Akhmetzyanov D, Plackmeyer J, Endeward B, Denysenkov V, Prisner T F
Goethe University Frankfurt am Main, Institute of Physical and Theoretical Chemistry and Center for Biomolecular Magnetic Resonance, Max von Laue Str. 7, 60438 Frankfurt am Main, Germany.
Phys Chem Chem Phys. 2015 Mar 14;17(10):6760-6. doi: 10.1039/c4cp05362a.
Pulsed Electron-Electron Double Resonance (PELDOR) has attracted considerable attention for biomolecular applications, as it affords precise measurements of distances between pairs of spin labels in the range of 1.5-8 nm. Usually nitroxide moieties incorporated by site-directed spin labelling with cysteine residues are used as spin probes in protein systems. Recently, naturally occurring cofactors and metal ions have also been explored as paramagnetic spin species for such measurements. In this work we investigate the performance of PELDOR between a nitroxide spin label and a high-spin Mn(2+) ion in a synthetic model compound at Q-band (34 GHz) and G-band (180 GHz). We demonstrate that the distances obtained with high-frequency PELDOR are in good agreement with structural predictions. At Q-band frequencies experiments have been performed by probing either the high-spin Mn(2+) ion or the nitroxide spin label. At G-band frequencies we have been able to detect changes in the dipolar oscillation frequency, depending on the pump-probe positions across the g-tensor resolved nitroxide EPR spectrum. These changes result from the restricted mobility of the nitroxide spin label in the model compound. Our results demonstrate that the high-spin Mn(2+) ion can be used for precise distance measurements and open the doors for many biological applications, as naturally occurring Mg(2+) sites can be readily exchanged for Mn(2+).
脉冲电子-电子双共振（PELDOR）在生物分子应用中引起了广泛关注，因为它能够精确测量1.5至8纳米范围内自旋标记对之间的距离。在蛋白质系统中，通常将通过半胱氨酸残基定点自旋标记引入的氮氧化物部分用作自旋探针。最近，天然存在的辅因子和金属离子也被探索用作此类测量的顺磁自旋物种。在这项工作中，我们研究了在合成模型化合物中，在Q波段（34吉赫兹）和G波段（180吉赫兹）下，氮氧化物自旋标记与高自旋Mn(2+)离子之间的PELDOR性能。我们证明，高频PELDOR获得的距离与结构预测结果吻合良好。在Q波段频率下，通过探测高自旋Mn(2+)离子或氮氧化物自旋标记进行了实验。在G波段频率下，我们能够检测到偶极振荡频率的变化，这取决于在g张量分辨的氮氧化物EPR谱上泵浦-探测位置。这些变化是由模型化合物中氮氧化物自旋标记的受限流动性引起的。我们的结果表明，高自旋Mn(2+)离子可用于精确的距离测量，并为许多生物应用打开了大门，因为天然存在的Mg(2+)位点可以很容易地被Mn(2+)取代。
