Gu Z, Opella S J
Department of Chemistry, University of Pennsylvania, Philadelphia, Pennsylvania, 19104, USA.
J Magn Reson. 1999 Jun;138(2):193-8. doi: 10.1006/jmre.1999.1709.
Triple-resonance experiments capable of correlating directly bonded and proximate carbon and nitrogen backbone sites of uniformly 13C- and 15N-labeled peptides in stationary oriented samples are described. The pulse sequences integrate cross-polarization from 1H to 13C and from 13C to 15N with flip-flop (phase and frequency switched) Lee-Goldburg irradiation for both 13C homonuclear decoupling and 1H-15N spin exchange at the magic angle. Because heteronuclear decoupling is applied throughout, the three-dimensional pulse sequence yields 13C shift/1H-15N coupling/15N shift correlation spectra with single-line resonances in all three frequency dimensions. Not only do the three-dimensional spectra correlate 13C and 15N resonances, they are well resolved due to the three independent frequency dimensions, and they can provide up to four orientationally dependent frequencies as input for structure determination. These experiments have the potential to make sequential backbone resonance assignments in uniformly 13C- and 15N-labeled proteins.
本文描述了一种三共振实验，该实验能够在固定取向样品中，将均匀 13C 和 15N 标记的肽段中直接相连以及相邻的碳和氮主链位点进行关联。脉冲序列将从 1H 到 13C 以及从 13C 到 15N 的交叉极化与用于 13C 同核去耦和魔角处 1H - 15N 自旋交换的触发器（相位和频率切换）Lee - Goldburg 照射相结合。由于全程应用了异核去耦，三维脉冲序列在所有三个频率维度上都产生具有单线共振的 13C 位移/1H - 15N 耦合/15N 位移相关光谱。三维光谱不仅能关联 13C 和 15N 共振，由于三个独立的频率维度，它们得到了很好的分辨，并且它们可以提供多达四个取向依赖频率作为结构测定的输入。这些实验有潜力对均匀 13C 和 15N 标记的蛋白质进行连续主链共振归属。
