title: Versatile Trityl Spin Labels for Nanometer Distance Measurements on Biomolecules In Vitro and within Cells.
abstract: Structure determination of biomacromolecules under in-cell conditions is a relevant yet challenging task. Electron paramagnetic resonance (EPR) distance measurements in combination with site-directed spin labeling (SDSL) are a valuable tool in this endeavor but the usually used nitroxide spin labels are not well-suited for in-cell measurements. In contrast, triarylmethyl (trityl) radicals are highly persistent, exhibit a long relaxation time and a narrow spectral width. Here, the synthesis of a versatile collection of trityl spin labels and their application in in vitro and in-cell trityl-iron distance measurements on a cytochrome P450 protein are described. The trityl labels show similar labeling efficiencies and better signal-to-noise ratios (SNR) as compared to the popular methanethiosulfonate spin label (MTSSL) and enabled a successful in-cell measurement.
authors.affiliation: Institute of Physical and Theoretical Chemistry, University of Bonn, Wegelerstr. 12, 53115, Bonn, Germany.
authors.forename: J Jacques
authors.initials: JJ
authors.lastname: Jassoy
authors.affiliation: Institute of Physical and Theoretical Chemistry, University of Bonn, Wegelerstr. 12, 53115, Bonn, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Berndhäuser
authors.affiliation: Institute of Physical and Theoretical Chemistry, University of Bonn, Wegelerstr. 12, 53115, Bonn, Germany.
authors.forename: Fraser
authors.initials: F
authors.lastname: Duthie
authors.affiliation: Institute of Physical and Theoretical Chemistry, University of Bonn, Wegelerstr. 12, 53115, Bonn, Germany.
authors.forename: Sebastian P
authors.initials: SP
authors.lastname: Kühn
authors.affiliation: Institute of Physical and Theoretical Chemistry, University of Bonn, Wegelerstr. 12, 53115, Bonn, Germany.
authors.forename: Gregor
authors.initials: G
authors.lastname: Hagelueken
authors.affiliation: Institute of Physical and Theoretical Chemistry, University of Bonn, Wegelerstr. 12, 53115, Bonn, Germany.
authors.forename: Olav
authors.identifier: 0000-0001-6346-9779
authors.initials: O
authors.lastname: Schiemann
mesh_terms: D000818:Animals; D003577:Cytochrome P-450 Enzyme System / Q000032:analysis*; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods*; D007501:Iron / Q000032:analysis; D009865:Oocytes / Q000201:enzymology; D016958:Pseudomonas putida / Q000201:enzymology*; D059629:Signal-To-Noise Ratio; D013113:Spin Labels / Q000138:chemical synthesis*; D014320:Triphenylmethyl Compounds / Q000138:chemical synthesis*; D029867:Xenopus Proteins / Q000032:analysis*; D014982:Xenopus laevis / Q000378:metabolism
chemical_list: D013113:Spin Labels; D014320:Triphenylmethyl Compounds; D029867:Xenopus Proteins; D003577:Cytochrome P-450 Enzyme System; D007501:Iron
keywords: EPR spectroscopy; distance measurements; in-cell spectroscopy; protein structures; spin labeling