Probing slow time scale dynamics at methyl-containing side chains in proteins by relaxation dispersion NMR measurements: Application to methionine residues in a cavity mutant of T4 lysozyme
Probing slow time scale dynamics at methyl-containing side chains in proteins by relaxation dispersion NMR measurements: Application to methionine residues in a cavity mutant of T4 lysozyme
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DOI：
10.1021/ja004179p
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发表时间：
2001-05-16
影响因子：
15
通讯作者：
Kay, LE
中科院分区：
文献类型：
--
作者：
Skrynnikov, NR;Mulder, FAA;Kay, LE
提出了一种基于弛豫色散的NMR实验，用于测量和定量蛋白质甲基侧链位置的mus-ms动力学过程。实验测量的交换贡献的C-13线宽的甲基使用恒定时间的CPMG计划。偶极-偶极和偶极-CSA相互作用之间的交叉相关自旋弛豫的影响以及在实验过程中负责混合磁化模式的标量耦合的影响已经在理论上和通过模拟详细研究。结果表明，甲基自旋系统的复杂的弛豫特性不复杂的提取准确的交换参数，只要小心，以确保适当的磁化模式在中间的恒定时间的CPMG期间互换。甲硫氨酸残基在Leu 99 Ala取代T4溶菌酶的松弛分散配置文件的测量的应用。所有的甲硫氨酸残基都对20 ℃下速率为1200 s（-1）的交换事件敏感，这可能与疏水配体能够快速结合到该突变体中存在的空腔的过程有关。
A relaxation dispersion-based NMR experiment is presented for the measurement and quantitation of mus-ms dynamic processes at methyl side-chain positions in proteins. The experiment measures the exchange contribution to the C-13 line widths of methyl groups using a constant-time CPMG scheme. The effects of cross-correlated spin relaxation between dipole-dipole and dipole-CSA interactions as well as the effects of scalar coupling responsible for mixing of magnetization modes during the course of the experiment have been investigated in detail both theoretically and through simulations. It is shown that the complex relaxation properties of the methyl spin system do not complicate extraction of accurate exchange parameters as long as care is taken to ensure that appropriate magnetization modes are interchanged in the middle of the constant-time CPMG period. An application involving the measurement of relaxation dispersion profiles of methionine residues in a Leu99Ala substitution of T4 lysozyme is presented. All of the methionine residues are sensitive to an exchange event with a rate on the order of 1200 s(-1) at 20 degreesC that may be linked to a process in which hydrophobic ligands are able to rapidly bind to the cavity that is present in this mutant.