Electrostatic model of S4 motion in voltage-gated ion channels - PubMed (original) (raw)

Comparative Study

Electrostatic model of S4 motion in voltage-gated ion channels

Harold Lecar et al. Biophys J. 2003 Nov.

Abstract

The S4 transmembrane domain of the family of voltage-gated ion channels is generally thought to be the voltage sensor, whose translocation by an applied electric field produces the gating current. Experiments on hSkMI Na(+) channels and both Shaker and EAG K(+) channels indicate which S4 residues cross the membrane-solution interface during activation gating. Using this structural information, we derive the steady-state properties of gating-charge transfer for wild-type and mutant Shaker K(+) channels. Assuming that the energetics of gating is dominated by electrostatic forces between S4 charges and countercharges on neighboring transmembrane domains, we calculate the total energy as a function of transmembrane displacement and twist of the S4 domain. The resulting electrostatic energy surface exhibits a series of deep energy minima, corresponding to the transition states of the gating process. The steady-state gating-charge distribution is then given by a Boltzmann distribution among the transition states. The resulting gating-charge distributions are compared to experimental results on wild-type and charge-neutralized mutants of the Shaker K(+) channel.

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Figures

FIGURE 1

Geometry of one voltage-sensing subunit, S1–S4, in the resting (left column) and activated states (right column). Here we illustrate the fully resting state a and the final salt-bridge state d both in Fig. 2. State e is related to state d by a pure axial translation with no new salt bridges being formed. (A, B) S1–S3 is pictured as an amorphous, gray structure surrounding an _α_-helical S4. Acidic, negative amino acids are shown in blue and placed in proximity to the red basic residues of S4, R362–K380. The extracellular space is at the top of the molecule, and R362 is closest to this space. The region from Z = _z_out to 0 corresponds roughly to the solvent-inaccessible portion of S4 (Mannuzzu et al., 1996; Larsson et al., 1996). The length from Z = 0 to _z_in corresponds to a region of S4 that is accessible to cysteine modification, but that most likely supports an electric field in the presence of a transmembrane potential. (C, D) A more precise representation of the charge geometry. Red S4 charges are pictured over a green cylinder for clarity. This cylinder is aligned along the _z_-axis, and the positive charges are placed at a radius of 10 Å. The fixed positions of the negative charges in both the resting and activated states were determined to be E283: −10.33, −11.75, and 12.6; E293: 15.28, −0.91, and 4.52; and D316: 9.13, 12.92, and 0.28. S4 charges participating in salt bridges are labeled. Key charges on S2−S3 are labeled in C. S4 undergoes an axial translation of 13.5 Å and a clockwise rotation of 180° (when viewed from the extracellular medium) in going from C to D. (E, F) A projection of the basic and acidic charge positions in C and D into the _x_-y plane. Two salt bridges are made in the resting state (indicated by gray ellipses), and three are made in the activated state. The positions of E283, E293, and D316 remain the same in both graphs. All lengths are in Ångstroms.

FIGURE 2

Energy landscapes of a wild-type subunit during independent translation, z, and twist, ω. (A) Total electrostatic energy (W) as a function of the position of S4. A clear valley is seen corresponding to the screw-helical twist motion of S4. (B) Energy contour plot, showing more clearly that there is a diagonal path from minimum to minimum in this energy valley. These minima are sequentially labeled a, b, c, d, and e. The minima a corresponds to the resting conformation whereas e corresponds to the fully activated state. Although a_–_d nearly fall along a diagonal path, e does not. (C) Energy barrier profile along the reaction path in B from states a to e. Transitions from well to well correspond to a stochastically lurching helical screw motion. The total electrostatic energy (solid line) is plotted along with the image force energy (dashed red) and the Coulomb energy (dashed green). (D) The reaction path in C has been drawn for several values of the membrane potential V = −90, −45, 0, 45, and 90 mV. The −90 mV contour is drawn in red for clarity. At hyperpolarized values, the resting state a is stabilized with respect to the outer states d and e. As the membrane potential depolarizes, the free energy minima shifts to the outer states, driving the translocation of the voltage sensor.

FIGURE 3

Effects of various charge neutralizations on the energy profile along the reaction path, with the right panels showing the approximate salt-bridge linkages between S4 charges and the countercharges in each of the four minima. As the position of neutralized charge is moved, different close-approach bridges are missing and the corresponding energy states are destabilized. (A) Wild-type ShB-IR channels. (B) Neutralization of the negative charge E283Q destabilizes the outer wells with respect to the inner wells a and b. In general the electrostatic energy of the system is greatly elevated upon charge neutralization of the negative charges. (C) D316N is the most severe charge neutralization in terms of overall energetics; however, the relative number of salt contacts made in each of the states is invariant, resulting in an energy profile that is similar to wild-type. (D) The most extracellular S4 charge, R362Q, is neutralized, and the deactivated state is destabilized. (E) R365Q is neutralized, stabilizing the middle states b and c with respect to the resting and activated states. This gives rise to the anomalous voltage-dependence in the gating charge Q seen in Fig. 4 E. The dashed curve in each panel is the wild-type energy, shown for comparison.

FIGURE 4

Voltage-dependence of gating for wild-type and mutant channels. Comparison with experimental data of Seoh et al. (1996) (diamonds) and theory (solid line). For each channel we show gating-charge displacement, Q, as a function of voltage, V. Experimental data for Q were taken directly from Seoh et al. (1996) using the program Data Thief II (Bas Tummers, Eindhoven, The Netherlands). (A) Wild-type ShB-IR channels, (B) E283Q neutralized, (C) D316N neutralized, (D) R362Q neutralized, and (E) R365Q neutralized. Fits to the data were performed as explained in the text, and all parameters are featured in Table 1. The original data for mutant R362Q was normalized to unity due to an inability to calibrate the currents. Here we have scaled the original data by a factor of 9.3 to compare with results from the model.

FIGURE 5

Reaction paths, state-dependent probabilities, and conductances for wild-type (top panels) and R365Q channels (bottom panels). (A, B) Contour plots of the total electrostatic energy with the four most significant wells labeled (red asterisk). Note that the reaction path for R365Q is nearly a pure axial translation with little rotation about the S4 axis. (C, D) State-dependent probabilities corresponding to the energy minima in the contour maps in A and B. Wild-type channels are nearly a pure two-state system between wells a and e. Subsidiary states reach no higher than 10% occupancy. Meanwhile, states b and c are significantly populated, at ∼40%, for the mutant R365Q. Moreover, even at very hyperpolarized potentials, state a is only at 80% occupancy. (E, F) Conductance from state-dependent probabilities. The experimental data of Seoh et al. (1996) (circles) is compared with theory (solid line). Experimental data for the open probabilities, _P_o, were taken directly from Seoh et al. (1996) using Data Thief II. The theory curves were generated using Eq. 10, with a value of κ = 0.76. When S4 was in state d or e it was assumed that the concerted transition could occur; therefore, the sum of the probabilities of being in states d and e were raised to the fourth power in Eq. 10. There is little deviation in the shapes of the G(V) curves if the concerted transitions can only occur from state e. Note that G(V) curves were not fit to the model; the excellent agreement between theory and experiment is a direct consequence of first fitting the gating-charge data and then assuming subunit cooperativity is required for channel opening.

References

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Electrostatic model of S4 motion in voltage-gated ion channels - PubMed (original) (raw)