Regulatory mechanisms of the Crk Adapter protein

Crk Adapter 蛋白的调控机制

• 批准号: 7688559
• 负责人: CHARALAMPOS KALODIMOS
• 金额: $ 27.71万
• 依托单位: RUTGERS, THE STATE UNIV OF N.J.
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-08-01 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7688559
• 关键词: ABL1 gene; Adaptor Signaling Protein; Apoptotic; Behavior; Binding; Biological Assay; Biological Process; Cancerous; Catalytic Domain; Cell physiology; Cells; Complex; Coupling; Cyclophilin A; Data; Differentiation and Growth; Enzymes; Event; Family; Human; In Vitro; Kinetics; Knowledge; Lead; Length; Ligand Binding; Malignant Neoplasms; Mediating; Methodology; Molecular; Molecular Conformation; Monitor; Mutation; Oncogenes; Oncogenic; Peptidylprolyl Isomerase; Phagocytosis; Phosphopeptides; Phosphorylation; Phosphotransferases; Physiological; Play; Process; Proline; Property; Proteins; Recruitment Activity; Regulation; Research; Research Proposals; Resolution; Role; SH3 Domains; Signal Transduction; Solutions; Specificity; Stimulus; Structure; Testing; Translating; Tyrosine; adapter protein; base; c-abl Proto-Oncogenes; cell transformation; cis trans isomerization; conformer; design; drug candidate; extracellular; in vivo; leukemia; novel; polypeptide; polyproline; protein complex; stem; tumorigenesis

DESCRIPTION (provided by applicant): Crk-family adaptors mediate the timely formation of protein complexes elicited by a variety of external and intrinsic stimuli and contribute to the specificity of signal transduction events by recruiting active enzymes into signaling networks. Despite the fact that adaptor proteins have no intrinsic catalytic or transcriptional activity, their aberrant behavior may lead to oncogenic transformation. Our long-term objectives are to (i) provide the structural basis of Crk-mediated signaling phenomena; (ii) determine the molecular features of the interaction between Crk and Abl, which results in Abl trans-activation; (iii) understand the physicochemical basis of regulatory phenomena in cell signaling; and (iv) design drug candidates that could control aberrant signal transduction events leading to cancer. The strong hypothesis behind the proposed research is that the binding activity of Crk is regulated by a novel autoinhibitory mechanism. Notably, autoinhibition is further controlled by an intrinsic switch afforded by prolyl cis-trans isomerization whose kinetics is effectively catalyzed by cyclophilin A (CypA). We have additionally identified that P221, which lies by the tyrosine residue (Y222) that becomes phosphorylated by c-Abl, undergoes independently cis/trans isomerization that is also catalyzed by CypA. We aim to: 1. Delineate the structural and dynamic features of both the cis and trans conformers of the linker-SH3C Crk fragment, to understand how cis/trans isomerization at P238 regulates the interaction between the linker and the SH3C domain. 2. Determine the molecular basis of the intramolecular SH3N-SH3C interaction, which forms the basis of an autoinhibitory mechanism. 3. Characterize the structural and dynamic properties of full-length Crk (c-Crkll), to understand how distinct regulatory mechanisms integrate in the context of the physiological cellular form of Crk. 4. Determine the effect of the autoinhibition and proline cis/trans isomerization on the biological function of Crk, both in vitro and in vivo. Crk proteins are involved in mediating many signaling events during the physiological cellular function, but it may also result in tumorigenesis. In this proposal, we will investigate the mechanisms that regulate the activity of the Crk protein, that is, how the protein is switched "on" and "off." We hope to ultimately translate the gained knowledge into new strategies for the rational design of drug candidates that could inhibit proliferation of cancerous cells.

描述（由申请人提供）：Crk家族衔接子介导由多种外部和内在刺激引发的蛋白质复合物的及时形成，并通过将活性酶募集到信号传导网络中而促进信号转导事件的特异性。尽管衔接蛋白没有内在的催化或转录活性，但它们的异常行为可能导致致癌转化。我们的长期目标是（i）提供Crk介导的信号传导现象的结构基础;（ii）确定Crk和Abl之间相互作用的分子特征，这导致Abl反式激活;（iii）了解细胞信号传导中调节现象的物理化学基础;以及（iv）设计可以控制导致癌症的异常信号转导事件的候选药物。提出的研究背后的强有力的假设是，Crk的结合活性由一种新的自抑制机制调节。值得注意的是，自身抑制进一步控制的内在开关提供脯氨酰顺反异构化，其动力学有效地催化亲环素A（CypA）。我们还发现，P221，这是由酪氨酸残基（Y222），成为磷酸化的c-Abl，经历独立的顺式/反式异构化，也是由CypA催化。我们的目标是： 1.描述linker-SH 3C Crk片段的顺式和反式构象的结构和动力学特征，以了解P238处的顺式/反式异构化如何调节linker与SH 3C结构域之间的相互作用。 2.确定分子内SH 3 N-SH 3C相互作用的分子基础，这形成了自抑制机制的基础。 3.表征全长Crk（c-Crkll）的结构和动力学特性，以了解不同的调控机制如何整合到Crk的生理细胞形式中。 4.在体外和体内测定自抑制和脯氨酸顺/反异构化对Crk生物学功能的影响。 Crk蛋白参与介导细胞生理功能过程中的许多信号转导事件，但也可能导致肿瘤的发生。在这个提议中，我们将研究调节Crk蛋白活性的机制，也就是说，蛋白质是如何“打开”和“关闭”的。“我们希望最终将获得的知识转化为新的策略，用于合理设计可以抑制癌细胞增殖的候选药物。