Structural Mechanism Underlying TRPV1 Activation By Pungent Compounds In Gingers

Objective:TRPV1?transient receptor potential vanilloid 1?channel is a non-selective cation channel on the cell membrane,which can participate in a variety of physiological and pathological processes in the body,especially in the aspects of pain production and conduction as well as pain sensitivity enhancement.It has become a new target for analgesics.Capsaicin in peppers gives a pungent sensation.Capsaicin is a specifically targeting and actvating TRPV1,the molecular mechanism underlying capsaicin activation at this receptor has been validated through a series of experiments.Like chili peppers,gingers produce pungent stimuli by a group of vanilloid compounds that also activate the nociceptive TRPV1 ion channel.How these compounds interact with TRPV1 remains unclear.Methods:We used computational structural modeling,functional tests?electrophysiology and calcium imaging?and mutagenesis to investigate the structural mechanism for ligand-channel interactions.Results:1.The vanilloid compounds in gingers can directly activate wild-type TRPV1 channel(6-shogaol:EC₅₀=1.4±0.1?M;6-gingerol:EC₅₀=2.9±0.3?M;zingerone:EC₅₀=2.6±0.9 mM).Capsaicin analog capsazepine can competitively antagonize the activation of TRPV1 by the ginger compound.2.Results from calcium imaging experiments showed that when the wild type mTRPV1 was mutated to T551V or E571A,the EC₅₀0 value of the ginger compound activating TRPV1 channels was significantly increased,causing the activation curve to shift to the right.These observatious indicat that T551 and E571 are key residues for the interaction of ginger compounds with the TRPV1 channel.3.Electrophysiological experiments further confirmed that the two residues T551 and E571 are key sites for the interaction of ginger compounds with TRPV1 channels.4.Results from molecular docking indicate that T551 and E571 can interact with the three active components in gingers.It was found that there may be a new interaction between zingerone and T671 residues that was not observed from capsaicin-TRPV1interactions.Results from electrophysiology showed that when a point mutation T671S was introduced,the EC₅₀0 of zingerone activation was significantly increased.Conclusions:Our data revealed fundamental mechanistic features shared by three principle ginger pungent compounds,shogaol,gingerol,zingerone,and the chili pepper pungent compound,capsaicin,including a“head-down,tail-up”bound-ligand orientation,two specific hydrogen bonds as well as important contributions of van der Waals interactions by the aliphatic tail.We also identified how unique molecular structures in the ginger compounds affect channel activation potency,as well as a new interaction mode by zingerone.Structural insights on TRPV1-ginger compound complexes should help guide pharmaceutic efforts to regulate nociception.