Common Structural Pattern for Flecainide Binding in Atrial-Selective Kv1.5 and Nav1.5 Channels: A Computational Approach

Figures

• 

  Scheme 1. General workflow describing steps in methodology. In color blue, software are displayed. The processes are indicated in yellow. PDB: Protein Data Bank, BS: binding site, MD: molecular dynamics.

• 

  Figure 1. Representation of flecainide binding mode in Kv1.5 channel. (a) Ribbon representation of Kv1.5 pore domain; each subunit is shown in a different color: orange (subunit A), pink (subunit B), blue (subunit C), green (subunit D), and flecainide bound in the inner cavity is depicted in ball and sticks. Residues forming the selectivity filter are labeled and indicated in gray boxes. (b) Side-chain representation of residues in contact with flecainide (distance ≤ 5 Å) having a frequency of interaction ≥70% along MD simulations. Residue names and numbers are indicated. The tertiary structure used corresponds to a frame from the holo MD trajectories. (c) Flecainide representation in 2D format. Flecainide and residues are colored per element with carbon atoms in dark gray, oxygen atoms in red, nitrogen atoms in blue, sulfur atoms in yellow, hydrogens in light gray and fluorine in green.

• 

  Figure 2. Flecainide interactions in Kv1.5 along MD simulations. The residue contacts and interactions were computed along the three 100 ns MD simulations’ replicas concatenated. (a) Residues in contact within 5Å of flecainide with a frequency of ≥70%. (b) Residues interacting with flecainide and the nature of such interactions predicted using PLIP software. Only the residues with number of interactions ≥20% of the maximum value of its type are considered.

• 

  Figure 3. Representation of flecainide binding mode in Nav1.5 channel. (a) Ribbon representation of Nav1.5 pore domain; each domain is shown in a different color: orange (DI), pink (DII), blue (DIII) and green (DIV), and flecainide bound in the inner cavity is depicted in ball and sticks. Residues forming the selectivity filter are labeled and indicated in gray boxes. (b) Side-chain representation of residues in contact with flecainide (distance ≤ 5 Å) having a frequency of interaction ≥70% along MD simulations. Residue names and numbers are indicated. The tertiary structure used corresponds to a frame from the holo MD trajectories. (c) Flecainide representation in 2D format. Flecainide and residues are colored per element with carbon atoms in dark gray, oxygen atoms in red, nitrogen atoms in blue, sulfur atoms in yellow, hydrogens in light gray and fluorine in green.

• 

  Figure 4. Flecainide interactions in Nav1.5 along MD simulations. The residue contacts and interactions were computed along the 100 ns MD simulations. (a) Residues in contact within 5 Å of flecainide with a frequency of ≥70%. (b) Residues interacting with flecainide and the nature of such interactions were predicted using PLIP software. Only the residues with number of interactions ≥20% of the maximum value of its type are considered.

• 

  Figure 5. Physicochemical characterization of flecainide binding site (BS). The flecainide BSs were computed in terms of (a) volume, (b) area, (c) hydrophobicity proportion, (d) Monera hydrophobicity score and (e) the proportion of apolar atoms in Kv1.5 and Nav1.5 along three replicas of molecular dynamics simulations. The colored points and vertical lines stand for the median value and the interquartile range (IQR) of each property measured, respectively. **** represent p-value adjusted (Bonferroni) = 0 on a Wilconsox Rank Sum test.

• 

  Figure 6. Structural alignment of equivalent residues in flecainide BS. The equivalent residues of flecainide BS in (a) Kv1.5, (b) Nav1.5 and (c) their superposition according to PocketAlign comparison are displayed. Only residues with a contact frequency ≥70% are shown. In (c), the equivalences are numbered from 1 to 10 similar to Table 4. The underlined residues were not previously reported in flecainide BS. In addition, the distances between the centers of mass of the equivalent residues are appended.

• 

  Figure 7. Mapping onto tertiary structure the equivalent residues in flecainide BS. Surface representation of the equivalent residues in flecainide BS in (a) Kv1.5 and (b) Nav1.5 as predicted by PocketAlign (listed in Table 4). Residues from the hydrophobic patch are shown in gray surface and they are labeled inside a gray box meanwhile those from the hydrophilic region are highlighted in light-blue and included in a box with similar color. The rest of equivalent residues are presented in green surface and they are enclosed in a green box. The equivalent residues with a similar physicochemical nature are highlighted in bold.––The * is used to denote residues relevant for flecainide binding according to mutagenesis in Kv1.5 and cryo-EM flecainide-bound structure in Nav1.5. The underlined residues were not previously reported in flecainide BS. Flecainide is shown in sticks and colored per element with oxygen atoms in red, nitrogen atoms in blue, fluorine atoms in green and carbon atoms in sand color.