Migration of PIP2 lipids on voltage-gated potassium channel surface influences channel deactivation

Abstract

Published studies of lipid-protein interactions have mainly focused on lipid binding to an individual site of the protein. Here, we show that a lipid can migrate between different binding sites in a protein and this migration modulates protein function. Voltage-gated potassium (Kv) channels have several potential binding sites for phosphatidylinositol-4,5-bisphosphate (PIP2). Our molecular dynamics (MD) simulations on the KCNQ2 channel reveal that PIP2 preferentially binds to the S4-S5 linker when the channel is in the open state while maintains a certain probability of migrating to the S2-S3 linker. Guided by the MD results, electrophysiological experiments using KCNQ2, KCNQ1, and hERG channels show that the migration of PIP2 toward the S2-S3 linker controls the deactivation rate of the channel. The data suggest that PIP2 can migrate between different binding sites in Kv channels with significant impacts on channel deactivation, casting new insights into the dynamics and physiological functions of lipid-protein interactions.

Early-onset epileptic encephalopathy caused by a reduced sensitivity of Kv7.2 potassium channels to phosphatidylinositol 4,5-bisphosphate

“…In Kv7.2 channels, as herein confirmed, PIP 2 up-regulates current density and facilitates voltage-dependent opening649; dynamic repositioning of PIP 2 from the VSD to the open pore gate occurs during activation49, whereas the reverse movement correlates with channel deactivation46. Thus, a critical PIP 2 binding site in Kv7.1 and Kv7.2 channels is located at the VSD-pore domain interface, where PIP 2 headgroups are engaged in electrostatic interactions with basic residues in the VSD and the proximal C-terminus (including the S 6 gate)341, thereby bridging these two domains and providing structural stabilization1.…”

“…Consistent with this view is also the decreased potency shown by the water-soluble PIP 2 analogue diC8-PIP 2 in activating Kv7.2 channel carrying the R325A mutation43. Noteworthy, at variance with Kv7.2 R325G channels, homomeric Kv7.2 R325A channels, similarly to Kv7.1 channels carrying the equivalent R360A mutation both in the absence44 or in the presence45 of KCNE1 subunits, were functional, although they displayed a reduced current amplitude4346; the α-helix-stabilizing propensity of alanine relative to glycine47 provides a plausible explanation for such relevant functional difference. Noteworthy, all Kv7.2 missense variants causing EE (including the c.973A > G leading to the R325G mutation herein investigated) are de novo substitutions of a single nucleotide748.…”

Predicting the Functional Impact of KCNQ1 Variants of Unknown Significance

“…The S4–S5 linker, which is believed to be responsible for transferring the conformational changes in the voltage sensor domain to the pore 29 and serve as binding sites for phosphatidylinositol-4,5-bisphosphate (PIP2) to modulate the deactivation rate of the channel, 30 has a mean rate of evolution of 0.92. The S2–S3 linker, proposed in a recent study to also bind PIP2, 31 is only moderately conserved. Interestingly, although most subdomains of the CD exhibit a low mean rate of evolution, two subdomains namely the S1–S2 linker and the S5-Pore linker, show substantially higher mean rates of evolution (2.5 and 1.9, respectively) than the rest of the CD.…”

Regulation of KCNQ/Kv7 family voltage-gated K + channels by lipids

“…Based on extensive structure-function studies of Kv7.1 and Kv7.2 complemented by molecular modeling, PIP 2 binding is known to be generally localized to the junction between the VSD and the PD, involving residues (most of which are positively charged) in the S2-S3 and S4-S5 linkers, and at the C-terminal (gate) end of the S6 helix [126, 142, 144–147]. Structural alignment between the PDs of the PIP 2 -Kir crystal structures and the Kv7.1 homology model [148] suggested the possibility of similar positioning of the PIP 2 binding surface in these two channels relative to the PD, as least in the channel open state [126].…”

“…However, whether 1 or 2 PIP 2 molecules can bind at a time at the VSD/S4-S5/S6 interface is not yet well established. One appealing possibility is that the stoichiometry is 1 PIP 2 per subunit, but that the location of the binding site in the closed state channel is different than in the open state channel [142, 146, 147]. Exactly how PIP 2 couples the VSD to the channel gate is a matter of intense study, as described above.…”