New Methods and Applications for the Dynamic Characterization of Proteins by the Combination of NMR Spectroscopy and Computer Simulations

本研究的目的是利用核磁共振（NMR）和计算方法研究蛋白质动力学。核磁共振弛豫参数现在通常用于测量蛋白质主链，但较少用于测量蛋白质侧链。虽然侧链弛豫研究主要集中在甲基弛豫上，但大多数侧链碳原子属于亚甲基。如果没有部分氘化，偶极-偶极相互关系通常会使它们的弛豫行为和解释变得困难。开发了新的弛豫实验，有效地抑制了全碳标记蛋白中的偶极子-偶极子交叉相关弛豫。弛豫数据将用重定向特征模动力学（RED）分析来解释，该分析将核磁共振弛豫数据与分子动力学模拟相结合，以研究ns和sub-ns时间尺度范围内的运动相关效应。直到最近，对于没有表现出交换展宽的自旋，蛋白质在数百纳秒到毫秒范围内较慢的时间尺度动力学一直很难评估。残余偶极耦合的出现现在为全面的实验研究打开了这个时间窗口。通过应用和改进最近引入的无模型方法，该方法在各种具有不同取向张量的取向介质中使用偶极耦合数据，将研究蛋白质主链和侧链在这些较慢时间尺度上的蛋白质动力学。动力学信息将以平均球面谐波的形式检索，这些谐波定义了有序参数以及运动的各向异性。现实的分析运动模型将被开发来表征局部蛋白质片段的运动，以及涉及氨基酸基团和整个二级结构元件的集体运动。蛋白质的结构动力学在生物化学过程中起着重要作用。在原子水平上关于蛋白质动力学的详细信息对于理解它们的功能是至关重要的。本研究的总体目标是探索和发展新的实验和计算方法，以在广泛不同的时间尺度上全面描述复杂的蛋白质动力学，并证明它们在生物学相关系统中的适用性。

The objective of this research is to study protein dynamics by nuclear magnetic resonance (NMR) and computational approaches. NMR relaxation parameters are now routinely measured for the protein backbone, but less so for protein side chains. While side-chain relaxation studies have mainly focused on methyl group relaxation, the majority of side-chain carbon atoms belong to methylene moieties. If not partially deuterated, dipole-dipole cross correlations generally make their relaxation behavior and their interpretation difficult. New relaxation experiments are developed that efficiently suppress dipole-dipole cross-correlated relaxation in fully carbon-labeled proteins. The relaxation data will be interpreted in terms of reorientational eigenmode dynamics (RED) analysis that integrates NMR relaxation data with molecular dynamics simulations for studying motional correlation effects in the ns and sub-ns time scale range. Until recently, slower time-scale dynamics of proteins in the hundreds of ns to ms range have been very hard to assess for spins that do not show exchange broadening. The advent of residual dipolar couplings is now opening up this time window for comprehensive experimental investigations. By applying and improving the recently introduced model-free approach that uses dipolar coupling data in a variety of alignment media with different alignment tensors, protein dynamics on these slower time scales will be investigated for the protein backbone as well as for the side chains. The dynamics information will be retrieved in the form of averaged spherical harmonics that define an order parameter as well as the anisotropy of motion. Realistic analytical motional models will be developed to characterize motions of local protein fragments as well as collective motions involving amino-acid groups and whole secondary structural elements. Structural dynamics of proteins play an important role in biochemical processes. Detailed information on protein dynamics at an atomic level is of fundamental importance for understanding their function. The overall goal of this research is the exploration and development of new experimental and computational methods for a comprehensive description of complex protein dynamics on widely different time scales and to demonstrate their applicability to biologically relevant systems.