An allosteric agonist activates BK channels by perturbing coupling between Ca ²⁺ binding and pore opening

BK type Ca ²⁺ -activated K ⁺ channels activate in response to both the membrane voltage and intracellular Ca ²⁺ with distinct mechanisms. Ca ²⁺ binds to the cytosolic domain (CTD) to open the pore across the membrane, but the mechanism that couples Ca ²⁺ binding to pore opening is not clear. Here we show that a compound, BC5, identified using in silico screening, interacts with BK channels at the interface between the CTD and the transmembrane voltage sensing domain (VSD) and enhances channel activity by specifically affecting the Ca ²⁺ dependent mechanism. BC5 activates the channel in the absence of Ca ²⁺ binding but Ca ²⁺ binding inhibits BC5 effects. Thus, BC5 perturbs the pathway that couples Ca ²⁺ binding to pore opening to allosterically affect both, which is supported by atomistic simulations and mutagensis. The results suggest that the CTD- VSD interaction makes a major contribution to the mechanism of Ca ²⁺ dependent activation and is an important site for allosteric agonists to modulate BK channel activation.