LAUSR

Vanilloids activation of TRPV1 represents an excellent model system for ligand gating of ion channels. Recent studies using cryo electron microcopy (cryo-EM), computational analysis and functional quantification revealed the location of capsaicin-binding site and critical residues mediating ligand-binding and channel activation. To validate our current working model for capsaicin activation, here we have successfully converted the vanilloid-insensitive TRPV2 channel to be resiniferatoxin-activated, using a rationally designed minimal set of four point mutations (F467S-S498F-L505T-Q525E, termed TRPV2_Quad). We found that resiniferatoxin binds to TRPV2_Quad with sub-micromolar affinity even though the ligand-induced open state is relatively unstable. Using Rosetta-based molecular docking, we observed a common structural mechanism underlying vanilloids activation of TRPV1 and TRPV2_Quad, where the ligand serves as a molecular “glue” that bridges the S4-S5 linker to the S1-S4 domain to open these channels. Furthermore, our analysis revealed previously unrecognized structural requirements needed for capsaicin binding to TRPV1. These results should help guide the design of drug candidate compounds for this important pain sensor.