scholarly journals Allosteric Voltage Gating of Potassium Channels II

1999 ◽  
Vol 114 (2) ◽  
pp. 305-336 ◽  
Author(s):  
Frank T. Horrigan ◽  
Richard W. Aldrich
Keyword(s):  
Preceding Paper ◽  
Voltage Sensor ◽  
Voltage Gating ◽  
Total Charge ◽  
Charge Movement ◽  
Gating Currents ◽  
Open Probability ◽  
K Channels ◽  
Virtual Absence ◽  
Gating Charge

Large-conductance Ca2+-activated K+ channels can be activated by membrane voltage in the absence of Ca2+ binding, indicating that these channels contain an intrinsic voltage sensor. The properties of this voltage sensor and its relationship to channel activation were examined by studying gating charge movement from mSlo Ca2+-activated K+ channels in the virtual absence of Ca2+ (<1 nM). Charge movement was measured in response to voltage steps or sinusoidal voltage commands. The charge–voltage relationship (Q–V) is shallower and shifted to more negative voltages than the voltage-dependent open probability (G–V). Both ON and OFF gating currents evoked by brief (0.5-ms) voltage pulses appear to decay rapidly (τON = 60 μs at +200 mV, τOFF = 16 μs at −80 mV). However, QOFF increases slowly with pulse duration, indicating that a large fraction of ON charge develops with a time course comparable to that of IK activation. The slow onset of this gating charge prevents its detection as a component of IgON, although it represents ∼40% of the total charge moved at +140 mV. The decay of IgOFF is slowed after depolarizations that open mSlo channels. Yet, the majority of open channel charge relaxation is too rapid to be limited by channel closing. These results can be understood in terms of the allosteric voltage-gating scheme developed in the preceding paper (Horrigan, F.T., J. Cui, and R.W. Aldrich. 1999. J. Gen. Physiol. 114:277–304). The model contains five open (O) and five closed (C) states arranged in parallel, and the kinetic and steady-state properties of mSlo gating currents exhibit multiple components associated with C–C, O–O, and C–O transitions.

scholarly journals Effective gating charges per channel in voltage-dependent K+ and Ca2+ channels.

1996 ◽  
Vol 108 (3) ◽  
pp. 143-155 ◽  
Author(s):  
F Noceti ◽  
P Baldelli ◽  
X Wei ◽  
N Qin ◽  
L Toro ◽  
...  
Keyword(s):  
Ionic Current ◽  
Variance Analysis ◽  
Beta Subunit ◽  
Charge Movement ◽  
Gating Currents ◽  
Open Probability ◽  
K Channels ◽  
Voltage Dependent ◽  
Pore Opening ◽  
Ca2 Channels

In voltage-dependent ion channels, the gating of the channels is determined by the movement of the voltage sensor. This movement reflects the rearrangement of the protein in response to a voltage stimulus, and it can be thought of as a net displacement of elementary charges (e0) through the membrane (z: effective number of elementary charges). In this paper, we measured z in Shaker IR (inactivation removed) K+ channels, neuronal alpha 1E and alpha 1A, and cardiac alpha 1C Ca2+ channels using two methods: (a) limiting slope analysis of the conductance-voltage relationship and (b) variance analysis, to evaluate the number of active channels in a patch, combined with the measurement of charge movement in the same patch. We found that in Shaker IR K+ channels the two methods agreed with a z congruent to 13. This suggests that all the channels that gate can open and that all the measured charge is coupled to pore opening in a strictly sequential kinetic model. For all Ca2+ channels the limiting slope method gave consistent results regardless of the presence or type of beta subunit tested (z = 8.6). However, as seen with alpha 1E, the variance analysis gave different results depending on the beta subunit used. alpha 1E and alpha 1E beta 1a gave higher z values (z = 14.77 and z = 15.13 respectively) than alpha 1E beta 2a (z = 9.50, which is similar to the limiting slope results). Both the beta 1a and beta 2a subunits, coexpressed with alpha 1E Ca2+ channels facilitated channel opening by shifting the activation curve to more negative potentials, but only the beta 2a subunit increased the maximum open probability. The higher z using variance analysis in alpha 1E and alpha 1E beta 1a can be explained by a set of charges not coupled to pore opening. This set of charges moves in transitions leading to nulls thus not contributing to the ionic current fluctuations but eliciting gating currents. Coexpression of the beta 2a subunit would minimize the fraction of nulls leading to the correct estimation of the number of channels and z.

