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Protein Spin Labeling with a Photocaged Nitroxide Using Diels-Alder Chemistry

submitted on 01.04.2019, 09:02 and posted on 02.04.2019, 16:30 by Anandi Kugele, Bjarne Silkenath, Jakob Langer, Valentin Wittmann, Malte Drescher
EPR spectroscopy of diamagnetic bio-macromolecules is based on site-directed spin labeling (SDSL). Here, we present a novel labeling strategy for proteins. We developed and synthesized a nitroxide-based spin label that can be ligated to proteins by an inverse-electron-demand Diels-Alder (DAinv) cycloaddition to genetically encoded non-canonical amino acids (ncAA). The nitroxide moiety is shielded by a photoremovable protecting group (PPG) with an attached tetraethylene glycol unit to achieve water solubility. We demonstrate SDSL of two model proteins with the PaNDA (Photoactivatable Nitroxide for DAinv reaction) label. Our strategy features high reaction rates combined with high selectivity, as well as high stability of the nitroxide in E. coli lysate.


European Research Council under the European Union's Horizon 2020 research and innovation programme (Grant Agreement Number: 772027 - SPICE - ERC-2017-COG)

Deutsche Forschungsgemeinschaft (SFB 969, Projects B5 and C3)


Email Address of Submitting Author


University of Konstanz



ORCID For Submitting Author


Declaration of Conflict of Interest

There are no conflicts of interest to declare.