Multi-Quantum Spin Counting and Long Range Distance Measurements by Pulsed EPR

脉冲EPR方法允许在1-10 nm范围内确定大分子的距离。这是通过测量两个共价连接的顺磁自旋标记之间的磁偶极-偶极相互作用来完成的。在这个项目中，我们想从两个方面将这些方法扩展到最常用的氮氧化物自旋标记：1)通过开发具有宽带幅度和频率调制的微波脉冲的Carr-Purcell-Siefter脉冲序列来扩展可测量的距离范围。这样的重新聚焦序列使电子自旋与邻近的核自旋解耦，从而延长了观测时间窗。这对于具有高内部柔性的大分子系统(例如，本质上未折叠的蛋白质、膜蛋白质复合体或蛋白质-核酸复合体)的应用是重要的。2)利用宽带调幅和调频微波脉冲的多量子EPR实验计算络合物中的偶极耦合电子自旋。这将允许例如确定膜蛋白络合物在其天然脂环境中的低聚状态。新型实验将在模型化合物上进行测试和优化，然后应用于生物系统。

Pulsed EPR methods allow to determine distances in the 1-10 nm range on macromolecules. This is done by measuring the magnetic dipole-dipole interactions between two covalently attached paramagnetic spin labels. In this project we want to extend these methods for the most common used nitroxide spin labels in two respects:1) Extending the measurable distance range by developing Carr-Purcell-SIFTER pulse sequences with broadband amplitude and frequency modulated microwave pulses. Such refocusing sequences decouple the electron spin from closeby nuclear spins and allow to prolong the observation time window. This is important for applications to large macromolecular systems with high internal flexibility (as for example intrinsically unfolded proteins, membrane protein complexes or protein-nucleic acid complexes). 2) Counting of the dipolar coupled electron spins in a complex by Multi-Quantum EPR experiments with broadband amplitude and frequency modulated microwave pulses.This would allow to determine for example oligomeric states of membrane protein complexes in their native lipid environment. The new type of experiments will be tested and optimized on model compounds and then applied to biological systems.