Solid-state NMR enhanced by dynamic nuclear polarization as a novel tool for ribosome structural biology
Solid-state NMR enhanced by dynamic nuclear polarization as a novel tool for ribosome structural biology
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DOI：
10.1007/s10858-013-9721-2
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发表时间：
2013-06-01
影响因子：
2.7
通讯作者：
Bodenhausen, Geoffrey
中科院分区：
文献类型：
--
作者：
Gelis, Ioannis;Vitzthum, Veronika;Bodenhausen, Geoffrey
核磁共振 (NMR) 对大分子复合物研究的影响取决于灵敏度和分辨率的提高。如果样品制备经过优化以保持光谱分辨率，固态动态核极化 (DNP) 可以提高灵敏度。对于几纳摩尔的完整核糖体和 800 kDa 核糖体复合物，我们证明 DNP 和魔角旋转 NMR (MAS-NMR) 的结合可以克服当前的灵敏度限制，从而可以记录同核和异核 C-13 和 N-15 NMR 相关光谱。直接沉淀并冷冻到 NMR 转子中的核糖体颗粒可产生大约 25 倍的 DNP 信号增强，并且光谱呈现窄线宽，适合获取位点特异性信息。我们预计相同的方法也适用于其他高分子量复合物。
The impact of Nuclear Magnetic Resonance (NMR) on studies of large macromolecular complexes hinges on improvements in sensitivity and resolution. Dynamic nuclear polarization (DNP) in the solid state can offer improved sensitivity, provided sample preparation is optimized to preserve spectral resolution. For a few nanomoles of intact ribosomes and an 800 kDa ribosomal complex we demonstrate that the combination of DNP and magic-angle spinning NMR (MAS-NMR) allows one to overcome current sensitivity limitations so that homo- and heteronuclear C-13 and N-15 NMR correlation spectra can be recorded. Ribosome particles, directly pelleted and frozen into an NMR rotor, yield DNP signal enhancements on the order of similar to 25-fold and spectra that exhibit narrow linewidths, suitable for obtaining site-specific information. We anticipate that the same approach is applicable to other high molecular weight complexes.