Approaching proteins structural dynamics and protein-protein interactions in the cellular environment is a fundamental challenge. Due to its absolute sensitivity and to its selectivity to paramagnetic species, Site-Directed Spin Labeling (SDSL) combined with Electron Paramagnetic Resonance (EPR) has the potential to evolve into an efficient method to follow conformational changes in proteins directly inside cells. Until now, the use of nitroxyde-based spin labels for in-cell studies has represented a major hurdle because of their short persistence in the cellular context. In this work we present the design and synthesis of the first maleimido-proxyl-based spin label (M-TETPO) resistant towards reduction and being efficient to probe protein dynamics by continuous wave and pulsed EPR. In particular, the extended lifetime of M-TETPO enabled the study of structural features of a chaperone in the absence and presence of its binding partner at endogenous concentration directly inside cells.
A combined EPR and MD simulation study of a nitroxyl spin label with restricted internal mobility sensitive to protein dynamics
From The DNP-NMR Blog:
A combined EPR and MD simulation study of a nitroxyl spin label with restricted internal mobility sensitive to protein dynamics
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Oganesyan, V.S., et al., A combined EPR and MD simulation study of a nitroxyl spin label with restricted internal mobility sensitive to protein dynamics. J. Magn. Reson., 2017. 274: p. 24-35.
www.sciencedirect.com/science/article/pii/S1090780716302270
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02-20-2017 03:39 PM
Nuclear spin-lattice relaxation in nitroxide spin-label EPR
From The DNP-NMR Blog:
Nuclear spin-lattice relaxation in nitroxide spin-label EPR
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Marsh, D., Nuclear spin-lattice relaxation in nitroxide spin-label EPR. J Magn Reson, 2016. 272: p. 166-171.
https://www.ncbi.nlm.nih.gov/pubmed/27712989
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12-02-2016 07:56 PM
[NMR paper] A combined EPR and MD simulation study of a nitroxyl spin label with restricted internal mobility sensitive to protein dynamics
A combined EPR and MD simulation study of a nitroxyl spin label with restricted internal mobility sensitive to protein dynamics
Publication date: January 2017
Source:Journal of Magnetic Resonance, Volume 274</br>
Author(s): Vasily S. Oganesyan, Fatima Chami, Gaye F. White, Andrew J. Thomson</br>
EPR studies combined with fully atomistic Molecular Dynamics (MD) simulations and an MD-EPR simulation method provide evidence for intrinsic low rotameric mobility of a nitroxyl spin label, Rn, compared to the more widely employed label MTSL (R1). Both experimental...
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11-19-2016 08:35 PM
The Spatial Effect of Protein Deuteration on Nitroxide Spin-label Relaxation: Implications for EPR Distance Measurement
The Spatial Effect of Protein Deuteration on Nitroxide Spin-label Relaxation: Implications for EPR Distance Measurement
Publication date: Available online 28 September 2014
Source:Journal of Magnetic Resonance</br>
Author(s): Hassane. El Mkami , Richard Ward , Andrew Bowman , Tom Owen-Hughes , David G. Norman</br>
Pulsed electron-electron double resonance (PELDOR) coupled with site-directed spin labeling is a powerful technique for the elucidation of protein or nucleic acid, macromolecular structure and interactions. The intrinsic high sensitivity of electron...
[NMR paper] Quantitative comparison of protein dynamics in live cells and in vitro by in-cell (19)F-NMR.
Quantitative comparison of protein dynamics in live cells and in vitro by in-cell (19)F-NMR.
Quantitative comparison of protein dynamics in live cells and in vitro by in-cell (19)F-NMR.
Chem Commun (Camb). 2013 Feb 26;
Authors: Takaoka Y, Kioi Y, Morito A, Otani J, Arita K, Ashihara E, Ariyoshi M, Tochio H, Shirakawa M, Hamachi I
Abstract
Here we describe how a (19)F-probe incorporated into an endogenous protein by a chemical biology method revealed protein dynamics. By explicit determination of ligand-bound and unbound structures with...
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02-27-2013 06:47 PM
Integrated analysis of the conformation of a protein-linked spin label by crystallography, EPR and NMR spectroscopy
Integrated analysis of the conformation of a protein-linked spin label by crystallography, EPR and NMR spectroscopy
Abstract Long-range structural information derived from paramagnetic relaxation enhancement observed in the presence of a paramagnetic nitroxide radical is highly useful for structural characterization of globular, modular and intrinsically disordered proteins, as well as proteinā??protein and protein-DNA complexes. Here we characterized the conformation of a spin-label attached to the homodimeric protein CylR2 using a combination of X-ray crystallography, electron...
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01-31-2011 06:03 AM
Integrated analysis of the conformation of a protein-linked spin label by crystallography, EPR and NMR spectroscopy.
Integrated analysis of the conformation of a protein-linked spin label by crystallography, EPR and NMR spectroscopy.
Integrated analysis of the conformation of a protein-linked spin label by crystallography, EPR and NMR spectroscopy.
J Biomol NMR. 2011 Jan 28;
Authors: Gruene T, Cho MK, Karyagina I, Kim HY, Grosse C, Giller K, Zweckstetter M, Becker S
Long-range structural information derived from paramagnetic relaxation enhancement observed in the presence of a paramagnetic nitroxide radical is highly useful for structural characterization of...
A bioresistant nitroxide spin label for in-cell EPR spectroscopy: in vitro and in oocytes protein structural dynamics studies.
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