Kehlet Cindie, Bjerring Morten, Sivertsen Astrid C, Kristensen Torsten, Enghild Jan J, Glaser Steffen J, Khaneja Navin, Nielsen Niels Chr
Center for Insoluble Protein Structures (inSPIN), Interdisciplinary Nanoscience Center (iNANO), University of Aarhus, Langelandsgade 140, DK-8000, Aarhus C, Denmark.
J Magn Reson. 2007 Oct;188(2):216-30. doi: 10.1016/j.jmr.2007.06.011. Epub 2007 Jul 7.
We present novel pulse sequences for magic-angle-spinning solid-state NMR structural studies of (13)C,(15)N-isotope labeled proteins. The pulse sequences have been designed numerically using optimal control procedures and demonstrate superior performance relative to previous methods with respect to sensitivity, robustness to instrumental errors, and band-selective excitation profiles for typical biological solid-state NMR applications. Our study addresses specifically (15)N to (13)C coherence transfers being important elements in spectral assignment protocols for solid-state NMR structural characterization of uniformly (13)C,(15)N-labeled proteins. The pulse sequences are analyzed in detail and their robustness towards spin system and external experimental parameters are illustrated numerically for typical (15)N-(13)C spin systems under high-field solid-state NMR conditions. Experimentally the methods are demonstrated by 1D (15)N-->(13)C coherence transfer experiments, as well as 2D and 3D (15)N,(13)C and (15)N,(13)C,(13)C chemical shift correlation experiments on uniformly (13)C,(15)N-labeled ubiquitin.
我们展示了用于对（13）C、（15）N同位素标记蛋白质进行魔角旋转固态核磁共振结构研究的新型脉冲序列。这些脉冲序列是使用最优控制程序进行数值设计的，并且在灵敏度、对仪器误差的鲁棒性以及典型生物固态核磁共振应用的带选择性激发谱方面相对于先前方法展现出卓越性能。我们的研究特别关注（15）N到（13）C的相干转移，这是对均匀（13）C、（15）N标记蛋白质进行固态核磁共振结构表征的谱峰归属协议中的重要元素。对这些脉冲序列进行了详细分析，并针对高场固态核磁共振条件下典型的（15）N - （13）C自旋系统，通过数值方式说明了它们对自旋系统和外部实验参数的鲁棒性。通过对均匀（13）C、（15）N标记的泛素进行一维（15）N→（13）C相干转移实验以及二维和三维（15）N、（13）C和（15）N、（13）C、（13）C化学位移相关实验，对这些方法进行了实验验证。
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