scholarly journals Specificity of Charge-carrying Residues in the Voltage Sensor of Potassium Channels

2004 ◽  
Vol 123 (3) ◽  
pp. 205-216 ◽  
Author(s):  
Christopher A. Ahern ◽  
Richard Horn
Keyword(s):  
Potassium Channels ◽  
Electric Field ◽  
Protein A ◽  
Membrane Lipid ◽  
Voltage Sensor ◽  
Charge Movement ◽  
Gating Currents ◽  
Gating Charge ◽  
Channel Opening ◽  
Positively Charged

Positively charged voltage sensors of sodium and potassium channels are driven outward through the membrane's electric field upon depolarization. This movement is coupled to channel opening. A recent model based on studies of the KvAP channel proposes that the positively charged voltage sensor, christened the “voltage-sensor paddle”, is a peripheral domain that shuttles its charged cargo through membrane lipid like a hydrophobic cation. We tested this idea by attaching charged adducts to cysteines introduced into the putative voltage-sensor paddle of Shaker potassium channels and measuring fractional changes in the total gating charge from gating currents. The only residues capable of translocating attached charges through the membrane-electric field are those that serve this function in the native channel. This remarkable specificity indicates that charge movement involves highly specialized interactions between the voltage sensor and other regions of the protein, a mechanism inconsistent with the paddle model.

scholarly journals Voltage Sensitivity and Gating Charge in Shaker and Shab Family Potassium Channels

1999 ◽  
Vol 114 (5) ◽  
pp. 723-742 ◽  
Author(s):  
Leon D. Islas ◽  
Fred J. Sigworth
Keyword(s):  
Single Channel ◽  
Delayed Rectifier ◽  
Voltage Sensitivity ◽  
Charge Movement ◽  
Gating Currents ◽  
Open Probability ◽  
Charge Voltage ◽  
Gating Charge ◽  
Charged Residues ◽  
Positively Charged

The members of the voltage-dependent potassium channel family subserve a variety of functions and are expected to have voltage sensors with different sensitivities. The Shaker channel of Drosophila, which underlies a transient potassium current, has a high voltage sensitivity that is conferred by a large gating charge movement, ∼13 elementary charges. A Shaker subunit's primary voltage-sensing (S4) region has seven positively charged residues. The Shab channel and its homologue Kv2.1 both carry a delayed-rectifier current, and their subunits have only five positively charged residues in S4; they would be expected to have smaller gating-charge movements and voltage sensitivities. We have characterized the gating currents and single-channel behavior of Shab channels and have estimated the charge movement in Shaker, Shab, and their rat homologues Kv1.1 and Kv2.1 by measuring the voltage dependence of open probability at very negative voltages and comparing this with the charge–voltage relationships. We find that Shab has a relatively small gating charge, ∼7.5 eo. Surprisingly, the corresponding mammalian delayed rectifier Kv2.1, which has the same complement of charged residues in the S2, S3, and S4 segments, has a gating charge of 12.5 eo, essentially equal to that of Shaker and Kv1.1. Evidence for very strong coupling between charge movement and channel opening is seen in two channel types, with the probability of voltage-independent channel openings measured to be below 10−9 in Shaker and below 4 × 10−8 in Kv2.1.

scholarly journals The voltage sensor is responsible for ΔpH dependence in Hv1 channels

2021 ◽  
Vol 118 (19) ◽  
pp. e2025556118
Author(s):  
Emerson M. Carmona ◽  
Miguel Fernandez ◽  
Juan J. Alvear-Arias ◽  
Alan Neely ◽  
H. Peter Larsson ◽  
...  
Keyword(s):  
Free Energy ◽  
Ph Dependence ◽  
Electrical Potential ◽  
Voltage Sensor ◽  
Proton Channel ◽  
Gating Currents ◽  
Open Probability ◽  
Charge Voltage ◽  
Gating Charge ◽  
Sensor Activation

