In silico analysis of conformational changes induced by mutation of aromatic binding residues: consequences for drug binding in the hERG K+ channel

Abstract

Pharmacological inhibition of cardiac hERG K(+) channels is associated with increased risk of lethal arrhythmias. Many drugs reduce hERG current by directly binding to the channel, thereby blocking ion conduction. Mutation of two aromatic residues (F656 and Y652) substantially decreases the potency of numerous structurally diverse compounds. Nevertheless, some drugs are only weakly affected by mutation Y652A. In this study we utilize molecular dynamics simulations and docking studies to analyze the different effects of mutation Y652A on a selected number of hERG blockers. MD simulations reveal conformational changes in the binding site induced by mutation Y652A. Loss of π-π-stacking between the two aromatic residues induces a conformational change of the F656 side chain from a cavity facing to cavity lining orientation. Docking studies and MD simulations qualitatively reproduce the diverse experimentally observed modulatory effects of mutation Y652A and provide a new structural interpretation for the sensitivity differences.

Figure 1. Location and flexibility of putative aromatic binding residues in hERG.

(A) π-π-stacking interactions between binding determinants Y652 and F656, located on helix S6 and residues F619 (P-helix) and F557 (S5 helix). Side chains are shown as green sticks (B). χ₁/χ₂ plot of Y652 and F656 (C) obtained from 50 ns MD simulations.

Figure 2. Stability of WT and mutant hERG channels.

(A) Backbone RMSD of the Y652A (blue) and the F656A mutant (brown) compared to WT channel (black). (B) Comparison of the root mean square fluctuations (RMSF) for WT and mutant channels. Only the P-helix and connecting loops are shown. (C) RMSF of S6 helix.

Figure 3. Side chain rearrangements of F656 induced by mutation Y652A.

(A) χ₁/χ₂ side-chain angles of F656 for WT (black) and Y652A (blue). The green and blue arrows indicate the approximate conformations of the F656 side chains shown in B and C. (B) Representative side-chain conformations of WT and Y652A mutant (C) channel snapshots taken from MD simulations. (D) F656 χ₁ dihedral angles for WT (black) and Y652A (blue) in all four domains as a function of time.

Figure 4. Structures of hERG blockers examined in this study.

Drugs are clustered into three groups: group 1 (orange frame) includes blockers which are relatively insensitive to mutation Y652A^22,28–30, group 2 (green frame) shows Y652 sensitive drugs^10,18,22,31 and group 3 (blue frame) shows capsaicin whose affinity is increased by mutation Y652A³².

Figure 5. Docking (cyan transparent sticks) and MD poses.

at the end of the simulation (blue sticks) of bepridil (AB), thioridazine (CD), propafenone (EF), cisapride (GH), terfenadine (IJ) and ibutilide (KL) in WT and Y652A (from left to right). Y652 and F656 are shown as green lines; A652 is shown as orange lines. The arrow displays the movement of the heptyl chain of ibutilide.

Figure 6. Interactions of capsaicin with the selectivity filter in WT (A) and Y652A mutant (B).

Selectivity filter residues involved in capsaicin binding are shown as green sticks; residues of the TSV motif not interacting with capsaicin are shown as grey lines. Hydrogen bonds are depicted as black dots.