HIGH FREQUENCY ELECTRON NUCLEAR DOUBLE RESONANCE

高频电子核双共振

• 批准号: 6279715
• 负责人: MARINA L BENNATI
• 金额: $ 1.18万
• 依托单位: MASSACHUSETTS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-05-01 至 1999-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6279715
• 关键词: biological products; biomedical equipment development; biomedical resource; enzymes; technology /technique

We have recently implemented a 140 GHz pulsed ENDOR spectrometer for detailed studies of the hyperfine structure and the electron spin density of different nuclei, specifically 13C, 2H and 1H. The spectra obtained are the highest frequency pulsed ENDOR spectra currently available. The high resolution of this technique allows for the complete separation of hyperfine spectra of different nuclei, in contrast to conventional ENDOR at low frequencies. For instance, the 2H ENDOR spectrum of deuterated BDPA (bisdiphenylene phenyl allyl) is almost not resolvable with 9 GHz ENDOR due to the low 2H-Larmor frequency (2.3 MHz) and overlap with 13C-resonances, whereas two distinct doublets are resolved at 140 GHz. The peaks arise from the hyperfine splittings of two inequivalent sets of deuterons. Different types of pulsed ENDOR excitation and detection schemes (Davies-, Mims-, TRIPLE-, Refocused ENDOR) have been used in order to demonstrate the sensitivity of the spectrometer. The performance of different pulse sequences is strongly influenced by the application of long soft pulses. The largest ENDOR effect was observed in Mims-ENDOR; however, the spectral line shape suffers from typical blindspots at A = n/t, where A is the hyperfine constant and t is the time between the two 900 excitation pulses. Blindspots can mostly be removed by use of the Refocused-Mims technique. With our current ENDOR spectrometer, we obtained the best sensitivity employing Davies-ENDOR with FID detection. A long, soft 1800 preparation pulse provides for high selectivity in the excitation of the EPR line and produces a narrow hole (< 1 MHz) in the ENDOR spectrum, leading to high sensitivity for small hyperfine couplings. The FID signal can be detected and integrated with minor error without phase cycling, due to the intrinsic short dead time of the 140 GHz spectrometer (< SO ns). The S/N of the FID detected spectra is a factor 10 higher than obtained with conventional spin-echo detection. The substantial g-anisotropy of most organic radicals at S T allows orientation selectivity in the ENDOR experiment. The greater resolution and sensitivity at high fields has been exploited to obtain Davies-ENDOR spectra of the tyrosyl radical in RNR-B2 from E. coli. ENDOR spectra were recorded at different field positions within the rhombic EPR powder pattern. In the small coupling region (A < 5 MHz) we observe resonances from the 2,6-ring- and the weakly coupled ~3-methylene protons. The structure of the spectra is complicated due to the overlapping of different lines, but a pair of doublets is resolved for excitation on the edges of the EPR spectrum (gz and gx), where only few orientations contribute to the ENDOR spectrum. A simulation program was developed which is capable of calculating the ENDOR spectrum for different preparation and detection schemes. The simulations reproduce the essential features of the experimental spectra. We obtained a set of hyperfine couplings in excellent agreement with the parameter set proposed by Hoganson et al.

我们最近实现了140 GHz脉冲ENDOR谱仪 详细研究超精细结构和电子自旋 不同核的密度，特别是13 C，2 H和1H。 光谱 获得了目前最高频率的脉冲ENDOR谱 available. 该技术的高分辨率允许 完全分离不同核的超精细光谱， 与传统的低频ENDOR形成对比。 你比如说 氘代BDPA（双二苯基苯基烯丙基）的2 H ENDOR光谱是 由于2 H-Larmor较低，使用9 GHz ENDOR几乎无法分辨 频率（2.3 MHz），并与13 C-共振重叠，而两个 在140 GHz分辨出不同的双峰。 这些峰来自于 两组不等价的氘核的超精细分裂。 不同类型的脉冲ENDOR激发和检测方案 （Davies-、Mims-、TRIPLE-、Refocused ENDOR）， 证明光谱仪的灵敏度。 的性能 不同的脉冲序列强烈地受到以下应用的影响： 长而柔和的脉搏 最大的ENDOR效应是在 然而，谱线形状遭受典型的 A = n/t处的盲点，其中A是超精细常数，t是 两个900激发脉冲之间的时间。 盲点大多可以是 通过使用重新聚焦-Mims技术去除。 以我们现在的 ENDOR光谱仪，我们获得了最佳的灵敏度， 带FID检测器的Davies-ENDOR。 一个长而柔和的1800准备脉冲 在EPR谱线的激发中提供高选择性， 在ENDOR频谱中产生窄孔（< 1 MHz），导致 高灵敏度，适用于小型超精细耦合。 FID信号可以是 检测并积分，误差较小，无相位循环，由于 140 GHz频谱仪的固有短死时间（<50 ns）。 FID检测光谱的S/N比 用常规的自旋回波检测获得。 大幅 大多数有机自由基在ST处的g-各向异性允许取向 ENDOR实验中的选择性。 更高的分辨率和 利用高场灵敏度获得Davies-ENDOR RNR-B2中酪氨酰自由基的光谱。杆菌 ENDOR谱 在菱形EPR中的不同场位置记录 粉末图案。 在小耦合区（A < 5 MHz），我们观察到 2，6-环-和弱耦合~3-亚甲基的共振 质子 光谱的结构是复杂的，因为 重叠的不同的线，但一对双峰被解决为 EPR谱边缘的激发（gz和gx），其中只有 少数取向对ENDOR谱有贡献。 仿真 开发了一个能够计算ENDOR的程序 不同制备和检测方案的光谱。 的 模拟再现了实验的基本特征， 谱 我们得到了一组极好的超精细耦合 与Hoganson等人提出的参数集的协议。