The dissipation of acute acid loads by the voltage-gated proton channel (Hv1) relies on regulating the channel’s open probability by the voltage and the ΔpH across the membrane (ΔpH = pHex − pHin). Using monomeric Ciona-Hv1, we asked whether ΔpH-dependent gating is produced during the voltage sensor activation or permeation pathway opening. A leftward shift of the conductance-voltage (G-V) curve was produced at higher ΔpH values in the monomeric channel. Next, we measured the voltage sensor pH dependence in the absence of a functional permeation pathway by recording gating currents in the monomeric nonconducting D160N mutant. Increasing the ΔpH leftward shifted the gating charge-voltage (Q-V) curve, demonstrating that the ΔpH-dependent gating in Hv1 arises by modulating its voltage sensor. We fitted our data to a model that explicitly supposes the Hv1 voltage sensor free energy is a function of both the proton chemical and the electrical potential. The parameters obtained showed that around 60% of the free energy stored in the ΔpH is coupled to the Hv1 voltage sensor activation. Our results suggest that the molecular mechanism underlying the Hv1 ΔpH dependence is produced by protons, which alter the free-energy landscape around the voltage sensor domain. We propose that this alteration is produced by accessibility changes of the protons in the Hv1 voltage sensor during activation.

scholarly journals Charge movement in gating-locked HCN channels reveals weak coupling of voltage sensors and gate

2012 ◽  
Vol 140 (5) ◽  
pp. 469-479 ◽  
Author(s):  
Sujung Ryu ◽  
Gary Yellen
Keyword(s):  
Voltage Dependence ◽  
Voltage Sensor ◽  
Hcn Channels ◽  
Charge Movement ◽  
Coupling Energy ◽  
K Channels ◽  
Voltage Sensors ◽  
Gating Charge ◽  
Pacemaker Channels ◽  
Open States

HCN (hyperpolarization-activated cyclic nucleotide gated) pacemaker channels have an architecture similar to that of voltage-gated K+ channels, but they open with the opposite voltage dependence. HCN channels use essentially the same positively charged voltage sensors and intracellular activation gates as K+ channels, but apparently these two components are coupled differently. In this study, we examine the energetics of coupling between the voltage sensor and the pore by using cysteine mutant channels for which low concentrations of Cd2+ ions freeze the open–closed gating machinery but still allow the sensors to move. We were able to lock mutant channels either into open or into closed states by the application of Cd2+ and measure the effect on voltage sensor movement. Cd2+ did not immobilize the gating charge, as expected for strict coupling, but rather it produced shifts in the voltage dependence of voltage sensor charge movement, consistent with its effect of confining transitions to either closed or open states. From the magnitude of the Cd2+-induced shifts, we estimate that each voltage sensor produces a roughly three- to sevenfold effect on the open–closed equilibrium, corresponding to a coupling energy of ∼1.3–2 kT per sensor. Such coupling is not only opposite in sign to the coupling in K+ channels, but also much weaker.

scholarly journals Electrostatics and the Gating Pore of Shaker Potassium Channels

2001 ◽  
Vol 117 (1) ◽  
pp. 69-90 ◽  
Author(s):  
Leon D. Islas ◽  
Fred J. Sigworth
Keyword(s):  
Charge Movement ◽  
Transmembrane Helices ◽  
Gating Currents ◽  
Conical Cavity ◽  
K Channels ◽  
Gating Charge ◽  
Voltage Dependent ◽  
Shaker Potassium Channels ◽  
Electrostatic Calculations ◽  
S4 Segment

Various experiments have suggested that the S4 segment in voltage-dependent Na+ and K+ channels is in contact with a solvent-accessible cavity. We explore the consequences of the existence of such a cavity through the electrostatic effects on the gating currents of Shaker K+ channels under conditions of reduced ionic strength S. We observe that ∼10-fold reductions of intracellular S produce reductions of the measured gating charge of ∼10%. These effects continue at even lower values of S. The reduction of gating charge when S is reduced by 10-fold at the extracellular surface is much smaller (∼2%). Shifts of the Q(V) curve because of a reduced S are small (<10 mV in size), which is consistent with very little fixed surface charge. Continuum electrostatic calculations show that the S effects on gating charge can be explained by the alteration of the local potential in an intracellular conical cavity of 20–24-Å depth and 12-Å aperture, and a smaller extracellular cavity of 3-Å depth and the same aperture. In this case, the attenuation of the membrane potential at low S leads to reduction of the apparent gating charge. We suggest that this cavity is made by a bundle of transmembrane helices, and that the gating charge movement occurs by translocation of charged residues across a thin septum of ∼3–7 Å thickness.

