Selectivity filter modalities and rapid inactivation of the hERG1 channel

Abstract

The human ether-á-go-go–related gene (hERG1) channel conducts small outward K+ currents that are critical for cardiomyocyte membrane repolarization. The gain-of-function mutation N629D at the outer mouth of the selectivity filter (SF) disrupts inactivation and K+-selective transport in hERG1, leading to arrhythmogenic phenotypes associated with long-QT syndrome. Here, we combined computational electrophysiology with Markov state model analysis to investigate how SF-level gating modalities control selective cation transport in wild-type (WT) and mutant (N629D) hERG1 variants. Starting from the recently reported cryogenic electron microscopy (cryo-EM) open-state channel structure, multiple microseconds-long molecular-dynamics (MD) trajectories were generated using different cation configurations at the filter, voltages, electrolyte concentrations, and force-field parameters. Most of the K+ permeation events observed in hERG1-WT simulations occurred at microsecond timescales, influenced by the spontaneous dehydration/rehydration dynamics at the filter. The SF region displayed conductive, constricted, occluded, and dilated states, in qualitative agreement with the well-documented flickering conductance of hERG1. In line with mutagenesis studies, these gating modalities resulted from dynamic interaction networks involving residues from the SF, outer-mouth vestibule, P-helices, and S5–P segments. We found that N629D mutation significantly stabilizes the SF in a state that is permeable to both K+ and Na+, which is reminiscent of the SF in the nonselective bacterial NaK channel. Increasing the external K+ concentration induced “WT-like” SF dynamics in N629D, in qualitative agreement with the recovery of flickering currents in experiments. Overall, our findings provide an understanding of the molecular mechanisms controlling selective transport in K+ channels with a nonconventional SF sequence.

Refinement of a Cryo-EM Structure of hERG: Bridging Structure and Function

“…This is different from the observation on the voltage-sensor flexibility found in a micro-seconds long MD simulation with the reported RMSDs reaching a plateau at around 5 Å (25). A study by Miranda et al was centered at the permeation across the PD and employed a strong biasing electrical field (750 mV) to enable ion movement which may impact VSD conformation (25). Applications of electrical field are known to induce significant conformational plasticity of VSD in the previously studied Kv channels (44).…”

“…The gating charge associated with hERG activation is also surprisingly small compared to the gating charge of the Kv1.2 channel, suggesting a different mechanism (5). MD simulations performed with the cryo-EM structure embedded in a lipid bilayer also highlighted a loosely packed VSD configuration (25), in line with the observation of Subbiah et al (11) However, the VSD in the cryo-EM structure is tightly packed against the PD. Shi et al also emphasized the importance of further refinement of this cryo-EM structure due to the uncertain positioning of D509, which acts as a proton sensor and stabilizes the open state (18).…”

“…We observed a compact configuration of VSD regardless of the environment during the fitting. This is different from the observation on the voltage-sensor flexibility found in a micro-seconds long MD simulation with the reported RMSDs reaching a plateau at around 5 Å (25). A study by Miranda et al was centered at the permeation across the PD and employed a strong biasing electrical field (750 mV) to enable ion movement which may impact VSD conformation (25).…”

“…Then we used the Molecular Dynamics Flexible Fitting (MDFF) (26)(27)(28) technique for structure refinement. We extracted the starting structure for fitting from our multi-microsecond MD simulation where the protein was embedded in a bilayer mimicking native environment (25). We extracted the structure towards the end of the simulation where the VSD was in fully expanded configuration.…”

K2Pchannel C-type gating involves asymmetric selectivity filter order-disorder transitions

“…The SF1 structural transformation affects a post-filter residue Asn147, a position that modulates C-type inactivation in four-fold symmetric potassium channels 39,40 and that undergoes similar changes in hERG simulations 41 . Hence, this class of C-type inactivation conformational change appears to be shared with four-fold symmetric potassium channels.…”