通过顺磁 NMR 光谱法分析蛋白质动力学的镧系元素标记验证。
Validation of a lanthanide tag for the analysis of protein dynamics by paramagnetic NMR spectroscopy.
机构信息
Leiden University, Institute of Chemistry, Gorlaeus Laboratories, P.O. Box 9502, 2300 RA Leiden, The Netherlands.
出版信息
J Am Chem Soc. 2010 Jul 28;132(29):9952-3. doi: 10.1021/ja909508r.
Paramagnetic lanthanide tags potentially can enhance the effects of microsecond to millisecond dynamics in proteins on NMR signals and provide structural information on lowly populated states encoded in the pseudocontact shifts. We have investigated the microsecond to millisecond mobility of a two-point attached lanthanide tag, CLaNP-5, using paramagnetic (1)H CPMG relaxation dispersion methods. CLaNP-5 loaded with Lu(3+), Yb(3+), or Tm(3+) was attached to three sites on the surface of two proteins, pseudoazurin and cytochrome c. The paramagnetic center causes large relaxation dispersion effects for two attachment sites, suggesting that local dynamics of the protein at the attachment site causes mobility of the paramagnetic center. At one site the relaxation dispersions are small and limited to the immediate environment of the tag. It is concluded that paramagnetic relaxation dispersion could represent a sensitive method to probe protein dynamics. However, the selection of a rigid attachment site is of critical importance.
顺磁镧系标签有可能增强蛋白质中微秒到毫秒动力学对 NMR 信号的影响，并提供伪接触位移中编码的低丰度状态的结构信息。我们使用顺磁 (1)H CPMG 弛豫分散方法研究了两点连接的镧系标签 CLaNP-5 的微秒到毫秒迁移率。负载 Lu(3+)、Yb(3+)或 Tm(3+)的 CLaNP-5 连接到两个蛋白质（假变青菌和细胞色素 c）表面的三个位点上。顺磁中心导致两个连接位点的大弛豫分散效应，表明连接位点处蛋白质的局部动力学导致顺磁中心的迁移。在一个位点上，弛豫分散较小，仅限于标签的直接环境。可以得出结论，顺磁弛豫分散可能是一种灵敏的探测蛋白质动力学的方法。然而，刚性连接位点的选择至关重要。
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