Conformational Dynamics of Protein Tyrosine Kinases Src and Abl

蛋白酪氨酸激酶 Src 和 Abl 的构象动力学

• 批准号: 7319435
• 负责人: Markus A Seeliger
• 金额: $ 7.98万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-07-15 至 2009-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7319435
• 关键词: ABL1 gene; Affinity; Amino Acids; Arts; Binding; Cancer Patient; Cell division; Characteristics; Chimeric Proteins; Chronic; Chronic Myeloid Leukemia; Collaborations; Complex; Condition; Data; Depth; Disease; Drug Delivery Systems; Drug Interactions; Drug Regulations; Drug resistance; Energy Metabolism; Event; Gleevec; Goals; Heteronuclear NMR; Imatinib; Link; Maps; Measures; Mediating; Methodology; Molecular Conformation; Motion; Movement; Mutation; Myeloid Leukemia; Nitrogen; Nuclear Magnetic Resonance; Patients; Peptides; Pharmaceutical Preparations; Phosphotransferases; Population; Preparation; Process; Protein Dynamics; Protein Kinase; Protein Region; Proteins; Proto-Oncogenes; Range; Rate; Regulation; Relative (related person); Research; Research Personnel; Resistance; Resolution; Roentgen Rays; Role; Sampling; Signal Transduction; Specificity; Structure; System; Therapeutic; Time; Time Study; Vertebral column; base; c-abl Proto-Oncogenes; drug sensitivity; inhibitor/antagonist; instrumentation; interest; leukemia; millisecond; nanosecond; programs; protein-tyrosine kinase c-src; research study; src-Family Kinases

DESCRIPTION (provided by applicant): Protein kinases mediate many cell signaling events, and their tight control is essential for regulating essential processes ranging from cell division to energy metabolism. Thus, it is not surprising that protein kinases are directly or indirectly involved in many diseases and that kinases are key drug targets. For example, Src kinase was the first identified proto-oncogene and the formation of a de-regulated Abl fusion protein (BCR-Abl) is the cause of disease in 95% of patients with chronic myeloid leukemia. X-ray crystal structures have shown that the same kinases can obtain an active and various inactive conformations, implying that kinases are inherently flexible. How the active and inactive states are stabilized and how the states interconvert are key questions in understanding kinase regulation. Because X-ray crystal structures provide only static snapshots, we will use nuclear magnetic resonance (NMR) experiments to study the time scales and amplitudes of structural interconversions in Abl and Src kinase domains. BCR-Abl is the target of the clinically highly successful drug imatinib (Gleevec(r), Novartis) in the treatment of chronic myelogenous leukemia (CML). Why does imatinib bind and inhibit c-Abl but not the structurally closely related c-Src kinase? The crystal structure of Src in complex with imatinib shows protein-drug interactions similar to that of Abl, even though the affinity of imatinib for Src is orders of magnitude lower than for Abl. Because imatinib binds only to the inactive conformation of the kinase, drug binding is intimately related to the interconversion between active and inactive states. The goal of this study is to examine whether differences in this interconversion underlie the differential sensitivities for imatinib. Therefore, we will compare the time scales and amplitudes of backbone motions between Src and Abl kinases in the presence of imatinib by NMR experiments. In preparation for these dynamics experiments, we have established expression systems and NMR conditions and will next pursue the assignment of the Src and Abl NMR spectra. Kinase inhibitory drugs such as imatinib have a great therapeutic potential because of the many signaling events that protein kinases mediate. However, these drugs have to be exceptionally specific for their target kinase and resistance mutations can render these drugs ineffective as seen in leukemia patients under imatinib treatment. The proposed experiments will clarify how inhibitors such as imatinib exploit the characteristic movements of kinase proteins, rather than just their structures, to achieve specificity. Furthermore, the results will have broader impact on the understanding of the fundamental mechanisms of kinase regulation and of drug resistance mutations that are known to arise in cancer patients undergoing kinase inhibitory treatment.

描述（由申请人提供）：蛋白激酶介导许多细胞信号事件，其严格控制对于调节从细胞分裂到能量代谢的基本过程至关重要。因此，毫不奇怪，蛋白激酶直接或间接地参与许多疾病，并且激酶是关键的药物靶标。例如，SRC激酶是第一个鉴定出的原始癌基因，并且在95％的慢性髓样白血病患者中，下降的ABL融合蛋白（BCR-ABL）的形成是疾病的原因。 X射线晶体结构表明，相同的激酶可以获得活跃和各种非活动构象，这意味着激酶本质上是柔性的。活性状态和非活动状态如何稳定以及状态如何互连是理解激酶调节的关键问题。由于X射线晶体结构仅提供静态快照，因此我们将使用核磁共振（NMR）实验来研究ABL和SRC激酶结构域中结构互连的时间尺度和振幅。 BCR-ABL是临床上非常成功的药物伊马替尼（Gleevec（r），诺华）的靶标，在慢性骨髓性白血病（CML）治疗中。伊马替尼为什么结合和抑制C-ABL，而不结合结构密切相关的C-SRC激酶？与伊马替尼复合物中SRC的晶体结构显示出类似于ABL的蛋白质 - 药物相互作用，即使伊马替尼对SRC的亲和力比ABL低的数量级。由于伊马替尼仅与激酶的非活性构象结合，因此药物结合与活性状态和非活性状态之间的互连密切相关。这项研究的目的是检查这种相互转换的差异是否是伊马替尼的差异敏感性的基础。因此，我们将在NMR实验存在伊马替尼的情况下比较SRC和ABL激酶之间主链运动的时间尺度和振幅。为了准备这些动力学实验，我们建立了表达系统和NMR条件，然后接下来追求SRC和ABL NMR光谱的分配。 激酶抑制性药物（例如伊马替尼）具有巨大的治疗潜力，因为蛋白激酶介导的许多信号传导事件。但是，这些药物必须特别具体，因为它们的靶激酶酶，耐药突变可能会使这些药物在伊马替尼治疗下在白血病患者中所见，这些药物无效。提出的实验将阐明诸如伊马替尼之类的抑制剂如何利用激酶蛋白的特征运动，而不仅仅是其结构，以达到特异性。此外，结果将对对激酶调节和耐药性突变的基本机制的理解产生更广泛的影响，这些突变是在接受激酶抑制治疗的癌症患者中出现的。