Department of Chemistry, North Carolina State University, 2620 Yarbrough Drive, Raleigh, NC 27695, USA.
Department of Chemistry, North Carolina State University, 2620 Yarbrough Drive, Raleigh, NC 27695, USA.
J Magn Reson. 2018 Aug;293:9-18. doi: 10.1016/j.jmr.2018.05.007. Epub 2018 May 17.
EPR pulsed dipolar spectroscopy (PDS) is indispensable for measurements of nm-scale distances between electronic spins in biological and other systems. While several useful modifications and pulse sequences for PDS have been developed in recent years, DEER experiments utilizing pump and observer pulses at two different frequencies remain the most popular for practical applications. One of the major drawbacks of all the available DEER approaches is the presence of a significant unmodulated fraction in the detected signal that arises from an incomplete inversion of the coupled spins by the pump pulse. The latter fraction is perceived as one of the major sources of error for the reconstructed distance distributions. We describe an alternative detection scheme - a Refocused Out-Of-Phase DEER (ROOPh-DEER) - to acquire only the modulated fraction of the dipolar DEER signal. When Zeeman splitting is small compared to the temperature, the out-of-phase magnetization components cancel each other and are not observed in 4-pulse DEER experiment. In ROOPh-DEER these components are refocused by an additional pump pulse while the in-phase component containing an unmodulated background is filtered out by a pulse at the observed frequency applied right at the position of the refocused echo. Experimental implementation of the ROOPh-DEER detection scheme requires at least three additional pulses as was demonstrated on an example of a 7-pulse sequence. The application of 7-pulse ROOPh-DEER sequence to a model biradical yielded the interspin distance of 1.94 ± 0.07 nm identical to the one obtained with the conventional 4-pulse DEER, however, without the unmodulated background present as a dominant fraction in the latter signal.
EPR 脉冲双共振（PDS）对于测量生物和其他系统中电子自旋之间的纳米级距离是不可或缺的。尽管近年来已经开发了几种有用的 PDS 改进和脉冲序列，但利用两个不同频率的泵浦和观测脉冲的 DEER 实验仍然是实际应用中最受欢迎的。所有可用的 DEER 方法的主要缺点之一是，由于泵浦脉冲不能完全反转耦合自旋，检测信号中存在显著的未调制部分。后一部分被认为是重构距离分布的主要误差源之一。我们描述了一种替代检测方案——重聚焦反相双共振（ROOPh-DEER），以仅获取双共振 DEER 信号的调制部分。当塞曼分裂与温度相比很小时，反相磁化分量相互抵消，在 4 脉冲 DEER 实验中观察不到。在 ROOPh-DEER 中，这些分量通过附加的泵浦脉冲重新聚焦，而包含未调制背景的同相分量则通过在观察频率下施加的脉冲过滤掉，该脉冲正好位于重新聚焦的回波位置。ROOPh-DEER 检测方案的实验实现需要至少三个附加脉冲，如 7 脉冲序列的示例所示。将 7 脉冲 ROOPh-DEER 序列应用于模型双自由基，得到的自旋间距离为 1.94±0.07nm，与传统的 4 脉冲 DEER 相同，但后者信号中不存在未调制背景。
