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Long-range EPR distance measurements with semi-rigid spin labels at Q-band frequencies

dc.contributor.advisorBennati, Marina Prof. Dr.
dc.contributor.authorHalbmair, Karin
dc.publisherNiedersächsische Staats- und Universitätsbibliothek Göttingende
dc.titleLong-range EPR distance measurements with semi-rigid spin labels at Q-band frequenciesde
dc.contributor.refereeBennati, Marina Prof. Dr.
dc.description.abstractengElucidating the structure of biomolecules is essential to understand their function. Pulsed electron double resonance (PELDOR) in conjunction with site-directed spin labelling has become a well-established experiment for measuring long-range distances between paramagnetic centres. In this thesis the capacity of PELDOR distance measurements performed Q-band frequency (34 GHz/1.2 T) employing a high power microwave amplifier in conjunction with spin labels of reduced mobility is demonstrated. It is shown that the high microwave power does not only enable high sensitivity in PELDOR experiments but also allows to supress artefacts arising from orientation selection. The power of this method is documented for two representative biological model system spin labelled with distinct semi-rigid nitroxides. Distances up to 8 nm could be recorded with high accuracy (error in the main distance ≤ 0.1 nm) on a ribonucleic acid (RNA) duplex labelled with a TEMPO-based nitroxide called CT that is attached to a cytosine. CT is preserving Watson-Crick base-pairing capability, which restricts it to a small conformational space. Therefore subtle dynamics of the RNA itself could be observed. The narrow width of the distance distributions obtained in the experiment (Δr ≤ 0.5 nm) suggest overall small deviations of the duplex from an ideal A-form RNA. A transmembrane model peptide (WALP24) labelled with the standard MTSSL as well as the unnatural amino acid TOPP was investigated by EPR distance measurements in solution and two different lipid environments. A comparison between WALP24-MTSSL and WALP24-TOPP, clearly reveals the superiority of the rigid TOPP label, as it delivers sharp one-peak inter-spin distances (Δr ≤ 0.4 nm) even in the lipid environment. In contrast MTSSL reported broad distances that do show a strong dependence on the hydrophobic thickness of the bilayer, suggesting an adaption of the flexible label to the lipid environment, which results in a loss of information on peptide
dc.contributor.coRefereeDiederichsen, Ulf Prof. Dr.
dc.contributor.thirdRefereeZweckstetter, Markus Prof. Dr.
dc.contributor.thirdRefereeHöbartner, Claudia Prof. Dr.
dc.contributor.thirdRefereeSteinem, Claudia Prof. Dr.
dc.contributor.thirdRefereeMeinecke, Michael Prof. Dr.
dc.subject.engDistance Measurementde
dc.subject.engSpin labelde
dc.subject.engtransmembrane peptidede
dc.subject.engq bandde
dc.affiliation.instituteFakultät für Chemiede
dc.subject.gokfullChemie  (PPN62138352X)de

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