Koharudin Leonardus M I, Bonvin Alexandre M J J, Kaptein Robert, Boelens Rolf
Bijvoet Center for Biomolecular Research, NMR Spectroscopy, Utrecht University, Padualaan 8, 3584 CH, Utrecht, The Netherlands.
J Magn Reson. 2003 Aug;163(2):228-35. doi: 10.1016/s1090-7807(03)00149-6.
The high sensitivity of modern NMR instrumentation, in combination with full deuteration, enabled the measurement of long-range NOEs between amide protons in a fully deuterated protein corresponding to distances up to 8A. These are beyond the limit normally observed in protonated samples. Such long-distance NOEs could be observed using long mixing times, which became possible due to reduced spin diffusion and T1 relaxation of the amide protons in the fully deuterated sample. This information was used in combination with secondary structure restraints derived from secondary chemical shifts for structure calculations. With these backbone amide proton NOEs only, a unique fold could be obtained with positional root mean square deviations from the average of 1.30 and 2.25 A for backbone and heavy atoms, respectively. Despite the low density of restraints, no mirror image problems were observed. Addition of sidechain NOE information increased the precision of the ensemble and in particular of the core packing. The structures obtained in this way were close to the published crystal structure. NOE completeness analysis revealed that the cumulative completeness is still more than 80% for an 8.0 A cut-off distance.
现代核磁共振仪器的高灵敏度，结合完全氘代，使得能够测量完全氘代蛋白质中酰胺质子之间对应距离达8埃的长程核Overhauser效应（NOE）。这些距离超出了质子化样品中通常观察到的极限。使用长混合时间可以观察到这种长距离NOE，这在完全氘代样品中由于酰胺质子的自旋扩散和T1弛豫降低而成为可能。该信息与源自二级化学位移的二级结构约束相结合用于结构计算。仅利用这些主链酰胺质子NOE，就可以得到一个独特的折叠结构，主链和重原子相对于平均值的位置均方根偏差分别为1.30埃和2.25埃。尽管约束密度较低，但未观察到镜像问题。添加侧链NOE信息提高了整体结构的精度，特别是核心堆积的精度。通过这种方式获得的结构与已发表的晶体结构接近。NOE完整性分析表明，对于8.0埃的截止距离，累积完整性仍超过80%。
