Coupling of Ca²⁺and voltage activation in BK channels through the αB helix/voltage sensor interface

Abstract

Office phone: 305 243-6236. AbstractLarge conductance Ca 2+ and voltage activated K + (BK) channels control membrane excitability in many cell types. BK channels are tetrameric. Each subunit is comprised of a voltage sensor domain (VSD), a central pore gate domain, and a large cytoplasmic domain (CTD) that contains the Ca 2+ sensors. While it is known that BK channels are activated by voltage and Ca 2+ , and that voltage and Ca 2+ activations interact, less is known about the mechanisms involved. We now explore mechanism by examining the gating contribution of an interface formed between the VSDs and the αB helices located at the top of the CTDs. Proline mutations in the αB helix greatly decreased voltage activation while having negligible effects on gating currents. Analysis with the HCA model indicated a decreased coupling between voltage sensors and pore gate. Proline mutations decreased Ca 2+ activation for both Ca 2+ bowl and RCK1 Ca 2+ sites, suggesting that both high affinity Ca 2+ sites transduce their effect, at least in part, through the αB helix. Mg 2+ activation was also decreased. The crystal structure of the CTD with proline mutation L390P showed a flattening of the first helical turn in the αB helix compared to WT, without other notable differences in the CTD, indicating structural change from the mutation was confined to the αB helix. These findings indicate that an intact αB helix/VSD interface is required for effective coupling of Ca 2+ binding and voltage depolarization to pore opening, and that shared Ca 2+ and voltage transduction pathways involving the αB helix may be involved. Slo1 channel | BK channel | Ca 2+ -activated K + channel | allosteric coupling | patch clamp SignificanceLarge conductance BK (Slo1) K + channels are activated by voltage, Ca 2+ , and Mg 2+ to modulate membrane excitability in neurons, muscle, and other cells. BK channels are of modular design, with pore-gate and voltage sensors as transmembrane domains and a large cytoplasmic domain CTD containing the Ca 2+ sensors. Previous observations suggest that voltage and Ca 2+ sensors interact, but less is known about this interaction and its involvement in the gating process. We show that a previously identified structural interface between the CTD and voltage sensors is required for effective activation by both voltage and Ca 2+ , suggesting that these processes may share common allosteric activation pathways. Such knowledge should help explain disease processes associated with BK channel dysfunction.

BK channels of five different subunit combinations underlie the de novo KCNMA1 G375R channelopathy

“…4A and C). The differences in observations with those of Liang et al (24) are unlikely to arise from the different expression systems, as we have found that both oocytes and HEK293 cells effectively express WT BK channels and also BK channels with various types of mutations (16,38,40,41).…”

A gating lever and molecular logic gate that couple voltage and calcium sensor activation to opening in BK potassium channels

“…First, it is conceivable that the linker interacts with S6 in a different conformational state, as only two states are represented by existing structures and questions remain whether the PGD in the “Ca 2+ -free closed” structure is fully closed and whether the VSD in the “Ca 2+ -bound open” structure (which also has Mg 2+ bound) is fully activated ( 16 ). Second, the S4-S5 linker region contacts the CTD ( 7 ) and represents the only resolved noncovalent interaction between VSD and CTD in the Ca 2+ -free hSlo1 structure, suggesting a role in indirect coupling ( 17 ). Last, owing to the nonswapped architecture of Slo1, contacts exist between the bottoms of S4, S5, and S6 within each subunit (Fig.…”

Regulatory mechanisms of mitochondrial BKCa channels

“…Simultaneously, the lateral tilting of the N-lobe moves the αB helix located at the top of each N-lobe (Figure 1) both upward and laterally to push on the bottom of the S4-S5 linker/VSD to potentially open the channel. In addition, it has recently been demonstrated that in human BK Ca , the αB helix links the binding of Ca 2+ at the RCK domains to the VGD of the channel, confirming that interaction between the αB helices at the top of the N-lobes of the CTD and the cytoplasmic surfaces of the S4-S5 linkers/VSD is required to open the channel [42]. In agreement with this new model for channel opening, early observations have shown that both RCK1 and 2 domains can move independently from each other upon binding of Ca 2+ , indicating a high degree of flexibility for this domain [43].…”