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A Bioresistant Nitroxide Spin Label for In‐Cell EPR Spectroscopy: In Vitro and In Oocytes Protein Structural Dynamics Studies
Author(s) -
Karthikeyan Ganesan,
Bonucci Alessio,
Casano Gilles,
Gerbaud Guillaume,
Abel Sébastien,
Thomé Virginie,
Kodjabachian Laurent,
Magalon Axel,
Guigliarelli Bruno,
Belle Valérie,
Ouari Olivier,
Mileo Elisabetta
Publication year - 2018
Publication title -
angewandte chemie
Language(s) - English
Resource type - Journals
eISSN - 1521-3757
pISSN - 0044-8249
DOI - 10.1002/ange.201710184
Subject(s) - electron paramagnetic resonance , site directed spin labeling , nitroxide mediated radical polymerization , spin label , chemistry , pulsed epr , context (archaeology) , biophysics , chaperone (clinical) , protein dynamics , nuclear magnetic resonance , protein structure , chemical physics , biochemistry , physics , biology , spin echo , magnetic resonance imaging , radical polymerization , organic chemistry , copolymer , polymer , medicine , paleontology , radiology , pathology
Approaching protein structural dynamics and protein–protein interactions in the cellular environment is a fundamental challenge. Owing 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 nitroxide‐based spin labels for in‐cell studies has represented a major hurdle because of their short persistence in the cellular context. 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 is presented. 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.