The Structural Basis of IKs Ion-Channel Activation: Mechanistic Insights from Molecular Simulations

Abstract

Relating ion-channel (iCh) structural dynamics to physiological function remains a challenge. Current experimental and computational techniques have limited ability to explore this relationship in atomistic detail over physiological timescales. A framework associating iCh structure to function is necessary for elucidating normal and disease mechanisms. We formulated a modeling schema that overcomes the limitations of current methods through applications of Artificial Intelligence Machine Learning (ML). Using this approach, we studied molecular processes that underlie human IKs voltage mediated gating. IKs malfunction underlies many debilitating and life-threatening diseases. Molecular components of IKs that underlie its electrophysiological function include KCNQ1 (pore forming tetramer) and KCNE1 (auxiliary subunit). Simulations, using the IKs structure-function model, reproduced experimentally recorded saturation of gating charge displacement at positive membrane voltages, twostep voltage sensor (VS) movement shown by fluorescence, iCh gating statistics, and current-voltage (I-V) relationship. New mechanistic insights include -(1) pore energy profile determines iCh subconductance (SC), (2) entire protein structure, not limited to the pore, contributes to pore energy and channel SC, (3) interactions with KCNE1 result in two distinct VS movements, causing gating charge saturation at positive membrane voltages and current activation delay, and (4) flexible coupling between VS and pore permits pore opening at lower VS positions, resulting in sequential gating. The new modeling approach is applicable to atomistic scale studies of other proteins on timescales of physiological function.

Modulation of the IKSchannel by PIP2requires two binding sites per monomer

“…As a matter of fact, a recent computational study, which consisted in the use of machine learning methods to generate a conformational space of IKS channels, have yielded to the design of a structure-based predictor of IKS channel experimental properties including its subconductance and gating current (Ramasubramanian and Rudy, 2018b). The two sequential translations and rotations of S4 and the rotations of KCNE1 leading to VSD activation we predicted from our models are supported by the results obtained with this structure-based predictor.…”

“…Since beside KCNQ1EM (Sun and MacKinnon, 2017), and the very recently published Cryo-EM structures of the human KV7.1 AO channel in the presence of PIP2 (Sun and MacKinnon, 2020) , no high-resolution structure of the IKS channel is available, we and others turn to molecular modeling. Quite surprisingly, among the molecular constructs of the IKS channel that have been published the last decade (Gofman et al, 2012;Kang et al, 2008;Ramasubramanian and Rudy, 2018a;Xu et al, 2013), very few were modeled in presence of PIP2. Moreover, those which were modeled with the lipid aimed at validating experiments were limited to the identification of the PIP2 binding sites (Eckey et al, 2014), without providing any molecular insight about the way the lipid interacts with KV7.1 or with its KCNE1 subunits.…”

I Ks ion-channel pore conductance can result from individual voltage sensor movements

“…1B, a simplified version of this allosteric model of gating is shown, where VSs undergo two transitions to full activation, and in the presence or absence of KCNE1 each VS state is more or less coupled to corresponding open states of the channel (13,27). Recent support for an allosteric I Ks gating pathway comes from the modeling studies of Ramasubramanian and Rudy (28) in which pore openings to small or intermediate subconductance levels are shown to be possible over a range of early and intermediate S4 translations.…”

Molecular determinants of the modulation of the VSD-PD coupling mechanism of the KV7.1 channel by the KCNE1 ancillary subunits

“…The inner pore surface contains the S4-S5L residues engaged in state-independent VdW interactions with both S5 and S6 residues from the same subunit, which suggests a pulling of S6 segments from the pore axis toward the inner membrane surface upon S4 and S4-S5L movements during the VSD activation. The specificity in our IKS models in relation with the various IKS models reported so far (Eckey et al, 2014;Jalily Hasani et al, 2017Kang et al, 2008;Kuenze et al, 2020b;Ramasubramanian and Rudy, 2018;Xu and Rudy, 2018) resides in the fact that the presence of PIP2 is acting as a hook to ensure the leverage effect of S4-S5L on S6 cytoplasmic region. Indeed, R249 from S4-S5L is interacting with PIP2 intra in all models, while PIP2 intra itself progressively anchors S2-S3L during S4 activation, while maintaining the S4-S5L close to the inner membrane surface.…”

“…These strips are assumed to pull PD segments farther from the pore axis than in KV7.1 AO state, which can explain the pore radii differences observed between KV7.1 and IKS AO models (Deyawe Kongmeneck et al, 2021). Interestingly, a recent computational study, which consisted in the use of machine learning methods to generate a conformational space of IKS channels, have yielded to the design of a structure-based predictor of IKS channel experimental properties including its subconductance and gating current (Ramasubramanian and Rudy, 2018). The two sequential translations and rotations of S4 and the rotations of KCNE1 leading to VSD activation we predicted from our models are supported by the results obtained with this structure-based predictor.…”