的识别位点。我们对Itk和CypA的详细分析将为鉴定控制免疫细胞信号传导的蛋白质中的其他脯氨酸开关奠定基础。

DESCRIPTION (provided by applicant): We have discovered a novel proline-driven conformational switch in the immunological protein kinase Interleukin-2 tyrosine kinase (Itk). Proline cis/trans isomerization within the Itk Src homology 2 (SH2) domain mediates conformer-specific recognition of Itk signaling partners and an important regulatory interaction with the ubiquitous peptidyl-prolyl isomerase cyclophilin A (CypA). This proposal is focused on clearly defining the role of proline isomerization in the Itk regulatory apparatus. Our extensive experience in working with Itk and CypA both at the structural and functional level places us in an ideal position to dissect the Itk regulatory mechanism. Proline isomerization is an intrinsic conformational exchange event that is most easily studied by NMR spectroscopy. Thus, our approach is anchored in structural studies that provide the indispensable atomic-level information required to fully examine the functions of these intracellular signaling proteins in T cells. Our structural insights have already been tested in mouse model systems providing further evidence that Itk regulation is linked to a single proline residue that is the target of CypA. Moreover, the proline isomerization event within Itk may represent another form of posttranslational 'modification' akin to protein phosphorylation. Like phosphorylated amino acid side chains, critical proline residues could control ligand binding and serve as recognition sites for regulatory partners such as the peptidyl-prolyl isomerases. Our detailed analysis of Itk and CypA will lay the groundwork for identification of other proline switches within the proteins that control immune cell signaling.