Determination of End-to-End Distances in a Series of TEMPO Diradicals of up to 2.8 nm Length with a New Four-Pulse Double Electron Electron Resonance Experiment

Martin, Rainer E.; Pannier, M.; Diederich, François; Gramlich, Völker; Hubrich, Michael; Spieß, Hans Wolfgang

doi:10.1002/(sici)1521-3773(19981102)37:20<2833::aid-anie2833>3.3.co;2-z

Cited by 110 publications

(155 citation statements)

References 0 publications

Supporting

Mentioning

154

Contrasting

Unclassified

Order By: Relevance

“…All EPR work mentioned was performed with the direct detection of static dipolar interaction between two paramagnetic centers [27][28][29][30]. The corresponding pulse double electron-electron resonance (DEER or PELDOR) method has developed into a 'semi-routine' approach with well established experimental schemes [30], data processing algorithms [31,32] and open source data analysis software [33].…”

Section: Introductionmentioning

confidence: 99%

Distance determination from dysprosium induced relaxation enhancement: a case study on membrane-inserted WALP23 polypeptides

et al. 2013

View full text Add to dashboard Cite

Abstract. Membrane incorporated synthetic α-helical polypeptides labeled with Dy(III) chelate complexes and nitroxide radicals were studied by the inversion recovery (IR) technique and Dy(III)-nitroxide distances were obtained. A comparison of obtained distances with the previously reported Gd(III)-nitroxide pulse double electron-electron resonance (DEER or PELDOR) calibration data was performed and revealed reliability of the IR-based technique for the distance determination in membrane-incorporated biomacromolecules. The presented distance determination technique is 'spectroscopically orthogonal' to DEER-based distance measurements and can be potentially combined with DEER to study multiply spin-labeled biomacromolecules. The key steps of the data processing, the types of obtained distance information and the areas of possible application of the technique are discussed.

show abstract

Section: Introductionmentioning

confidence: 99%

Distance determination from dysprosium induced relaxation enhancement: a case study on membrane-inserted WALP23 polypeptides

et al. 2013

View full text Add to dashboard Cite

show abstract

Section: Introductionmentioning

confidence: 99%

“…There is an intrinsic cysteine at position 218, but it is buried, leading to <10% labeling even with a 100-fold molar excess of the spin label. The mutant proteins and their spin labeled derivatives catalyze DNA cleavage and have DNA binding affinities similar to that of wild type EcoRI, indicating that they are functionally active (Supporting Information).DEER experiments [11] were performed on spin labeled S180C specific and non-specific complexes, and on R131C and K249C-S180C specific, miscognate, and non-specific complexes. The four-pulse DEER data for each of the mutant complexes are shown in Figure 2a.…”

mentioning

confidence: 99%

“…DEER experiments [11] were performed on spin labeled S180C specific and non-specific complexes, and on R131C and K249C-S180C specific, miscognate, and non-specific complexes. The DEER experiment [11] is now well established for measuring distance constraints in membrane proteins, [12,13] soluble proteins, [14,15] peptides, [16] oligonucleotides, [17,18] and synthetic oligomers.…”

mentioning

confidence: 99%

“…The DEER experiment [11] is now well established for measuring distance constraints in membrane proteins, [12,13] soluble proteins, [14,15] peptides, [16] oligonucleotides, [17,18] and synthetic oligomers. [19][20][21] Recently, the DEER experiment has been used to probe structural rearrangements upon metal binding in the anthracis repressor, a DNA binding protein.…”

mentioning

confidence: 99%

See 2 more Smart Citations

Electron Spin Resonance Shows Common Structural Features for Different Classes of EcoRI–DNA Complexes