scholarly journals Molecular Coupling between Voltage Sensor and Pore Opening in the Arabidopsis Inward Rectifier K+ Channel KAT1

2003 ◽  
Vol 122 (4) ◽  
pp. 459-469 ◽  
Author(s):  
Ramon Latorre ◽  
Riccardo Olcese ◽  
Claudia Basso ◽  
Carlos Gonzalez ◽  
Fabian Muñoz ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Structural Similarity ◽  
Ionic Currents ◽  
Inward Rectifier ◽  
Voltage Sensor ◽  
K Channel ◽  
Gating Currents ◽  
Open Probability ◽  
Voltage Sensing ◽  
K Channels

Animal and plant voltage-gated ion channels share a common architecture. They are made up of four subunits and the positive charges on helical S4 segments of the protein in animal K+ channels are the main voltage-sensing elements. The KAT1 channel cloned from Arabidopsis thaliana, despite its structural similarity to animal outward rectifier K+ channels is, however, an inward rectifier. Here we detected KAT1-gating currents due to the existence of an intrinsic voltage sensor in this channel. The measured gating currents evoked in response to hyperpolarizing voltage steps consist of a very fast (τ = 318 ± 34 μs at −180 mV) and a slower component (4.5 ± 0.5 ms at −180 mV) representing charge moved when most channels are closed. The observed gating currents precede in time the ionic currents and they are measurable at voltages (less than or equal to −60) at which the channel open probability is negligible (≈10−4). These two observations, together with the fact that there is a delay in the onset of the ionic currents, indicate that gating charge transits between several closed states before the KAT1 channel opens. To gain insight into the molecular mechanisms that give rise to the gating currents and lead to channel opening, we probed external accessibility of S4 domain residues to methanethiosulfonate-ethyltrimethylammonium (MTSET) in both closed and open cysteine-substituted KAT1 channels. The results demonstrate that the putative voltage–sensing charges of S4 move inward when the KAT1 channels open.

scholarly journals Gating charge displacement in a monomeric voltage-gated proton (Hv1) channel

2018 ◽  
Vol 115 (37) ◽  
pp. 9240-9245 ◽  
Author(s):  
Emerson M. Carmona ◽  
H. Peter Larsson ◽  
Alan Neely ◽  
Osvaldo Alvarez ◽  
Ramon Latorre ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Time Course ◽  
Voltage Sensor ◽  
Total Charge ◽  
Gating Currents ◽  
Voltage Gated ◽  
Central Transition ◽  
Physiological Processes ◽  
Gating Charge ◽  
Voltage Dependent

The voltage-gated proton (Hv1) channel, a voltage sensor and a conductive pore contained in one structural module, plays important roles in many physiological processes. Voltage sensor movements can be directly detected by measuring gating currents, and a detailed characterization of Hv1 charge displacements during channel activation can help to understand the function of this channel. We succeeded in detecting gating currents in the monomeric form of the Ciona-Hv1 channel. To decrease proton currents and better separate gating currents from ion currents, we used the low-conducting Hv1 mutant N264R. Isolated ON-gating currents decayed at increasing rates with increasing membrane depolarization, and the amount of gating charges displaced saturates at high voltages. These are two hallmarks of currents arising from the movement of charged elements within the boundaries of the cell membrane. The kinetic analysis of gating currents revealed a complex time course of the ON-gating current characterized by two peaks and a marked Cole–Moore effect. Both features argue that the voltage sensor undergoes several voltage-dependent conformational changes during activation. However, most of the charge is displaced in a single central transition. Upon voltage sensor activation, the charge is trapped, and only a fast component that carries a small percentage of the total charge is observed in the OFF. We hypothesize that trapping is due to the presence of the arginine side chain in position 264, which acts as a blocking ion. We conclude that the movement of the voltage sensor must proceed through at least five states to account for our experimental data satisfactorily.

scholarly journals A Dipeptidyl Aminopeptidase–like Protein Remodels Gating Charge Dynamics in Kv4.2 Channels

2006 ◽  
Vol 128 (6) ◽  
pp. 745-753 ◽  
Author(s):  
Kevin Dougherty ◽  
Manuel Covarrubias
Keyword(s):  
Membrane Potentials ◽  
Charge Movement ◽  
Transmembrane Segment ◽  
Gating Currents ◽  
Voltage Sensing ◽  
Charge Voltage ◽  
Charge Dynamics ◽  
Gating Charge ◽  
Kv4 Channels ◽  
Dipeptidyl Aminopeptidase

