Multiscale Modeling of Protein Kinase Structure, Catalysis and Allostery

蛋白激酶的结构、催化和变性的多尺度模拟 项目总结/摘要 这项研究的长期目标是促进我们对催化和调节机制的理解 复杂的酶系统和蛋白质动力学在酶功能中的作用。蛋白激酶（PK）， 这项研究的重点，是一个有吸引力的系统，因为它涉及大规模的构象， 变化和酶催化。此外，由于其病理意义，了解PK的 分子机制在激酶研究中具有根本的重要性，也可能为激酶的研究提供新的见解。 开发针对激酶的改良疗法。根据我们最近的研究， 和酶动力学数据，是PK的催化活性与其调节功能密切相关。 因此，调节活性的任何变化都会影响激酶的催化活性，这通过以下方式发生： 潜在的蛋白质动力学的变构调节，并一起控制PK的总体活性。此相反 传统观点认为，非活性到活性的构象变化是调节 激酶活性我们的目标是在这个补助金是检查这两个截然不同的观点调节激酶 通过对两种重要激酶，胰岛素受体激酶（IRK）和腺苷酸激酶（AdK）的平行研究， 其在细胞内稳态中起关键作用，并阐明其完整的分子机制。这些 目标将通过定量模拟它们的构象变化、配体结合和 关键功能状态下的催化，包括完全活性和非活性状态。拟议的研究涉及 结合量子、分子和统计力学的新的多尺度模拟方法的发展 方法及其扩展，以允许快速和准确地确定激酶机制。我们的具体 目标是：（1）发展有效的多尺度模拟方法，结合从头算/密度 泛函理论（AI/DFT）和半经验（SE）QM/MM方法与弦模拟方法， CHARMM，通过高级并行化和加速器算法以及特定于反应的 参数化，并开发有效的炼金术自由能模拟方法，克服 现有方法的局限性; 2）阐明IRK催化和构象变化的机制 以及其内在蛋白质运动与IRK催化作用之间的关系; 3）测定IRK的催化活性。 AdK的作用机制以及活性位点残基在控制活性位点和整体蛋白质动力学中的作用 和AdK的整体活性。完成拟议的研究将加深对机制的认识 这些激酶和蛋白质动力学在其催化中的作用。计算的实验验证 结果也将通过表征其动力学和结构参数，通过与 实验组最后，所发展的理论方法是通用的，可以很容易地应用于 许多涉及构象变化和催化的酶系统，如ATP/GTP酶和 各种马达蛋白。

Multiscale Modeling of Protein Kinase Structure, Catalysis and Allostery PROJECT SUMMARY/ABSTRACT The long-term goal of this research is to advance our understanding of the catalytic and regulatory mechanisms of complex enzymatic systems and the roles of protein dynamics in enzyme function. Protein kinase (PK), the focus of this study, is an attractive system for this purpose, because it involves both large-scale conformational change and enzymatic catalysis. Moreover, because of its pathological significance, understanding PK’s molecular mechanism is of fundamental importance in kinase research and also may provide new insights into the development of improved therapies against kinases. Our central hypothesis, based on our recent studies and enzyme kinetic data, is that the catalytic activity of PK is closely associated with its regulatory function. Therefore, any change in regulatory activity affects the catalytic activity of the kinase, which occurs through allosteric modulation of underlying protein dynamics, and together control overall activity of PK. This contrasts with the conventional view that the inactive-to-active conformational change is the main mechanism for regulating kinase activity. Our objective in this grant is to examine these two contrasting views on the regulation of kinase activity by the parallel study of two important kinases, insulin receptor kinase (IRK) and adenylate kinase (AdK), which play critical roles in cell homeostasis, and elucidate their complete molecular mechanisms. These objectives will be accomplished through quantitative modeling of their conformational change, ligand binding and catalysis in key functional states, including the fully active and inactive states. The proposed research involves development of new multiscale simulation methods combining quantum, molecular and statistical mechanical methods and their extension to permit rapid and accurate determination of kinase mechanisms. Our specific AIMs are: (1) the development of effective multiscale simulation methods integrating the ab initio/density functional theory (AI/DFT) and semiempirical (SE) QM/MM methods with the string simulation methods in CHARMM, their acceleration through advanced parallelization and accelerator algorithms and reaction-specific parameterizations, and development of efficient alchemical free energy simulation methods overcoming the limitations of existing methods; 2) elucidation of the mechanisms of IRK catalysis and conformational change and the connection between its intrinsic protein motions and catalysis of IRK; and 3) determination of the catalytic mechanism of AdK and the role of active site residues in controlling the active site and global protein dynamics and the overall activity of AdK. The completion of the proposed study will deepen the mechanistic understanding of these kinases and the role of protein dynamics in their catalysis. Experimental verification of computational results will also be made by characterization of their kinetic and structural parameters via collaboration with an experimental group. Finally, the theoretical methods developed are general and can be applied readily to numerous enzymatic systems involving conformational changes and catalysis, such as ATP/GTPases and various motor proteins.