PMC

Mutant cycles cube of Sr²⁺-induced (50 mM) shifts. Data taken from .

Effect of Sr²⁺ on G(V) curves for chimeras measured as midpoint shifts. Data taken from . The hatched vertical lines indicate the mean shift value of the Kv1.2 and Kv2.1 channels, respectively.

Correlations in higher-order effects. (A) Alterations in Sr²⁺-induced G(V) shifts plotted versus alterations in V_1/2 caused by transplantation of extracellular linkers from Kv1.2 to Kv2.1. Based on data taken from (slope = −0.49, p < 0.0001). (B) Alterations in alterations of Sr²⁺-induced G(V) shifts plotted versus alterations in alterations of V_1/2 caused by transplantation of extracellular linkers from Kv1.2 to Kv2.1. Based on data from (slope = −0.53, p = 0.0001).

Sequences and structure of the extracellular linkers of a Kv1.2/2.1 channel. (A) Sequences of the extracellular linkers of Kv1.2 and Kv2.1 are presented. Negatively charged residues are red and positively charged residues are blue. The numbers 1–8 above the sequences denote the residues where the Kv2.1 channel is more positively charged than Kv1.2. (B) Extracellular view of the Kv1.2/2.1 chimera () is shown. The numbered residues in (A) are colored and numbered in the molecular structure. The three top gating charges in S4 (R1–R3) are colored in gold. To see this figure in color, go online.

Sr²⁺ effects on Kv1.2 channel opening kinetics. (A) Currents at a step from −80 mV to 0 mV in control solution are shown. Rise time measured as difference between times at 10% and 90% of steady-state value (t₁₀₋₉₀ = t_90% − t_10%). (B) Plots of t₁₀₋₉₀ versus voltage are shown. Additional 50 mM SrCl₂ (solid symbols) to the control solution (open symbols) shifts the curve along the voltage axis. The continuous lines are fitted exponential curves where the exponent and the plateau are constrained to be equal.

Midpoint voltage (V_1/2) among wild-type Kv channels as a function of the electric field components. (A) The influence of ψ_out on V_1/2 of seven Kv channels is shown. Thick continuous line is the estimated regression line (V_1/2 = 0.94 × φ_out + 32.7), surrounded by a shaded area representing a 95% confidence interval. The line was obtained by Deming regression (n = 53, r = 0.62). The dashed line represents a linear function with a slope of +1. (B) The influence of the residual field potential (ΔG₀ / F + ψ_in) on V_1/2 is shown. Thick continuous line is the estimated regression line (V_1/2 = −1.12 × (ΔG₀ / F + ψ_in) − 41.9), surrounded by a shaded area representing a 95% confidence interval. The line was obtained by Deming regression (n = 53, r = 0.70). The dashed line represents a linear function with a slope of −1.

Midpoint voltage (V_1/2) among mutant Kv channels as a function of electric field components. (A) Dependence on ψ_out when external linkers are mutated (n = 81 distributed among seven mutation sites, r = 0.87). Deming regression line shown by the thick continuous line. Slope is 0.75 and intercept is 24.2 mV. The dashed line shows a slope of +1. A 95% confidence band is represented by shaded gray areas. (B) Dependence on the residual field potential (ΔG₀ / F + ψ_in) is shown for the same channels as in (A). Slope and intercept of the Deming regression line are −1.15 and −36.4 mV, respectively (r = 0.16). The dashed line shows a slope of −1. A 95% confidence band is represented by shaded gray areas.

Similarity in the amino acid sequence of the studied Kv channels. Using the Smith-Waterman algorithm (), the segments were pairwise locally aligned and the fraction of identical amino acids was taken as a measure of similarity (see for details). The bars show the mean similarity between the seven Kv channels studied (21 pairwise comparisons). The white bars represent intracellular segments, the gray bars represent the transmembrane segments, the black bars represent the extracellular segments, and the striped bar represents the pore. The pore is the most conserved part, whereas the extra- and intracellular linkers are the least-conserved parts of the protein (p < 0.05). Significance assessed by bootstrapping. Error bars represent mean ± SE.

Effect of Sr²⁺ on steady-state conductance of wild-type Kv channels. (A) K currents through Kv1.2 channels are elicited from a holding voltage of –80 mV by voltage steps of 500 ms duration up to +60 mV in steps of 10 mV under control conditions and with 50 mM SrCl₂ added to the control solution. The dots mark the currents at 0 mV. One voltage pulse is illustrated below the current traces. (B) G(V) curves in control (open symbols) and 50 mM SrCl₂ (solid symbols) (n = 16) are shown. The conductance values were normalized to their respective maximal value. The error bars represent mean ± SE. (C) Summary of Sr²⁺-induced shifts of midpoint values (ΔV_1/2) of the G(V) curves from seven wild-type channels are shown.