Protein dynamics, entropy and function

描述（由申请人提供）：蛋白质的分子识别是几乎所有生物过程的基础，特别是细胞信号转导基础的蛋白质关联。要了解蛋白质-蛋白质相互作用的基础，就必须充分描述它们相互作用的热力学。从历史上看，通过实验估计剩余蛋白质熵的变化几乎是不可能的，剩余蛋白质熵是蛋白质结合自由能的潜在重要组成部分。然而，溶液核磁共振光谱最近成为表征蛋白质动力学的有力工具，从而获得了其剩余熵。核磁共振弛豫研究提供的钙调素中蛋白质动力学（熵）的功能依赖性的初步观点是惊人的。钙调素的内部动力学变化在结合多种靶结构域时变化显著。令人惊讶的是，相应残差熵的明显变化与总体结合熵呈线性关系。这些结果表明，蛋白质构象熵的变化对蛋白质与配体结合的自由能有重要影响。因此，似乎很明显，蛋白质熵可以通过生物进化或人为干预（如设计蛋白质靶向药物）在高亲和相互作用的成熟中被利用。这一建议旨在更充分地描述蛋白质内部动力学的性质，并确定相应的熵对控制配体结合的热力学的贡献程度。为了实现这一目标，我们将扩展基于核磁共振的内部运动探针库，并将其应用于各种环境中。钙调素与调节蛋白的钙调素结合域的相互作用将继续是一个中心模型系统。钙调素对钙介导的信号转导至关重要，并密切参与关键生理和细胞反应的调节，包括控制消化道平滑肌收缩。它与300多种蛋白质相互作用并影响其活性。它也是一个表现良好的基于核磁共振的弛豫研究系统，并有望为蛋白质-配体相互作用中蛋白质动力学（熵）的性质和作用提供丰富的见解。此外，为了评估从钙调素系统中获得的见解是否具有普遍性，我们将对其他蛋白质-配体相互作用进行类似的研究。这将包括白血病抑制因子和肿瘤抑制素M细胞因子与GP130受体的相互作用。GP130是许多重要信号网络的交叉点，具有重要的临床意义，特别是在炎症性肠病和癌症中。在合作工作中，我们将比较晶体学分析和分子动力学模拟与实验结果，并对结合的热力学起源进行计算分析。总的来说，这些研究有望揭示蛋白质运动的物理起源及其生物学意义的新见解，并将开始阐明它们在伪变构药物抑制剂的背景下开发的潜力。

DESCRIPTION (provided by applicant): Molecular recognition by proteins is fundamental to almost every biological process, particularly the protein associations underlying cellular signal transduction. Understanding the basis for protein- protein interactions requires the full characterization of the thermodynamics of their association. Historically it has been virtually impossible to experimentally estimate changes in residual protein entropy, a potentially important component of the free energy of protein association. However, solution NMR spectroscopy has recently emerged as a powerful tool for characterizing the dynamics of proteins and has thereby gained access to their residual entropy. The initial view provided by NMR relaxation studies of the functional dependence of protein dynamics (entropy) in calmodulin is startling. The change in internal dynamics of calmodulin varies significantly upon binding a variety of target domains. Surprisingly, the apparent change in the corresponding residual entropy is linearly related to the overall binding entropy. These results indicate that changes in protein conformational entropy can contribute significantly to the free energy of protein-ligand association. It therefore seems evident that protein entropy can be exploited in the maturation of high affinity interactions either by biological evolution or by human intervention such as in the design of protein-targeted pharmaceuticals. This proposal seeks to more fully characterize the nature of the internal dynamics of proteins and to establish the degree to which the corresponding entropy contributes to the thermodynamics governing the binding of ligands. To achieve this, we will expand the library of NMR-based probes for internal motion and apply them in a variety of contexts. The interaction of calmodulin with calmodulin-binding domains of regulated proteins will continue to be a central model system. Calmodulin is critical to calcium-mediated signal transduction and is intimately involved in the regulation of critical physiological and cellular responses including control of smooth muscle contraction in the digestive tract. It interacts with and influences the activity of over 300 proteins. It is also a well-behaved system for NMR-based relaxation studies and promises to provide a wealth of insight into the nature and role of protein dynamics (entropy) in protein-ligand interactions. In addition, to assess whether the insights learned from the calmodulin system are general we will carry out similar investigations of other protein-ligand interactions. This will include the interaction of the Leukemia Inhibitory Factor and Oncostatin M cytokines with the GP130 receptor. GP130 it as the intersection of a number of important signaling networks of significant clinical interest, particularly in inflammatory bowel disease and cancer. In collaborative work, we will compare crystallographic analysis and molecular dynamics simulations with the experimental results obtained and undertake computational analyses of the thermodynamic origins of binding. Overall these studies promise to reveal novel insights into the physical origins of protein motion, their biological significance and will begin to illuminate the potential for their exploitation in the context of pseudo-allosteric drug inhibitors.