Atomistic basis of opening and conduction in mammalian inward rectifier potassium (Kir2.2) channels

Abstract

18 simulations, single channel recordings. 19 20 21 22 Potassium ion conduction through open potassium channels is essential to 23 control of membrane potentials in all cells. To elucidate the open conformation 24 and hence the mechanism of K + ion conduction in the classical inward rectifier 25 Kir2.2, we introduced a negative charge (G178D) at the crossing point of the inner 26 helix bundle (HBC), the location of ligand-dependent gating. This 'forced open' 27 mutation generated channels that were active even in the complete absence of 28 phosphoinositol-4,5-bisphosphate (PIP 2 ), an otherwise essential ligand for Kir 29 channel opening. Crystal structures were obtained at a resolution of 3.6 Å without 30 PIP 2 bound, or 2.8 Å in complex with PIP 2 . The latter revealed a slight widening at 31 the HBC, through backbone movement. Molecular dynamics (MD) simulations 32 showed that subsequent spontaneous wetting of the pore through the HBC gate 33 region allowed K + ion movement across the HBC and conduction through the 34 channel. Further simulations reveal atomistic details of the opening process and 35 highlight the role of pore lining acidic residues in K + conduction through Kir2 36 channels.

Determinants of trafficking, conduction, and disease within a K+ channel revealed through multiparametric deep mutational scanning

“…Intriguingly, this network involves interfaces between adjacent subunits (N-terminus<>βLM loop interface, βI<>βCD loop, βD<>βEG loop). Subunit interactions determining gating is consistent with recent crystal structures of a forced-opened Kir2.2 in which inner helix gate opening requires CTD subunits to move relative to each other 77 .…”

Defining how multiple lipid species interact with inward rectifier potassium (Kir2) channels

“…However, the MARTINI model fixes the secondary structure of the protein, which limits the opportunity to probe the effects of lipid interactions of conformational changes underlying channel gating would proceed, and thus on how lipid complexity might influence channel activity. Root-mean-squared fluctuations of the protein in simulations with and without PIP 2 are in agreement with the proposed effect of PIP 2 on channel opening; however, further work is required to understand how the interplay of PIP 2 and PS interactions might affect channel gating, especially as progress has been made recently in understanding the effect of phospholipids on gating of Kir2.2 channels via a combined structural and computational approach (50).…”

Improved Parameterization of Phosphatidylinositide Lipid Headgroups for the Martini 3 Coarse Grain Force Field

“…These channels, were found to be regulated by several lipid species, especially PI(4,5)P2 which was found to activate mammalian Kir channels 100,101 . Resolved crystal structures of Kir channels with bound PI(4,5)P2 59 , as well as previous Martini 2 simulation studies 23,62 , show a binding pocket from which the PI(4,5)P2 headgroup may interact with both the transmembrane and cytoplasmic domain, favoring the open conformation.…”

“…A Kir2.2 channel was positioned roughly as described in previous simulation studies 23,62 . Over the course of the simulation, PI(4,5)P2 lipids diffused freely and converged upon a site common to all subunits (Figure 8D and E) which overlaps with the PI(4,5)P2 binding site detected by X-ray crystallography 59 (Figure 8C). Indeed, PI(4,5)P2 lipids were found bound to the described Kir binding site (Figure 8B), and established amino acid contacts (K189, K186, K188, K183, R78, R80, W79) which are in agreement with those observed in the resolved X-ray structure 59 as well as in previous Martini 2 studies 23,62 .…”

“…Two different proteins were also studied. Protein structures were obtained from the Protein Data Bank 57 (PDB) (PLCδ1 PH domain PDB: 1MAI 58 , and the Kir2.2 channel PDB: 6M84 59 ). All CG protein models were constructed using Martinize2 60 , with an applied elastic network with a bond force constant of 700 kJ/mol and a cut off distance of 0.8 nm.…”