Replication Data for: Sigmatropic rearrangement enables access to a highly stable spirocyclic nitroxide for protein spin labelling

Nitroxides are stable organic radicals with exceptionally long lifetimes, which render them uniquely suitable as observable probes or polarising agents for spectroscopic investigation of biomolecular structure and dynamics. Spin labelling enables the study of biomolecules using electron paramagnetic resonance (EPR) spectroscopy. Here, we describe the synthesis of a spin label based on a spirocyclic pyrrolidinyl nitroxide containing an iodoacetamide moiety. The spin label was successfully used for double labelling of a calmodulin mutant, and was shown to be highly persistent under reducing conditions while maintaining excellent relaxation parameters up to a temperature of 180 K. Interspin distances measured by double electron-electron resonance (DEER) were in good agreement with the protein tertiary structure.