Liu A, Riek R, Wider G, von Schroetter C, Zahn R, Wüthrich K
Institut für Molekularbiologie und Biophysik, Eidgenössische Technische Hochschule Hönggerberg, Zürich, Switzerland.
J Biomol NMR. 2000 Feb;16(2):127-38. doi: 10.1023/a:1008305022907.
A combination of three heteronuclear three-dimensional NMR experiments tailored for sequential resonance assignments in uniformly 15N, 13C-labeled flexible polypeptide chains is described. The 3D (H)N(CO-TOCSY)NH, 3D (H)CA(CO-TOCSY)NH and 3D (H)CBCA(CO-TOCSY)NH schemes make use of the favorable 15N chemical shift dispersion in unfolded polypeptides, exploit the slow transverse 15N relaxation rates of unfolded polypeptides in high resolution constant-time [1H, 15N]-correlation experiments, and use carbonyl carbon homonuclear isotropic mixing to transfer magnetization sequentially along the amino acid sequence. Practical applications are demonstrated with the 100-residue flexible tail of the recombinant human prion protein, making use of spectral resolution up to 0.6 Hz in the 15N dimension, simultaneous correlation with the two adjacent amino acid residues to overcome problems associated with spectral overlap, and the potential of the presently described experiments to establish nearest-neighbor correlations across proline residues in the amino acid sequence.
本文描述了三种异核三维核磁共振实验的组合，这些实验专为对均匀 15N、13C 标记的柔性多肽链进行序列共振归属而设计。3D (H)N(CO-TOCSY)NH、3D (H)CA(CO-TOCSY)NH 和 3D (H)CBCA(CO-TOCSY)NH 方案利用了未折叠多肽中有利的 15N 化学位移分散，在高分辨率恒时 [1H, 15N] 相关实验中利用未折叠多肽缓慢的横向 15N 弛豫速率，并使用羰基碳同核各向同性混合来沿着氨基酸序列顺序转移磁化。通过重组人朊病毒蛋白的 100 个残基的柔性尾部展示了实际应用，利用了 15N 维度高达 0.6 Hz 的光谱分辨率、与两个相邻氨基酸残基的同时相关性来克服与光谱重叠相关的问题，以及当前所述实验在氨基酸序列中建立跨越脯氨酸残基的近邻相关性的潜力。