Dipeptidyl aminopeptidase–like proteins (DPLPs) interact with Kv4 channels and thereby induce a profound remodeling of activation and inactivation gating. DPLPs are constitutive components of the neuronal Kv4 channel complex, and recent observations have suggested the critical functional role of the single transmembrane segment of these proteins (Zagha, E., A. Ozaita, S.Y. Chang, M.S. Nadal, U. Lin, M.J. Saganich, T. McCormack, K.O. Akinsanya, S.Y. Qi, and B. Rudy. 2005. J. Biol. Chem. 280:18853–18861). However, the underlying mechanism of action is unknown. We hypothesized that a unique interaction between the Kv4.2 channel and a DPLP found in brain (DPPX-S) may remodel the channel's voltage-sensing domain. To test this hypothesis, we implemented a robust experimental system to measure Kv4.2 gating currents and study gating charge dynamics in the absence and presence of DPPX-S. The results demonstrated that coexpression of Kv4.2 and DPPX-S causes a −26 mV parallel shift in the gating charge-voltage (Q-V) relationship. This shift is associated with faster outward movements of the gating charge over a broad range of relevant membrane potentials and accelerated gating charge return upon repolarization. In sharp contrast, DPPX-S had no effect on gating charge movements of the Shaker B Kv channel. We propose that DPPX-S destabilizes resting and intermediate states in the voltage-dependent activation pathway, which promotes the outward gating charge movement. The remodeling of gating charge dynamics may involve specific protein–protein interactions of the DPPX-S's transmembrane segment with the voltage-sensing and pore domains of the Kv4.2 channel. This mechanism may determine the characteristic fast operation of neuronal Kv4 channels in the subthreshold range of membrane potentials.

scholarly journals Correlation between Charge Movement and Ionic Current during Slow Inactivation in Shaker K+ Channels

1997 ◽  
Vol 110 (5) ◽  
pp. 579-589 ◽  
Author(s):  
Riccardo Olcese ◽  
Ramón Latorre ◽  
Ligia Toro ◽  
Francisco Bezanilla ◽  
Enrico Stefani
Keyword(s):  
Time Course ◽  
Voltage Dependence ◽  
Ionic Current ◽  
K Channel ◽  
Charge Movement ◽  
Slow Inactivation ◽  
Gating Currents ◽  
Similar Time ◽  
K Channels ◽  
Recovery From Inactivation

Prolonged depolarization induces a slow inactivation process in some K+ channels. We have studied ionic and gating currents during long depolarizations in the mutant Shaker H4-Δ(6–46) K+ channel and in the nonconducting mutant (Shaker H4-Δ(6–46)-W434F). These channels lack the amino terminus that confers the fast (N-type) inactivation (Hoshi, T., W.N. Zagotta, and R.W. Aldrich. 1991. Neuron. 7:547–556). Channels were expressed in oocytes and currents were measured with the cut-open-oocyte and patch-clamp techniques. In both clones, the curves describing the voltage dependence of the charge movement were shifted toward more negative potentials when the holding potential was maintained at depolarized potentials. The evidences that this new voltage dependence of the charge movement in the depolarized condition is associated with the process of slow inactivation are the following: (a) the installation of both the slow inactivation of the ionic current and the inactivation of the charge in response to a sustained 1-min depolarization to 0 mV followed the same time course; and (b) the recovery from inactivation of both ionic and gating currents (induced by repolarizations to −90 mV after a 1-min inactivating pulse at 0 mV) also followed a similar time course. Although prolonged depolarizations induce inactivation of the majority of the channels, a small fraction remains non–slow inactivated. The voltage dependence of this fraction of channels remained unaltered, suggesting that their activation pathway was unmodified by prolonged depolarization. The data could be fitted to a sequential model for Shaker K+ channels (Bezanilla, F., E. Perozo, and E. Stefani. 1994. Biophys. J. 66:1011–1021), with the addition of a series of parallel nonconducting (inactivated) states that become populated during prolonged depolarization. The data suggest that prolonged depolarization modifies the conformation of the voltage sensor and that this change can be associated with the process of slow inactivation.

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