Pulse EPR distance measurements to study multimers and multimerisation
Pulse EPR distance measurements to study multimers and multimerisation
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DOI：
10.1080/00268976.2017.1421324
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发表时间：
2018-01-01
影响因子：
1.7
通讯作者：
Bode, Bela E.
中科院分区：
文献类型：
--
作者：
Ackermann, Katrin;Bode, Bela E.
脉冲偶极电子顺磁共振（PD-EPR）已成为结构生物学在纳米尺度上确定距离的有力工具。硬件、方法和数据分析的最新进展已经将范围扩大到复杂的生物系统。PD-EPR可应用于含有低填充构象或显示大的固有灵活性的系统，使它们几乎难以用于冷冻电子显微镜和晶体学。由于获得所有相关构象的高分辨率结构的固有困难，膜蛋白的应用特别令人感兴趣。许多涉及关键细胞功能的药物靶标是多聚体通道或转运蛋白。在这里，用于引入PD-EPR的自旋标记的常见方法导致每个多聚体复合物存在多于两个电子自旋。这需要仔细的实验设计，以克服有害的多自旋效应，并确保在存在多个距离的情况下有足够的距离分辨率。除了获得纯粹的距离，PD-EPR还可以提供多聚化程度的信息，从而研究结合平衡并确定解离常数。[图形]。
Pulse dipolar electron paramagnetic resonance (PD-EPR) has become a powerful tool for structural biology determining distances on the nanometre scale. Recent advances in hardware, methodology, and data analysis have widened the scope to complex biological systems. PD-EPR can be applied to systems containing lowly populated conformers or displaying large intrinsic flexibility, making them all but intractable for cryo-electron microscopy and crystallography. Membrane protein applications are of particular interest due to the intrinsic difficulties for obtaining high-resolution structures of all relevant conformations. Many drug targets involved in critical cell functions are multimeric channels or transporters. Here, common approaches for introducing spin labels for PD-EPR cause the presence of more than two electron spins per multimeric complex. This requires careful experimental design to overcome detrimental multi-spin effects and to secure sufficient distance resolution in presence of multiple distances. In addition to obtaining mere distances, PD-EPR can also provide information on multimerisation degrees allowing to study binding equilibria and to determine dissociation constants.[GRAPHICS].