Generic tags for Mn(ii) and Gd(iii) spin labels for distance measurements in proteins.
Generic tags for Mn(ii) and Gd(iii) spin labels for distance measurements in proteins.
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用于蛋白质距离测量的 Mn(ii) 和 Gd(iii) 自旋标签的通用标签。
DOI：
10.1039/c7cp04311b
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发表时间：
2017-10
期刊：
影响因子：
--
通讯作者：
Goldfarb Daniella
中科院分区：
文献类型：
--
作者：
Yang Yin;Gong Yan-Jun;Litvinov Aleksei;Liu Hong-Kai;Yang Feng;Su Xun-Cheng;Goldfarb Daniella
高亲和力的螯合标记Gd（III）和Mn（II）离子，提供有价值的高分辨率的距离限制生物分子被用作自旋标记的双电子-电子共振（DEER）测量。由于它们不同的EPR相关特性，可以结合两种金属离子并为两种离子提供窄且可预测的距离分布的通用标签的可用性是有吸引力的。在本文中，我们引入了两个顺磁性标签，4PSPyMTA和4PSPyNPDA，它们通过稳定的硫醚键与半胱氨酸残基缀合，形成短的并且取决于金属离子配位模式的与蛋白质的刚性系链。这些标签对Mn（ii）和Gd（iii）离子都表现出高亲和力。针对泛素的三种双半胱氨酸突变体，评估了与Gd（iii）或Mn（ii）复合的4PSPyMTA和4PSPyNPDA标签的DEER性能，并比较了它们产生的Gd（iii）-Gd（iii）和Mn（ii）-Mn（ii）距离分布。泛素-PyMTA和泛素-PyNPDA缀合物的所有三种Gd（iii）复合物产生相似的和预期的距离分布。与此相反，观察到的Mn（ii）类似物的距离分布的最大值和宽度的显着变化。此外，虽然PyNPDA-Gd（iii）和PyNPDA-Mn（ii）提供了相似的距离分布，但对于具有PyMTA的两个突变体观察到明显的差异，Mn（ii）类似物表现出更宽的距离分布和更短的距离。ELDOR（电子-电子双共振）检测的NMR测量揭示了一些分布在Mn（II）的两个标签的蛋白质缀合物的配位环境，但不是自由的标签。与Gd（iii）相比，4PSPyMTA-Mn（ii）产生的更宽的距离分布归因于Mn（ii）离子在PyMTA螯合物内的分布位置，这是由于其较小的尺寸和较低的配位数使得吡啶氮未配位。因此，在距离分辨率方面，4PSPyNPDA可以用作Gd（iii）和Mn（ii）的有效通用标签，而4PSPyMTA仅对Gd（iii）有效。Gd（iii）和Mn（ii）之间的这种比较表明，PyMTA模型化合物可能不能充分预测PyMTA-Mn（ii）作为蛋白质中高分辨率距离测量的标记的性能，因为蛋白质环境可以影响其配位模式。
High-affinity chelating tags for Gd(iii) and Mn(ii) ions that provide valuable high-resolution distance restraints for biomolecules were used as spin labels for double electron-electron resonance (DEER) measurements. The availability of a generic tag that can bind both metal ions and provide a narrow and predictable distance distribution for both ions is attractive owing to their different EPR-related characteristics. Herein we introduced two paramagnetic tags, 4PSPyMTA and 4PSPyNPDA, which are conjugated to cysteine residues through a stable thioether bond, forming a short and, depending on the metal ion coordination mode, a rigid tether with the protein. These tags exhibit high affinity for both Mn(ii) and Gd(iii) ions. The DEER performance of the 4PSPyMTA and 4PSPyNPDA tags, in complex with Gd(iii) or Mn(ii), was evaluated for three double cysteine mutants of ubiquitin, and the Gd(iii)-Gd(iii) and Mn(ii)-Mn(ii) distance distributions they generated were compared. All three Gd(iii) complexes of the ubiquitin-PyMTA and ubiquitin-PyNPDA conjugates produced similar and expected distance distributions. In contrast, significant variations in the maxima and widths of the distance distributions were observed for the Mn(ii) analogs. Furthermore, whereas PyNPDA-Gd(iii) and PyNPDA-Mn(ii) delivered similar distance distributions, appreciable differences were observed for two mutants with PyMTA, with the Mn(ii) analog exhibiting a broader distance distribution and shorter distances. ELDOR (electron-electron double resonance)-detected NMR measurements revealed some distribution in the Mn(ii) coordination environment for the protein conjugates of both tags but not for the free tags. The broader distance distributions generated by 4PSPyMTA-Mn(ii), as compared with Gd(iii), were attributed to the distributed location of the Mn(ii) ion within the PyMTA chelate owing to its smaller size and lower coordination number that leave the pyridine nitrogen uncoordinated. Accordingly, in terms of distance resolution, 4PSPyNPDA can serve as an effective generic tag for Gd(iii) and Mn(ii), whereas 4PSPyMTA is efficient for Gd(iii) only. This comparison between Gd(iii) and Mn(ii) suggests that PyMTA model compounds may not predict sufficiently well the performance of PyMTA-Mn(ii) as a tag for high-resolution distance measurements in proteins because the protein environment can influence its coordination mode.
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影响因子：
2
作者：
R. Dmochowski
通讯作者：
R. Dmochowski
DOI：
10.1016/j.respe.2020.09.004
发表时间：
2020-10-10
期刊：
Revue D'Epidemiologie et De Sante Publique
影响因子：
--
作者：
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通讯作者：
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影响因子：
9.3
作者：
通讯作者：
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影响因子：
5.9
作者：
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通讯作者：
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