Glycine, a determinant of paxilline block in BK channels: A novel bend in the BK S6 helix

The tremorogenic fungal metabolite, paxilline, is widely used as a potent: and relatively specific blocker of Ca ²⁺- and voltage-activated Slo1 (or BK) K⁺ channels. The pH-regulated Slo3 K⁺ channel, a Slol homologue, is resistant to blockade by paxilline. Taking advantage of the marked differences in paxilline sensitivity and the homology between subunits, we have examined The paxilline sensitivity of a set of chimeric Slol/SIoS subunits. Paxilline sensitivity is associated with elements of the S5-P loop-S6 module of the Slo1 channel. Replacement of the Slo1 S5 segment or the second half of the P loop results in modest, changes in paxilline sensitivity, Replacing the Slol S6 segment with the Slo3 sequence abolishes paxilline sensitivity. An increase in paxilline affinity and changes in block kinetics also result, from replacing the first part of the Slo1 P loop, the so-called turret, with Slo3 sequence. The Slol and SloS S6 segments differ at 10 residues. Slo1-GS11S was found to markedly reduce paxilline block. In constructs with a SloS S6 segment, S300G restored paxilline block, but most effectively when paired with a Slo1 P loop. Other S6 residues differing between Slo1 and SloS had little influence on paxilline block. The involvement of Slo1 GS11 in paxilline sensitivity suggests that paxilline may occupy a position within the central cavity or access its blocking position through the central cavity. To explain the differences in paxilline sensitivity between Slol and SloS, we propose that the GS11/S300 position in Slo1 and SloS underlies a structural difference between subunits in the bend of S6, which influences the occupancy by paxilline,