Kraulis P J
Center for Structural Biochemistry, Karolinska Institutet, Huddinge, Sweden.
J Mol Biol. 1994 Nov 4;243(4):696-718. doi: 10.1016/0022-2836(94)90042-6.
The sequence-specific assignment of resonances is considered to be a requirement for the determination of the three-dimensional (3D) structure of a protein in solution by nuclear magnetic resonance methods. The main source of structural information is the nuclear Overhauser effect spectroscopy (NOESY) spectrum, which contains information about spatially close pairs of protons. Currently, various J-correlated spectra must be recorded in order to obtain the sequence-specific assignments necessary to interpret the NOESY spectra. In this work, a novel procedure to determine the 3D structure and the sequence-specific assignments of a protein using only data from 13C and 15N-separated multidimensional NOESY spectra is described. No information from J-correlated spectra is required. The algorithm is called ANSRS (Assignment of NOESY Spectra in Real Space) and is based on an inversion of the traditional strategy. A 3D real-space structure of detected, but unassigned, 1H spins is calculated from the nuclear Overhauser effect (NOE) distance restraints using a dynamical simulated annealing procedure. The sequence-specific assignments are then determined by searching among the 1H spins in the 3D real-space structure for plausible residue assignments. The search uses a Monte Carlo simulated annealing algorithm based on assignment probabilities derived from the 1H, 15N and 13C chemical shifts, various spatial constraints, and the known sequence of the protein. The procedure has been tested on semi-synthetic data sets comprising published experimental chemical shifts and NOE distance restraints derived from the known 3D structures of the two proteins GAL4 (residues 9 to 41) and bovine pancreatic trypsin inhibitor. The ANSRS procedure was able to determine the sequence-specific assignments for more than 95% of the spins, and was fairly robust with respect to missing NOE data. The potential of the ANSRS approach with respect to automated assignment, reduction of the number of NMR spectra required for a structure determination, assignment of homologous and mutant proteins, and the possibility of analysing spectra recorded at high pH is discussed.
共振的序列特异性归属被认为是通过核磁共振方法测定溶液中蛋白质三维（3D）结构的必要条件。结构信息的主要来源是核Overhauser效应光谱（NOESY）谱，它包含有关空间上接近的质子对的信息。目前，必须记录各种J相关谱，以获得解释NOESY谱所需的序列特异性归属。在这项工作中，描述了一种仅使用来自13C和15N分离的多维NOESY谱的数据来确定蛋白质3D结构和序列特异性归属的新方法。不需要来自J相关谱的信息。该算法称为ANSRS（实空间中NOESY谱的归属），基于传统策略的反转。使用动态模拟退火程序，根据核Overhauser效应（NOE）距离约束计算检测到但未归属的1H自旋的3D实空间结构。然后通过在3D实空间结构中的1H自旋中搜索合理的残基归属来确定序列特异性归属。该搜索使用基于从1H、15N和13C化学位移、各种空间约束以及蛋白质的已知序列导出的归属概率的蒙特卡罗模拟退火算法。该方法已在半合成数据集上进行了测试，该数据集包含已发表的实验化学位移和从两种蛋白质GAL4（残基9至41）和牛胰蛋白酶抑制剂的已知3D结构导出的NOE距离约束。ANSRS方法能够确定超过95%的自旋的序列特异性归属，并且对于缺失的NOE数据相当稳健。讨论了ANSRS方法在自动归属、减少结构测定所需的NMR谱数量、同源和突变蛋白质的归属以及分析在高pH下记录的谱的可能性方面的潜力。
