'C-type' closed state and gating mechanisms of K2P channels revealed by conformational changes of the TREK-1 channel

Abstract

Two-pore domain potassium (K2P) channels gate primarily within the selectivity filter, termed 'C-type' gating. Due to the lack of structural insights into the nonconductive (closed) state, 'C-type' gating mechanisms remain elusive. Here, molecular dynamics (MD) simulations on TREK-1, a K2P channel, revealed that M4 helix movements induce filter closing in a novel 'deeper-down' structure that represents a 'C-type' closed state. The 'down' structure does not represent the closed state as previously proposed and instead acts as an intermediate state in gating. The study identified the allosteric 'seesaw' mechanism of M4 helix movements in modulating filter closing. Finally, guided by this recognition of K2P gating mechanisms, MD simulations revealed that gain-of-function mutations and small-molecule activators activate TREK-1 by perturbing state transitions from open to closed states. Together, we reveal a 'C-type' closed state and provide mechanical insights into gating procedures and allosteric regulations for K2P channels.

Keywords: C-type; K2P; gating; selectivity filter; state.

Figure 1

Conformation transitions of the TREK-1 channel from the ‘up’ to ‘deeper-down’ structure in the SMD simulation. (A) Schematic diagram of stretching M4 helix in the simulation. (B) Changes in ‘fenestration’ (blue), ‘zipper’ (green), and ‘expansion’ (yellow) distances in the simulation. (C) Superimposition of the state 2 conformation (gray) and the ‘up’ (blue) or ‘down’ (orange) structures. (D) Pore radius of the initial and final snapshots of the simulation. (E) Definitions of the distant SF diameter and TM diameter. (F) Cluster analysis of the SMD trajectories with the distant SF diameter and TM diameter as two variables. (G) Typical conformations. (H) The ‘seesaw’ movements of M4 and the influences of the movements on the networks within the filter region. Potassium ions and water molecules in the SF region are shown as purple or red spheres. Key residues are highlighted by colored sticks. Potential hydrogen bonds are indicated by dotted lines. Directions of conformational changes are indicated by dashed arrows.

Figure 2

Conformation transitions of the TREK-1 channel from the ‘down’ to ‘deeper-down’ structure in the MD simulation. (A) Cluster analysis of the MD simulation trajectory with the distant SF diameter and TM diameter as two variables. (B) Typical conformations. (C) The ‘seesaw’ movements of M4 and the influences of the movements on the networks within the filter region.

Figure 3

Conformation transitions of the channel in the SMD simulations of the gain-of-function mutants W275S and G137I. (A) Mapping of the mutant W275S in the ‘up’ state of the TREK-1 channel. The Ser residue is shown. (B) Cluster analysis of the W275S SMD trajectory with the distant SF diameter and TM diameter as two variables. (C) Typical conformations of the W275S SMD trajectory. (D) The ‘seesaw’ movements of M4 and the influences of the movements on the networks within the filter region. (E) Time traces of distances between the hydroxyl group of S275 and the hydroxyl group of T141 during the W275S SMD simulation. (F) Mapping of the mutant G137I in the ‘up’ state of the TREK-1 channel. A few of the residues are highlighted by the green sticks. (G) Cluster analysis of the G137I SMD trajectory with the distant SF diameter and TM diameter as two variables. (H) Typical conformations of the G137I SMD trajectory. (I) The ‘seesaw’ movements of M4 and the influences of the movements on the networks within the filter region. The display form is shown as in D. (J) Time traces of distances between the indole NH group of W275 and the hydroxyl group of T141 during the G137I SMD simulation.

Figure 4

Conformation transitions of the TREK-1 channel in the SMD simulation of the ML335/TREK-1 complex. (A) Cluster analysis of the ML335/TREK-1 complex SMD trajectory with the distant SF diameter and TM diameter as two variables. (B) Typical conformations of the ML335/TREK-1 complex SMD trajectory. (C) The ‘seesaw’ movements of M4 and the influences of the movements on the networks within the filter region. (D) Time traces of distances between the indole NH group of W275 and the hydroxyl group of T141 during the ML335/TREK-1 system (liganded) or the TREK-1 system (unliganded) SMD simulation.