Figure 2

Epilepsy mutations cluster at CaM-binding helix B and the helix B-C linker of K_v7.2. (a) Tetrameric human K_v7.2 (ribbons) in complex with four CaM subunits (transparent green surfaces) was modeled based on the structure of Xenopus K_v7.1 (Protein Data Bank: 5VMS)²⁸. Sites of pathogenic mutations in K_v7.2 C-terminal tail are highlighted with colored spheres on the C-alpha atoms of one subunit: mild or BFNE (blue), uncertain severity (purple), severe or EE (red). In addition to epilepsy mutation hotspots (S4, the pore loop, and S6), “severe or EE” mutations cluster near the inner leaflet of the plasma membrane and the C-terminus of S6 at the base of the gate. (b,c) The MHF statistical algorithm on the intracellular C-terminal tail of K_v7.2 identified helix B and the helix B-C linker as hotspots for all pathogenic epilepsy mutations (brackets, **p < 0.01, ***p < 0.005, Supplementary Table S2) (b) and EE mutations (brackets, *p < 0.05, ***p < 0.005, Supplementary Table S3) (c). (d) Location of amino acids mutated in mild or BFNE (blue), uncertain severity (purple), or severe or EE (red) in helix A containing a consensus IQ motif for binding CaM (underlined), helix B, and the helix B-C linker of K_v7.2. (e) Mutated amino acids are highlighted on a model of K_v7.2 helix A and B (grey) bound to Ca²⁺-CaM (green), which was modeled after the crystal structure of chimeric K_v7.3 helix A - K_v7.2 helix B protein in complex with Ca²⁺-CaM (Protein Data Bank: 5J03)³². Side chains for residues with pathogenic mutations are colored spheres: mild or BFNE (blue), uncertain severity (purple), severe or EE (red). (f) Predicted changes in Ca²⁺-CaM binding energy of pathogenic K_v7.2 missense mutations within helices A and B. The higher the energy the weaker the predicted affinity for Ca²⁺-CaM. (g) Positively charged basic residues at proximal helix A, distal helix B and the helix B-C linker (purple) are located close to basic residues from the S2–3 linker (yellow), S4, S6 (blue), and CaM (green).