19F NMR chemical shift encoded peptide screening targeting the potassium channel Kv1.3†

¹⁹F nuclear magnetic resonance (¹⁹F-NMR) is a pivotal technique for protein dynamic studies and drug screening because of its high sensitivity. Herein, we report a ¹⁹F-NMR chemical shift-based ligand screening strategy targeting membrane proteins using multiple peptide ligands containing different ¹⁹F-incorporating unnatural amino acids. Five different ¹⁹F-labelled unnatural amino acids (^3FF, ^4FF, ^5FW, ^6FW and ^7FW) with distinctive ¹⁹F chemical shift values were applied to chemically synthesize multiple toxin peptides, which can potentially bind to and inhibit the conductance function of the autoimmune disease-related potassium channel Kv1.3. The ¹⁹F NMR relaxation-filtered one-dimensional spectra of competitively eluted peptides from the Kv1.3-peptide complex directly revealed the high-affinity binding of the peptides, which was verified using patch-clamp electrophysiological analysis. This ¹⁹F-NMR chemical shift-based encoded peptide screening method can be directly extended for large-scale peptide library screening targeting membrane proteins.