ADDING KU-BAND DEER CAPABILITY TO THE DQC SPECTROMETER

为 DQC 光谱仪添加 KU 波段鹿功能

• 批准号: 8172090
• 负责人: PETER P BORBAT
• 金额: $ 0.29万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-01 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8172090
• 关键词: Back; Computer Retrieval of Information on Scientific Projects Database; Deer; Diffusion; Electrons; Frequencies; Funding; Grant; Heat shock proteins; Institution; Lipids; Liposomes; Membrane Proteins; Phase; Physiologic pulse; Positioning Attribute; Potassium Channel; Pump; Research; Research Personnel; Resources; Sampling; Signal Transduction; Solid; Source; Spin Labels; System; Techniques; United States National Institutes of Health; Water; Work; base; cold temperature; cytochrome c; microwave electromagnetic radiation; protein oligomer; reconstitution; voltage

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Pulsed double electron-electron resonance (PELDOR or DEER) is an established technique for the determination of distances and their distributions in low-temperature solids. The technique is based on recording the amplitude of primary or refocused echo as a function of the position of an additional microwave pulse applied at a different frequency. The technique is useful for sufficiently broad ESR spectra as the pulses at the two frequencies should have small or no overlap. Typically, it is realized with two independent pulse sources and bimodal resonators or else with a single source and low-Q dielectric or loop-gap resonator. The technique is useful when spin concentrations are not small. In some cases of spin-labeled membrane proteins reconstituted in liposomes with large excess water there are increased local spin concentrations in the lipid phase leading to a loss of the signal by the mechanism of instantaneous diffusion. Also, there are cases such as spin clusters, e.g. protein oligomers, when the use of strong pulses leads to a loss of the signal. DEER based on weak pulses is less sensitive to instantaneous diffusion and can compete favorably with DQC in such cases. In order to accommodate a wider range of systems, we set up DEER at Ku-band (17.4 GHz), by adding a second pulse channel to the existing 9/17GHz FT-ESR spectrometer currently used for DQC work. The use of a higher frequency than standard 9 GHz leads to a higher sensitivity for typical sample volumes. The pumping pulse at the second frequency is injected via a directional coupler into the path to the dielectric resonator, which is tuned for DQC and therefore has the sufficiently low Q needed to accommodate pulses with a frequency difference of 65-70 MHz. Switching back and forth between DEER and DQC can be accomplished in a minute. We have successfully applied Ku-band DEER to the study of the small heat-shock protein Hsp16.5, voltage-dependent K+ channel KvAP, and unfolded Cytochrome-c.

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 脉冲双电子-电子共振（PELDOR或DEER）是一种用于测定低温固体中距离及其分布的成熟技术。该技术是基于记录的幅度的主要或重新聚焦的回波作为一个功能的位置的附加微波脉冲施加在一个不同的频率。该技术对于足够宽的ESR谱是有用的，因为在两个频率处的脉冲应该具有小的重叠或没有重叠。通常，它是用两个独立的脉冲源和双模谐振器实现的，或者用单个源和低Q电介质或环隙谐振器实现的。当自旋浓度不小时，该技术是有用的。在某些情况下，自旋标记的膜蛋白与大量过量的水在脂质体中重建，在脂质相中有增加的局部自旋浓度，导致瞬时扩散机制的信号损失。此外，当使用强脉冲导致信号丢失时，存在诸如自旋簇（例如蛋白质寡聚体）的情况。基于弱脉冲的DEER对瞬时扩散不太敏感，在这种情况下可以与DQC竞争。为了适应更广泛的系统，我们在Ku波段（17.4 GHz）设置了DEER，通过在现有的9/17 GHz FT-ESR谱仪上增加第二个脉冲通道，目前用于DQC工作。使用比标准9 GHz更高的频率可提高典型样品体积的灵敏度。第二频率的泵浦脉冲通过定向耦合器注入到介质谐振器的路径中，该介质谐振器被调谐用于DQC，因此具有容纳具有65-70 MHz频率差的脉冲所需的足够低的Q。在DEER和DQC之间来回切换可以在一分钟内完成。 我们已经成功地将Ku波段DEER应用于小分子热休克蛋白Hsp16.5、电压依赖性K+通道KvAP和未折叠细胞色素c的研究。