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1.
Various time resolved EPR methods are applied to different test samples to demonstrate the abilities of pulsed high-field EPR spectroscopy. Two-pulse-echo field swept EPR spectroscopy on a nitroxide radical shows the increased spectral resolution by separating different spin systems by their relaxation properties. Additionally N14 electron-spin-echo-envelope-modulation (ESEEM) is observed for these systems at fields as high as 3.5 T. Thus, the N14 hyperfine interaction couplings can be probed by ESEEM and pulsed ENDOR (electron-nuclear-double-resonance) experiments. The sensitivity of pulsed ENDOR experiments is compared with cw-ENDOR. The different linewidths and amplitudes of the two methods are discussed. Transient nutation experiments on light induced triplet states demonstrate the high sensitivity and time resolution of high-field pulsed EPR. The sensitivity and time resolution of our 95 GHz spectrometer are determined and compared with pulsed X-band EPR spectrometer performances. 相似文献
2.
3.
Bode BE Dastvan R Prisner TF 《Journal of magnetic resonance (San Diego, Calif. : 1997)》2011,211(1):11-17
Pulsed electron-electron double resonance (PELDOR) spectroscopy is a powerful tool for measuring nanometer distances in spin-labeled systems. A common approach is doubly covalent spin-labeling of a macromolecule and measurement of the inter-spin distance, or to use singly-labeled components of a system that forms aggregates or oligomers. This situation has been described as a spin-cluster. The PELDOR signal, however, does not only contain the desired dipolar coupling between the spin-labels of the molecule or cluster under study. In samples of finite concentration the dipolar coupling between the spin-labels of the randomly distributed molecules or spin-clusters also contributes significantly. In homogeneous frozen solutions or lipid vesicle membranes this second contribution can be considered to be an exponential or stretched exponential decay, respectively. In this study, we show that this assumption is not valid in detergent micelles. Spin-labeled fatty acids that are randomly partitioned into different detergent micelles give rise to PELDOR time traces which clearly deviate from stretched exponential decays. The obtained signals can be modeled quantitatively based on the size of the micelles, their aggregation number, the spin-label concentration and the degree of spin-labeling. As a main conclusion a PELDOR signal deviating from a stretched exponential decay does not necessarily prove the observation of specific distance information on the molecule or cluster. These results are important for the interpretation of PELDOR experiments on membrane proteins or lipophilic peptides solubilized in detergent micelles or small vesicles, which often do not show pronounced dipolar oscillations in their time traces. 相似文献
4.
Erich Prisner 《Discrete Mathematics》2000,220(1-3):263-269
5.
A graphG is calledrepresentable in a tree T, ifG is isomorphic to the intersection graph of a family of subtrees ofT. In this paper those graphs are characterized which are representable in some subdivision of theK
1,n. In the finite case polynomial-time recognition algorithms of these graphs are given. But this concept can be generalized
to essentially infinite graphs by using no more trees but ‘tree-like’ posets and representability of graphs in these posets. 相似文献
6.
For aromatic organic radicals, pulsed electron-electron double resonance (PELDOR) experiments at high magnetic fields offer
the possibility to achieve orientation-selective pumping and detection that could allow one not only to determine the distance
between paramagnetic species but also their relative orientation with respect to the interconnecting dipolar axis. We present
a PELDOR two-frequency setup that was introduced into our homebuilt 180 GHz pulsed electron paramagnetic resonance (EPR) spectrometer
and we discuss its technical and experimental features. The capability of 180 GHz PELDOR has been tested using the three-pulse
ELDOR sequence on the protein RNR-R2 (ribonucleotide reductase) fromEscherichia coli, which contains two tyrosyl radicals at a distance of 3.3 nm. At 180 GHz, orientation selectivity is observed and the modulation
frequency was found in good agreement with theoretical predictions, which take into account the relative orientation of the
radicals from X-ray data. 相似文献
7.
Erich Prisner 《Results in Mathematics》1996,29(1-2):137-148
Given a graph-valued function ?, a graph G is called ? -periodic if there is some integer p ≥ 1 such that G is isomorphic to ?p(G). In this paper it is shown how to construct ?-periodic graphs, given essentially an embedding of a graph H as a subgraph of some iterated ?-graph ?p (H), provided ? obeys certain axioms. Then the results are applied to some graph-valued functions known in the literature. 相似文献
8.
