Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892-0520, USA.
J Magn Reson. 2010 Aug;205(2):304-14. doi: 10.1016/j.jmr.2010.05.013. Epub 2010 May 25.
We describe a computational approach to sequential resonance assignment in solid state NMR studies of uniformly (15)N,(13)C-labeled proteins with magic-angle spinning. As input, the algorithm uses only the protein sequence and lists of (15)N/(13)C(alpha) crosspeaks from 2D NCACX and NCOCX spectra that include possible residue-type assignments of each crosspeak. Assignment of crosspeaks to specific residues is carried out by a Monte Carlo/simulated annealing algorithm, implemented in the program MC_ASSIGN1. The algorithm tolerates substantial ambiguity in residue-type assignments and coexistence of visible and invisible segments in the protein sequence. We use MC_ASSIGN1 and our own 2D spectra to replicate and extend the sequential assignments for uniformly-labeled HET-s(218-289) fibrils previously determined manually by Siemer et al. (J. Biomol. NMR, 34 (2006) 75-87) from a more extensive set of 2D and 3D spectra. Accurate assignments by MC_ASSIGN1 do not require data that are of exceptionally high quality. Use of MC_ASSIGN1 (and its extensions to other types of 2D and 3D data) is likely to alleviate many of the difficulties and uncertainties associated with manual resonance assignments in solid state NMR studies of uniformly labeled proteins, where spectral resolution and signal-to-noise are often sub-optimal.
我们描述了一种计算方法，用于对具有魔角旋转的均匀（15）N、（13）C 标记的蛋白质的固态 NMR 研究中的序列共振分配。作为输入，该算法仅使用蛋白质序列和来自 2D NCACX 和 NCOCX 谱的（15）N/（13）C（alpha）交叉峰列表，其中包括每个交叉峰的可能残基类型分配。交叉峰的分配是通过蒙特卡罗/模拟退火算法完成的，该算法在程序 MC_ASSIGN1 中实现。该算法可以容忍残基类型分配的大量歧义以及蛋白质序列中可见和不可见片段的共存。我们使用 MC_ASSIGN1 和我们自己的 2D 光谱来复制和扩展先前由 Siemer 等人（J. Biomol. NMR，34（2006）75-87）通过更广泛的 2D 和 3D 光谱手动确定的均匀标记 HET-s（218-289）纤维的序列分配。MC_ASSIGN1 的准确分配不需要质量特别高的数据。在固态 NMR 研究中，使用 MC_ASSIGN1（及其对其他类型的 2D 和 3D 数据的扩展）可能会减轻与均匀标记蛋白质的手动共振分配相关的许多困难和不确定性，在这些研究中，光谱分辨率和信噪比通常不理想。