et al. 2008

View full text Add to dashboard Cite

KeywordsEPR spectroscopy; DEER; DNA binding protein; structure elucidation; protein flexibility; noncognate complexIn this communication, we show that the EcoRI restriction endonuclease binds different classes of DNA sites in the same binding cleft. EcoRI generates widespread interest because it exhibits an extraordinary sequence selectivity to carry out its function of cleaving incoming foreign DNA without causing potentially lethal cleavage of cellular DNA. For example, EcoRI binds to its correct recognition site GAATTC up to 90,000-fold better than to miscognate sites that have one incorrect base pair. [1,2] The ∼650 specific sites in the E. coli genome are protected from cleavage by double-strand methylation. The ∼21,000 miscognate sites are not methylated, but are still cleaved by EcoRI with a second-order rate constant that is ∼10 9 -fold lower.[1,2] EcoRI forms only non-specific complexes with no cleavage at sites that differ from GAATTC by two or more base pairs. [1,2] In order to understand the source of such high specificity, it is necessary to determine how the structures of EcoRI complexes differ at specific, miscognate (5/6 bp match), and nonspecific (≤4/6 bp match) DNA sites. This effort is timely given the extensive genetic, biochemical and biophysical data on EcoRI.[1-9] Footprinting results [1] suggest that the three classes of complexes are "structurally" distinct, and thermodynamic profiles (ΔG°, ΔH°, ΔS°, ΔC°P) [3,4] suggest that the specific complex has more restricted conformationalvibrational mobility of the protein and DNA. There are crystal structures of the free protein, Figure 1 shows the structure of the EcoRI specific complex. [6,7] The protein contains a large, relatively rigid and structured globular "main" domain and a smaller "arm" region. The protein arms are invisible in the free protein[6] but become ordered and enfold the DNA in the specific complex, where they play a role in modulating specificity.[2,4] Mutations R131C, S180C, and K249C-S180C were chosen based on the crystal structure. [6,7] These sites are solvent accessible and therefore likely to spin label with minimal perturbation to protein structure. Residues R131 and S180 lie in the inner and outer arms, respectively. Residue K249 is in the main domain, which has very restricted movement [6] and acts as a reference point. Since EcoRI is a 62 kDa homodimer, single cysteine mutations provide two sites for spin labeling, and double mutations provide four sites.The proteins were spin labeled at the cysteines with the methanethiosulfonate spin label (MTSSL). There is an intrinsic cysteine at position 218, but it is buried, leading to <10% labeling even with a 100-fold molar excess of the spin label. The mutant proteins and their spin labeled derivatives catalyze DNA cleavage and have DNA binding affinities similar to that of wild type EcoRI, indicating that they are functionally active (Supporting Information).DEER experiments [11] were performed on spin labeled S180C specific and non-specific complexes, and on R131C and...

show abstract

Determination of the Nanostructure of Polymer Materials by Electron Paramagnetic Resonance Spectroscopy

Jeschke

2002

Macromol. Rapid Commun.

174

164

View full text Add to dashboard Cite

Electron paramagnetic resonance (EPR) spectroscopy is one the few methods that can characterize structural features in the range between 0.5 and 5 nm in systems that lack long‐range order. Approaches based on EPR spectroscopy provide good structural contrast even in complex materials, as the sites of interest can be selectively labeled or addressed by suitably functionalized spin probes using well established techniques. This article assesses the EPR experiments available for distance measurements on nanoscales in terms of the accessible distance range, precision, and sensitivity. Recommendations are derived for the proper choice of experiment for a given problem. Both simple and sophisticated methods for data analysis are described and their limitations are evaluated. It is discussed which assumptions must be made to extract a pair correlation function from EPR data. Finally, applications to the study of polymer chain conformation and the structure of ionically functionalized diblock copolymers are highlighted.

show abstract

Determination of End-to-End Distances in a Series of TEMPO Diradicals of up to 2.8 nm Length with a New Four-Pulse Double Electron Electron Resonance Experiment

Cited by 110 publications

References 0 publications

Distance determination from dysprosium induced relaxation enhancement: a case study on membrane-inserted WALP23 polypeptides

Distance determination from dysprosium induced relaxation enhancement: a case study on membrane-inserted WALP23 polypeptides

Electron Spin Resonance Shows Common Structural Features for Different Classes of EcoRI–DNA Complexes

Determination of the Nanostructure of Polymer Materials by Electron Paramagnetic Resonance Spectroscopy

Contact Info

Product

Resources

About