Dynamics & energetics of p38a kinase regulation by ligands

动力学

• 批准号: 8608555
• 负责人: Wolfgang Peti
• 金额: $ 32.05万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-02-01 至 2017-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8608555
• 关键词: Active Sites; Affinity; Alzheimer' s Disease; Amino Acid Sequence; Amino Acids; Architecture; Binding; Binding Sites; C-terminal; Calorimetry; Complex; Coupled; Crystallography; Data; Development; Disease; Docking; Drug Design; Drug Interactions; Entropy; Family; Foundations; Goals; Growth Factor; Heart; Human Biology; Hydrogen; Knowledge; Laboratories; Libraries; Ligands; Light; Lobe; MAP Kinase Gene; MAP2K6 gene; MAPK Signaling Pathway Pathway; MAPK14 gene; MAPK8 gene; Malignant Neoplasms; Measures; Mediating; Methods; Mitogen-Activated Protein Kinases; Modeling; Molecular Conformation; Molecular Structure; Motion; NMR Spectroscopy; Nuclear Magnetic Resonance; Pathway interactions; Pharmaceutical Preparations; Pharmacologic Substance; Phosphoric Monoester Hydrolases; Phosphotransferases; Positioning Attribute; Process; Protein Tyrosine Phosphatase; Proteins; Proxy; Radial; Regulation; Relaxation; Renaissance; Reporting; Resolution; Rest; Rheumatoid Arthritis; Role; Sampling; Side; Signal Transduction; Site; Specificity; Stimulus; Structure; Techniques; Tertiary Protein Structure; Testing; Thermodynamics; Time; Titrations; Vertebral column; base; cell type; cost effective; cytokine; design; extracellular; genetic regulatory protein; human disease; improved; information gathering; inhibitor/antagonist; insight; man; nervous system disorder; novel; pressure; protein complex; protein structure; public health relevance; research study; response; small molecule

DESCRIPTION (provided by applicant): High-affinity protein complexes are critical to a large number of intricate regulatory processes. Their formation involves a complicated manifold of interactions that are diverse and complex. This complexity is reflected in the difficulty of computing the energetics of interactions between proteins using molecular structure alone. Indeed, the structure-based design of pharmaceuticals has been significantly impeded by this barrier. Understanding the fundamental origins of the energetics and dynamics of the interactions of proteins with both natural and pharmacological ligands is clearly critical to the optimization of "rational" drug design. Recent advances in nuclear magnetic resonance (NMR) relaxation methods have enabled the use of measures-of-motion between conformational states of a protein as a proxy for conformational entropy. There is now a strong indication from recent studies utilizing this approach that changes in conformational entropy can significantly influence the thermodynamics of the interaction of small molecule ligands with proteins. Therefore, we will examine this and related issues in the context of the ser/thr kinase p38¿. p38¿ is intimately associated with a variety of disease states, including cancer and neurological diseases, and is an active target for pharmaceutical development. Experiments are proposed to examine the changes in fast internal motion in this protein upon interaction with both natural and pharmacological small molecule ligands. Advanced NMR relaxation methods will be employed to measure main chain and side chain motion. A variety of analytical strategies will be used to gain insight into the quantitative contributions to the thermodynamics of complex formation and to discover their structural origins. In addition, the dynamical effects of regulatory protein bindng will also be examined. These data will go to the heart of the physical mechanism for activation and deactivation of this critical kinase by both natural effector proteins and man-made molecules. Complementary hydrogen exchange studies will also be carried out with the goal of exposing cooperative interactions within p38¿. This view will be particularly informative with respect to the emerging class of pseudo-allosteric drugs. A novel NMR-based approach using high-pressure perturbation and rapid three dimensional radial sampling will be employed to overcome limitations in the standard "native state" hydrogen exchange method in the context of large proteins, such as p38¿. Overall, the proposal rests on a significant foundation of preliminary results including an unusually deep and robust library of resonance assignments for a ser/thr kinase.

描述（由申请人提供）：高亲和力蛋白质复合物对于大量复杂的监管过程至关重要。它们的形成涉及多种复杂的相互作用。这种复杂性反映在仅使用分子结构计算蛋白质之间相互作用的能量学的难度上。事实上，基于结构的药物设计已受到这一障碍的严重阻碍。了解蛋白质与天然配体和药理学配体相互作用的能量学和动力学的基本起源对于优化“合理”药物设计显然至关重要。核磁共振 (NMR) 弛豫方法的最新进展使得能够使用蛋白质构象状态之间的运动测量作为构象熵的代表。最近利用这种方法的研究强烈表明，构象熵的变化可以显着影响小分子配体与蛋白质相互作用的热力学。因此，我们将在 ser/thr 激酶 p38 的背景下研究这个问题和相关问题。 p38¿与多种疾病状态密切相关，包括癌症和神经系统疾病，并且是药物开发的活跃靶点。建议进行实验来检查该蛋白质在与天然和药理学小分子配体相互作用时快速内部运动的变化。将采用先进的核磁共振弛豫方法来测量主链和侧链运动。将使用各种分析策略来深入了解复杂形成热力学的定量贡献并发现其结构起源。此外，还将检查调节蛋白结合的动力学效应。这些数据将深入研究天然效应蛋白和人造分子激活和失活这一关键激酶的物理机制的核心。互补的氢交换研究也将进行，目的是揭示 p38¿ 内的合作相互作用。这种观点对于新兴的伪变构药物类别尤其有用。将采用一种基于 NMR 的新型方法，该方法使用高压扰动和快速三维径向采样，以克服大蛋白质（例如 p38¿）中标准“天然状态”氢交换方法的局限性。总体而言，该提案建立在初步结果的重要基础上，包括异常深入且强大的丝氨酸/苏氨酸激酶共振分配库。