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

Abstract

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.