Detection of dynamic water molecules in a microcrystalline sample of the SH3 domain of a-spectrin by MAS solid-state NMR
Detection of dynamic water molecules in a microcrystalline sample of the SH3 domain of a-spectrin by MAS solid-state NMR
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DOI：
10.1007/s10858-005-1718-z
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发表时间：
2005-04-01
影响因子：
2.7
通讯作者：
Reif, B
中科院分区：
文献类型：
--
作者：
Chevelkov, V;Faelber, K;Reif, B
水分子是蛋白质稳定性的主要决定因素，对于理解蛋白质 - 蛋白质相互作用非常重要。我们介绍了两个实验，第一个实验能够测量单个酰胺质子的有效T - 2衰减率，第二个实验能够测量以自旋扩散作为混合元素从H₂O到H - N的选择性转移的磁化建立率。这些实验是在一个均匀的H - 2、N - 15标记的微晶SH3结构域样品上进行的，其中可交换的氘原子被质子反向取代。为了评估核磁共振实验数据，使用与固态核磁共振样品相同的结晶方案确定了该蛋白质的X射线结构。核磁共振实验数据与根据从蛋白质的X射线结构中提取的H₂O - H - N距离计算出的偶极耦合相关。我们发现H - N的T - 2 O - 衰减率和H(₂) - HN的建立率对检测到的水分子相对于蛋白质的距离和动力学敏感。我们表明，通过解释报告酰胺质子的自旋动力学，可以在固态下获得有关内部水分子的定位和动力学的定性信息。
Water molecules are a major determinant of protein stability and are important for understanding protein-protein interactions. We present two experiments which allow to measure first the effective T-2 decay rate of individual amide proton, and second the magnetization build-up rates for a selective transfer from H2O to H-N using spin diffusion as a mixing element. The experiments are demonstrated for a uniformly H-2, N-15 labeled sample of a microcrystalline SH3 domain in which exchangeable deuterons were back-substituted with protons. In order to evaluate the NMR experimental data, as X-ray structure of the protein was determined using the same crystallization protocol as for the solid-state NMR sample. The NMR experimental data are correlated with the dipolar couplings calculated from H2O-H-N distances which were extracted from the X-ray structure of the protein. We find that the H-N T-2 O-decay rates and H(2)-HN build-up rates are sensitive to distance and dynamics of the detected water molecules with respect to the protein. We show that qualitative information about localization and dynamics of internal water molecules can be obtained in the solid-state by interpretation of the spin dynamics of a reporter amide proton.