Bode BE Plackmeyer J Prisner TF Schiemann O 《The journal of physical chemistry. A》2008,112(23):5064-5073
Metal ions are functionally or structurally important centers in metalloproteins or RNAs, which makes them interesting targets for spectroscopic investigations. In combination with site-directed spin labeling, pulsed electron-electron double resonance (PELDOR or DEER) could be a well-suited method to characterize and localize them. Here, we report on the synthesis, full characterization, and PELDOR study of a copper(II) porphyrin/nitroxide model system. The X-band PELDOR time traces contain besides the distance information a convolution of orientational selectivity, conformational flexibility, exchange coupling, and spin density distribution, which can be deconvoluted by experiments with different frequency offsets and simulations. The simulations are based on the known experimental and spin Hamiltonian parameters and make use of a geometric model as employed for structurally similar bis-nitroxides and spin density parameters as obtained from density functional theory calculations. It is found that orientation selection with respect to dipolar angles is only weakly resolvable at X-band frequencies due to the large nitrogen hyperfine coupling of the copper porphyrin. On the other hand, the PELDOR time traces reveal a much faster oscillation damping than observed for structurally similar bis-nitroxides, which is mainly assigned to a small distribution in exchange couplings J. Taking the effects of orientation selectivity, distribution in J, and spin density distribution into account leads finally to a narrow distance distribution caused solely by the flexibility of the structure, which is in agreement with distributions from known bis-nitroxides of similar structure. Thus, X-band PELDOR measurements at different frequency offsets in combination with explicit time trace simulations allow for distinguishing between structural models and quantitative interpretation of copper-nitroxide PELDOR data gives access to localization of copper(II) ions. 相似文献
9.
Lyubenova S Siddiqui MK Vries MJ Ludwig B Prisner TF 《The journal of physical chemistry. B》2007,111(14):3839-3846
The complex formed between cytochrome c oxidase from Paracoccus denitrificans and its electron-transfer partner cytochrome c has been studied by multi-frequency pulse electron paramagnetic resonance spectroscopy. The dipolar relaxation of a fast-relaxing paramagnetic center induced on a more slowly relaxing center can be used to measure their distance in the range of 1-4 nm. This method has been used here for the first time to study transient protein-protein complex formation, employing soluble fragments for both interacting species. We observed significantly enhanced transversal relaxation of the CuA center in cytochrome c oxidase due to the fast-relaxing iron of cytochrome c upon complex formation. The possibility to measure cytochrome c oxidase in the presence and absence of cytochrome c permitted us to separate the dipolar relaxation from other relaxation contributions. This allowed a quantitative simulation and interpretation of the relaxation data. The specific temperature dependence of the dipolar relaxation together with the high orientational selectivity achieved at high magnetic field values may provide detailed information on distance and relative orientation of the two proteins with respect to each other in the complex. Our experimental results cannot be explained by any single well-defined structure of the complex of cytochrome c oxidase with cytochrome c, but rather suggest that a broad distribution in distances and relative orientations between the two proteins exist within this complex. 相似文献
10.
Hellmich UA Lyubenova S Kaltenborn E Doshi R van Veen HW Prisner TF Glaubitz C 《Journal of the American Chemical Society》2012,134(13):5857-5862
Members of the ATP binding cassette (ABC) transporter superfamily translocate various types of molecules across the membrane at the expense of ATP. This requires cycling through a number of catalytic states. Here, we report conformational changes throughout the catalytic cycle of LmrA, a homodimeric multidrug ABC transporter from L. lactis. Using site-directed spin labeling and pulsed electron-electron double resonance (PELDOR/DEER) spectroscopy, we have probed the reorientation of the nucleotide binding domains and transmembrane helix 6 which is of particular relevance to drug binding and part of the dimerization interface. Our data show that LmrA samples a very large conformational space in its apo state, which is significantly reduced upon nucleotide binding. ATP binding but not hydrolysis is required to trigger this conformational change, which results in a relatively fixed orientation of both the nucleotide binding domains and transmembrane helices 6. This orientation is maintained throughout the ATP hydrolysis cycle until the protein cycles back to its apo state. Our data present strong evidence that switching between two dynamically and structurally distinct states is required for substrate translocation. 相似文献