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Voltage sensitivity and gating charge in Shaker and Shab family potassium channels.

Islas LD, Sigworth FJ - J. Gen. Physiol. (1999)

Bottom Line: We find that Shab has a relatively small gating charge, approximately 7.5 e(o).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 e(o), 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 x 10(-8) in Kv2.1.

View Article: PubMed Central - PubMed

Affiliation: Department of Cellular and Molecular Physiology, Yale University School of Medicine, New Haven, Connecticut 06520, USA.

ABSTRACT
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, approximately 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, approximately 7.5 e(o). 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 e(o), 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 x 10(-8) in Kv2.1.

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Shab channels appear to have only one fully open state. (A) Single-channel traces recorded from a cell-attached patch at the indicated voltages. The patch contained only one channel as judged from the absence of overlapping openings. S indicates subconductance events; F indicates full conductance events. Bath solution was the standard composition. The pipette contained (mM): 5 K-aspartate, 5 KCl, 1.8 CaCl2, 100 NMDG-aspartate, 10 HEPES, pH 7.4. (B) Dwell-time distributions in the fully open (left) or closed (right) states. Superimposed on the histograms are maximum-likelihood fits to an exponential function for the open-time histogram, or a mixture of three exponentials for the closed time histogram. (C) Voltage dependence of the time constants. Filled symbols are the open-time constant and open symbols represent the three detected closed-time constants. The fitted lines represent exponential functions of voltage with the effective charges shown. (D) Prepulse inactivation in Shab channels. A prepulse of 500-ms duration was given at the indicated membrane potential and the current at a subsequent fixed depolarization of 40 mV was measured. The continuous curve is a fit to the same function as in Fig. 7 from −90 to −20 mV. The fit parameters are: A = 0.39, q = 8.1 eo, and Vo = −47 mV.
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Figure 10: Shab channels appear to have only one fully open state. (A) Single-channel traces recorded from a cell-attached patch at the indicated voltages. The patch contained only one channel as judged from the absence of overlapping openings. S indicates subconductance events; F indicates full conductance events. Bath solution was the standard composition. The pipette contained (mM): 5 K-aspartate, 5 KCl, 1.8 CaCl2, 100 NMDG-aspartate, 10 HEPES, pH 7.4. (B) Dwell-time distributions in the fully open (left) or closed (right) states. Superimposed on the histograms are maximum-likelihood fits to an exponential function for the open-time histogram, or a mixture of three exponentials for the closed time histogram. (C) Voltage dependence of the time constants. Filled symbols are the open-time constant and open symbols represent the three detected closed-time constants. The fitted lines represent exponential functions of voltage with the effective charges shown. (D) Prepulse inactivation in Shab channels. A prepulse of 500-ms duration was given at the indicated membrane potential and the current at a subsequent fixed depolarization of 40 mV was measured. The continuous curve is a fit to the same function as in Fig. 7 from −90 to −20 mV. The fit parameters are: A = 0.39, q = 8.1 eo, and Vo = −47 mV.

Mentions: As a further check for the possible existence of multiple open states, we examined the single-channel kinetics of Shab channels. Fig. 10 A shows recordings from a patch containing a single Shab channel. Over the range of voltages from −30 to +40 mV, the open time distribution showed a single exponential component (Fig. 10 B). Data from multichannel patches also show no evidence for a second open-time component at voltages down to −80 mV. The mean burst duration at −60 mV is 3.7 ± 0.1 ms (n = 3) and has a very weak voltage dependence corresponding to a partial charge of 0.1 eo. The single-exponential open-time distributions support the idea that Shab has only one fully open state. Thus again, we find no reason to expect multiple open states, and find the simplest conclusion to be that Shab channels have a reduced gating charge of ∼7.5 e0.


Voltage sensitivity and gating charge in Shaker and Shab family potassium channels.

Islas LD, Sigworth FJ - J. Gen. Physiol. (1999)

Shab channels appear to have only one fully open state. (A) Single-channel traces recorded from a cell-attached patch at the indicated voltages. The patch contained only one channel as judged from the absence of overlapping openings. S indicates subconductance events; F indicates full conductance events. Bath solution was the standard composition. The pipette contained (mM): 5 K-aspartate, 5 KCl, 1.8 CaCl2, 100 NMDG-aspartate, 10 HEPES, pH 7.4. (B) Dwell-time distributions in the fully open (left) or closed (right) states. Superimposed on the histograms are maximum-likelihood fits to an exponential function for the open-time histogram, or a mixture of three exponentials for the closed time histogram. (C) Voltage dependence of the time constants. Filled symbols are the open-time constant and open symbols represent the three detected closed-time constants. The fitted lines represent exponential functions of voltage with the effective charges shown. (D) Prepulse inactivation in Shab channels. A prepulse of 500-ms duration was given at the indicated membrane potential and the current at a subsequent fixed depolarization of 40 mV was measured. The continuous curve is a fit to the same function as in Fig. 7 from −90 to −20 mV. The fit parameters are: A = 0.39, q = 8.1 eo, and Vo = −47 mV.
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Figure 10: Shab channels appear to have only one fully open state. (A) Single-channel traces recorded from a cell-attached patch at the indicated voltages. The patch contained only one channel as judged from the absence of overlapping openings. S indicates subconductance events; F indicates full conductance events. Bath solution was the standard composition. The pipette contained (mM): 5 K-aspartate, 5 KCl, 1.8 CaCl2, 100 NMDG-aspartate, 10 HEPES, pH 7.4. (B) Dwell-time distributions in the fully open (left) or closed (right) states. Superimposed on the histograms are maximum-likelihood fits to an exponential function for the open-time histogram, or a mixture of three exponentials for the closed time histogram. (C) Voltage dependence of the time constants. Filled symbols are the open-time constant and open symbols represent the three detected closed-time constants. The fitted lines represent exponential functions of voltage with the effective charges shown. (D) Prepulse inactivation in Shab channels. A prepulse of 500-ms duration was given at the indicated membrane potential and the current at a subsequent fixed depolarization of 40 mV was measured. The continuous curve is a fit to the same function as in Fig. 7 from −90 to −20 mV. The fit parameters are: A = 0.39, q = 8.1 eo, and Vo = −47 mV.
Mentions: As a further check for the possible existence of multiple open states, we examined the single-channel kinetics of Shab channels. Fig. 10 A shows recordings from a patch containing a single Shab channel. Over the range of voltages from −30 to +40 mV, the open time distribution showed a single exponential component (Fig. 10 B). Data from multichannel patches also show no evidence for a second open-time component at voltages down to −80 mV. The mean burst duration at −60 mV is 3.7 ± 0.1 ms (n = 3) and has a very weak voltage dependence corresponding to a partial charge of 0.1 eo. The single-exponential open-time distributions support the idea that Shab has only one fully open state. Thus again, we find no reason to expect multiple open states, and find the simplest conclusion to be that Shab channels have a reduced gating charge of ∼7.5 e0.

Bottom Line: We find that Shab has a relatively small gating charge, approximately 7.5 e(o).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 e(o), 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 x 10(-8) in Kv2.1.

View Article: PubMed Central - PubMed

Affiliation: Department of Cellular and Molecular Physiology, Yale University School of Medicine, New Haven, Connecticut 06520, USA.

ABSTRACT
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, approximately 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, approximately 7.5 e(o). 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 e(o), 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 x 10(-8) in Kv2.1.

Show MeSH
Related in: MedlinePlus