Protein conformation by EPR spectroscopy using gadolinium tags clicked to genetically encoded p-azido-L-phenylalanine

Quantitative cysteine-independent ligation of a Gd3+ tag to genetically encoded p-azido-L-phenylalanine via Cu(I)-catalyzed click chemistry is shown to deliver an exceptionally powerful tool for Gd3+-Gd3+ distance measurements by double electron-electron resonance (DEER) experiments, as the position of the Gd3+ ion relative to the protein can be predicted with high accuracy.