![The role of S4 mutations in the Shaker KV channel for the effect of some resin-acid derivatives.(A and B) Top (A) and side (B) views of the S4 helical-screw motion (gray arrows) during VSD activation. Endogenous gating charge arginines are denoted by solid or open blue circles (R362 [=R1], R365 [=R2], R368 [=R3], and R371 [=R4]). Additional charges in the 2R motif are denoted by semitransparent blue circles (M356R [=R-1], A359R [=R0]). Binding of a negatively charged compound (red triangle) in the cleft between S3 and S4 (the S3/S4 site) is hypothesized to favor S4 activation through an electrostatic effect that stabilizes the S4 activated state. The homotetrameric channel assembly is shown in A, with each subunit (SU) represented in a different color. A single subunit is shown in B (the pore domain and the VSD are from two different subunits). (C) Activation of Shaker channels proceeds in several activation steps between closed CX and open O states, during which the S4 helix moves in a ratchet-like upward and rotative movement (gray arrows). The purple circle marks the position of a bound lipid headgroup in the absence of compound in the open state. (D) 100 µM Wu50 at pH 9.0 shifted the G(V) of the Shaker KV channel with the 2R motif by −40.7 mV. (E)G(V) shifts for R362Q versus WT Shaker KV channels for four compounds. Solid symbols represent compounds for which the shift is similar in the WT and in the R362Q mutant, while empty symbols represent compounds for which the shift is larger in the mutant. The dotted line marks an equal shift in WT and R362Q. (F–H) Concentration-response curves (Eq. 3). Black, WT Shaker KV. Red, Shaker KV with the 2R motif. (F) Wu50, pH = 9.0. WT: EC50 = 29.1 ± 3.5 µM, ΔVMAX = −36.1 ± 1.3 mV, n = 3–5. 2R motif: EC50 = 30.0 ± 5.0 µM, ΔVMAX = −56.5 ± 2.6 mV, n = 4–10. (G) Wu161, pH = 7.4. WT: EC50 = 43.7 ± 6.3 µM, ΔVMAX = −13.9 ± 0.6 mV, n = 3–4. 2R motif: EC50 = 36.4 ± 9.0 µM, ΔVMAX = −45.5 ± 3.5 mV, n = 3–6. (H) Wu181, pH = 7.4. WT: EC50 = 1.6 ± 0.6 µM, ΔVMAX = −7.8 ± 0.5 mV, n = 2–3. 2R motif: EC50 = 6.1 ± 1.7 µM, ΔVMAX = −53.6 ± 4.0 mV, n = 4–5. All data mean ± SEM.](https://cdn.rupress.org/rup/content_public/journal/jgp/153/4/10.1085_jgp.202012676/4/jgp_202012676_fig1.png?Expires=1767771718&Signature=gIS6ghmtAxKxNnEVl77syGzaVaBTCFtO7ZMZU9VQsc816zRM3aod3G1XVPnKwVAsP8WgWrtA0IwmzdwrSXGL3-qt~CmseT-cTH7VIDIgDTsLqI-4T3omJuLqEghnvvL5bw0l79RAUp-HntEQ6g1veT7f~DdSuKyhRrlp03vVORuqU8acplStfJOEXKSEari0M7eMUU1gKqvQKY7JQvfJyEPDBIGn0VL-LBwijttAPOPOtqHqSBp4loFGNgBmYl3Yy7WYxutZblU~SVr6Bk9eX5gvAQaSFRAuVPBUAqmi5d6yY4hID42T47TH58zI1Yn1T0OtGvGvdxJqhr-C9oJFxg__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
The role of S4 mutations in the Shaker KV channel for the effect of some resin-acid derivatives. (A and B) Top (A) and side (B) views of the S4 helical-screw motion (gray arrows) during VSD activation. Endogenous gating charge arginines are denoted by solid or open blue circles (R362 [=R1], R365 [=R2], R368 [=R3], and R371 [=R4]). Additional charges in the 2R motif are denoted by semitransparent blue circles (M356R [=R-1], A359R [=R0]). Binding of a negatively charged compound (red triangle) in the cleft between S3 and S4 (the S3/S4 site) is hypothesized to favor S4 activation through an electrostatic effect that stabilizes the S4 activated state. The homotetrameric channel assembly is shown in A, with each subunit (SU) represented in a different color. A single subunit is shown in B (the pore domain and the VSD are from two different subunits). (C) Activation of Shaker channels proceeds in several activation steps between closed CX and open O states, during which the S4 helix moves in a ratchet-like upward and rotative movement (gray arrows). The purple circle marks the position of a bound lipid headgroup in the absence of compound in the open state. (D) 100 µM Wu50 at pH 9.0 shifted the G(V) of the Shaker KV channel with the 2R motif by −40.7 mV. (E)G(V) shifts for R362Q versus WT Shaker KV channels for four compounds. Solid symbols represent compounds for which the shift is similar in the WT and in the R362Q mutant, while empty symbols represent compounds for which the shift is larger in the mutant. The dotted line marks an equal shift in WT and R362Q. (F–H) Concentration-response curves (Eq. 3). Black, WT Shaker KV. Red, Shaker KV with the 2R motif. (F) Wu50, pH = 9.0. WT: EC50 = 29.1 ± 3.5 µM, ΔVMAX = −36.1 ± 1.3 mV, n = 3–5. 2R motif: EC50 = 30.0 ± 5.0 µM, ΔVMAX = −56.5 ± 2.6 mV, n = 4–10. (G) Wu161, pH = 7.4. WT: EC50 = 43.7 ± 6.3 µM, ΔVMAX = −13.9 ± 0.6 mV, n = 3–4. 2R motif: EC50 = 36.4 ± 9.0 µM, ΔVMAX = −45.5 ± 3.5 mV, n = 3–6. (H) Wu181, pH = 7.4. WT: EC50 = 1.6 ± 0.6 µM, ΔVMAX = −7.8 ± 0.5 mV, n = 2–3. 2R motif: EC50 = 6.1 ± 1.7 µM, ΔVMAX = −53.6 ± 4.0 mV, n = 4–5. All data mean ± SEM.