The diagram is divided into four panels, each representing a stage in the inhibition process. In the first panel, labeled OCS docking, DCPIB initially binds at the outer constriction site (OCS) near R103, with its hydrophobic moiety in orange and the carboxylate group in red. The second panel, labeled Fenestration partitioning, shows the hydrophobic portion of DCPIB partitioning into membrane-facing inter-subunit fenestrations, engaging the TM1-TM2 interface. The third panel, labeled Deep pore engagement, depicts DCPIB repositioning along the pore-lining interface toward deeper regions of the channel, influenced by TM2 residues such as L136 and M139. The final panel, labeled Inactivation, illustrates the stabilization of a nonconducting, inactivated channel conformation through coupling to pore geometry, lipid-associated gating mechanisms, TM2 packing, and N-terminal constriction elements.
Proposed multistep mechanism of DCPIB inhibition. DCPIB (orange, hydrophobic moiety; red, carboxylate group) inhibits LRRC8 channels through a dynamic, state-dependent process rather than by acting as a simple static pore plug. (OCS docking) DCPIB initially binds at the OCS, where the negatively charged carboxylate can interact with residues in EL1, including R103 in LRRC8A-containing channels. Fenestration partitioning: Following initial docking, the hydrophobic portion of DCPIB partitions into membrane-facing inter-subunit fenestrations, engaging the TM1–TM2 interface. Deep pore engagement: DCPIB subsequently repositions along the pore-lining interface toward deeper regions of the channel, where inhibition is influenced by TM2 residues such as L136 and M139 and by the local electrostatic environment. Inactivation: Pore engagement stabilizes a nonconducting, inactivated channel conformation through coupling to pore geometry, lipid-associated gating mechanisms, TM2 packing, and N-terminal constriction elements.