Direct NMR Methods for Protein Structures and Assignment

描述(申请人提供)：在蛋白质的核磁共振研究中，对单个原子的核自旋共振的分配是所有后续生物医学应用的先决条件，例如研究配体结合、蛋白质-DNA相互作用和动力学。即使当蛋白质的3D X射线结构可用时，共振分配也是最耗时和最费力的步骤之一。提出了一种新的策略来寻找共振分配，从而优化利用X射线结构、剩余偶极耦合测量和化学位移。通过这种方式，核磁共振、X射线结晶学和量子化学的互补优势得到了协同使用。与标准分配协议相比，不需要关于顺序连通性的核磁共振信息。分配问题以加权匹配问题的数学形式表示，该加权匹配问题可以使用计算效率高的组合优化算法来解决。分配提供的化学位移信息可直接用于各种核磁共振应用，包括有助于表征结合部位、强度和专一性的配体结合研究，这将有助于合理指导药物设计。这项拟议的工作将使大量蛋白质的结构存放在蛋白质数据库(PDB)中，以便进行详细的生物医学核磁共振研究。 结构基因组学的目标是从其三维结构中推导出蛋白质的功能，它需要快速的结构确定方法。对于不能通过X射线结晶学确定结构的蛋白质，提出了一种利用核磁共振剩余偶极耦合信息、不完全序列连接性和建模技术同时确定结构和共振归属的有效方法。该方法通过直接建立与剩余偶极偶联、氨基酸类型特定化学位移和稀疏顺序主干连接性信息一致的3D蛋白质片段文库，避免了缓慢的分配步骤。单个碎片具有良好的分辨率，并且它们不偏向于沉积在PDB中的已知结构。然后，通过使用数据库导出的惩罚函数的高性能计算方法，将蛋白质片段组装成完整的3D蛋白质结构。与标准的核磁共振方法相比，该方法不依赖于NOESY派生的距离约束。该方法的速度将显著快于标准的核磁共振方法，将在模型蛋白质上进行测试和改进，然后应用于生物相关蛋白质。

DESCRIPTION (provided by applicant): In nuclear magnetic resonance (NMR) studies of proteins, the assignment of nuclear spin resonances to individual atoms is a prerequisite for all subsequent biomedical applications, such as studies of ligand binding, protein-DNA interactions, and dynamics. Resonance assignment is one of the most time consuming and labor intensive steps even when the 3D xray structure of the protein is available. A new strategy is proposed to find resonance assignments that makes optimal use of the x-ray structure, residual dipolar coupling measurements, and chemical shifts. In this way, the complementary strengths of NMR, x-ray crystallography, and quantum chemistry are synergetically used. In contrast to standard assignment protocols, no NMR information about sequential connectivities is required. The assignment problem is mathematically formulated in terms of a weighted matching problem that can be solved using a computationally efficient combinatorial optimization algorithm. The chemical shift information provided by the assignment can be directly used for a wide variety of NMR applications including ligand binding studies that help to characterize binding sites, strengths, and specificity that will help to rationally guide drug design. The proposed work will make a large number of proteins, whose structure is deposited in the protein database (PDB), amenable to detailed biomedical NMR investigations. Structural genomics, which aims at the derivation of protein function from its 3D structure, requires rapid structure determination methods. For proteins whose structure is not determined by x-ray crystallography, an efficient method is proposed for the simultaneous structure determination and resonance assignment using NMR residual dipolar coupling information, incomplete sequential connnectivities, and modeling techniques. The method avoids the slow assignment step by directly building a library of 3D protein fragments that are consistent with residual dipolar couplings, amino-acid type specific chemical shifts, and sparse sequential backbone connectivity information. The individual fragments have good resolution and they are not biased towards known structures deposited in the PDB. The protein fragments are then assembled to complete 3D protein structures by high-performance computional methods using database derived penalty functions. In contrast to standard NMR methods, this approach does not rely on NOESY-derived distance constraints. The method, which promises a significant speed-up over standard NMR methods, will be tested and refined on model proteins and then applied to biologically relevant proteins.