Relationship between auxiliary gamma subunits and mallotoxin on BK channel modulation

Abstract

The large-conductance, calcium-and voltage-Activated K + (BK) channel consists of the pore-forming α subunits (BKα) and auxiliary subunits. The auxiliary I 31-3 subunits potently modulate the BK channel by shifting its voltage-dependence of channel activation toward the hyperpolarizing direction by approximately 145 mV (I 31), 100 mV (I 32), and 50 mV (I 33). Mallotoxin is a potent small-molecule BK channel activator. We analyzed the relationship between mallotoxin and the I 3 subunits in their BK channel-Activating effects in membrane patches excised from HEK-293 cells. We found that mallotoxin, when applied extracellularly, shifted the half-Activation voltage (V 1/2) of BKα channels by â '72 mV. The channel-Activating effect of mallotoxin was greatly attenuated in the presence of the I 31, I 32, or I 33 subunit, with resultant I "V 1/2 (+/â ' mallotoxin) values of â '9, â '28, or â '15 mV, respectively. Most examined I 31 mutant subunits antagonized mallotoxin's channel-Activating effect in a manner that was largely dependent on its own modulatory function. However, mallotoxin caused an irreversible functional and structural disengagement of the I 31-F273S mutant from BK channels. We infer that the auxiliary I 3 subunit effectively interferes with mallotoxin on BK channel modulation via either a direct steric competition or an indirect allosteric influence on mallotoxin's binding and action on BKα.