On the Use of Q-Band Double Electron-Electron Resonance To Resolve the Relative Orientations of Two Double Histidine-Bound Cu2+ Ions in a Protein
On the Use of Q-Band Double Electron-Electron Resonance To Resolve the Relative Orientations of Two Double Histidine-Bound Cu2+ Ions in a Protein
复制标题
DOI：
10.1021/acs.jpcb.8b07727
复制
发表时间：
2018-11-29
影响因子：
3.3
通讯作者：
Saxena, Sunil
中科院分区：
文献类型：
--
作者：
Jarvi, Austin Gamble;Ranguelova, Kalina;Saxena, Sunil
在这项工作中，我们探索了刚性Cu2+自旋标记技术的潜力，双组氨酸（dHis）基序，以及q波段电子顺磁共振来报告自旋标记的相对方向。我们发现dHis基序的精度，加上q波段频率的灵敏度和分辨率，可以使用双电子电子共振（DEER）直接确定dHis- cu2 +标签的相对取向。我们在不同磁场下对含有两个dHis Cu2+位点的蛋白质进行了q波段DEER测量。这些测量显示出定向选择性，使得每个离散磁场产生一个独特的DEER信号。我们通过模拟定向选择性DEER数据确定了两个金属中心的相对取向。这些相对取向通过dHis位点修饰的蛋白质晶体结构的视觉分析得到了验证。简单的视觉分析结果与实验数据的模拟结果吻合较好。dHis-Cu2+基模结合Q-band的优势可以帮助精确测量蛋白质的结构和构象动力学。
In this work, we explore the potential of a rigid Cu2+ spin-labeling technique, the double histidine (dHis) motif, along with Q-band electron paramagnetic resonance to report on the relative orientations of the spin labels. We show that the precision of the dHis motif, coupled with the sensitivity and resolution of Q-band frequencies, may allow for the straightforward determination of the relative orientation of the dHis-Cu2+ labels using double electron electron resonance (DEER). We performed Q-band DEER measurements at different magnetic fields on a protein containing two dHis Cu2+ sites. These measurements exhibited orientational selectivity such that each discrete magnetic field yielded a unique DEER signal. We determined the relative orientation of the two metal centers by simulating the orientationally selective DEER data. These relative orientations were validated by visual analysis of the protein crystal structure modified with dHis sites. The simple visual analysis was shown to agree well with the angular values determined via simulation of the experimental data. The combination of the dHis-Cu2+ motif along with the advantages of the Q-band can aid in the accurate measurement of protein structural and conformational dynamics.