S4 Charges Move Close to Residues in the Pore Domain during Activation in a K Channel

2001 ◽  
Vol 118 (1) ◽  
pp. 1-10 ◽  
Author(s):  
Fredrik Elinder ◽  
Roope Männikkö ◽  
H. Peter Larsson
Keyword(s):  
Close Approach ◽  
Voltage Sensor ◽  
K Channel ◽  
Slow Inactivation ◽  
Transmembrane Voltage ◽  
Activated State ◽  
Ion Conducting ◽  
Voltage Gated Ion Channels ◽  
Modification Rate ◽  
Positively Charged

Voltage-gated ion channels respond to changes in the transmembrane voltage by opening or closing their ion conducting pore. The positively charged fourth transmembrane segment (S4) has been identified as the main voltage sensor, but the mechanisms of coupling between the voltage sensor and the gates are still unknown. Obtaining information about the location and the exact motion of S4 is an important step toward an understanding of these coupling mechanisms. In previous studies we have shown that the extracellular end of S4 is located close to segment 5 (S5). The purpose of the present study is to estimate the location of S4 charges in both resting and activated states. We measured the modification rates by differently charged methanethiosulfonate regents of two residues in the extracellular end of S5 in the Shaker K channel (418C and 419C). When S4 moves to its activated state, the modification rate by the negatively charged sodium (2-sulfonatoethyl) methanethiosulfonate (MTSES−) increases significantly more than the modification rate by the positively charged [2-(trimethylammonium)ethyl] methanethiosulfonate, bromide (MTSET+). This indicates that the positive S4 charges are moving close to 418C and 419C in S5 during activation. Neutralization of the most external charge of S4 (R362), shows that R362 in its activated state electrostatically affects the environment at 418C by 19 mV. In contrast, R362 in its resting state has no effect on 418C. This suggests that, during activation of the channel, R362 moves from a position far away (>20 Å) to a position close (8 Å) to 418C. Despite its close approach to E418, a residue shown to be important in slow inactivation, R362 has no effect on slow inactivation or the recovery from slow inactivation. This refutes previous models for slow inactivation with an electrostatic S4-to-gate coupling. Instead, we propose a model with an allosteric mechanism for the S4-to-gate coupling.

scholarly journals The trajectory of discrete gating charges in a voltage-gated potassium channel

2020 ◽  
Author(s):  
Michael F. Priest ◽  
Elizabeth E.L. Lee ◽  
Francisco Bezanilla
Keyword(s):  
Potassium Channel ◽  
Voltage Sensor ◽  
Drive Voltage ◽  
Shaker Potassium Channel ◽  
Voltage Gated ◽  
Charged Amino Acids ◽  
Voltage Gated Ion Channels ◽  
Sensing Membrane ◽  
Positively Charged ◽  
Gating Charges

AbstractPositively-charged amino acids respond to membrane potential changes to drive voltage sensor movement in voltage-gated ion channels, but determining the trajectory of voltage sensor gating charges has proven difficult. We optically tracked the movement of the two most extracellular charged residues (R1, R2) in the Shaker potassium channel voltage sensor using a fluorescent positively-charged bimane derivative (qBBr) that is strongly quenched by tryptophan. By individually mutating residues to tryptophan within the putative trajectory of gating charges, we observed that the charge pathway during activation is a rotation and a tilted translation that differs between R1 and R2 and is distinct from their deactivation pathway. Tryptophan-induced quenching of qBBr also indicates that a crucial residue of the hydrophobic plug is linked to the Cole-Moore shift through its interaction with R1. Finally, we show that this approach extends to additional voltage-sensing membrane proteins using the Ciona intestinalis voltage sensitive phosphatase (CiVSP).

scholarly journals Tracking the movement of discrete gating charges in a voltage-gated potassium channel

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Michael F Priest ◽  
Elizabeth EL Lee ◽  
Francisco Bezanilla
Keyword(s):  
Potassium Channel ◽  
Voltage Sensor ◽  
Charge Motion ◽  
Shaker Potassium Channel ◽  
Voltage Gated ◽  
Charged Amino Acids ◽  
Voltage Gated Ion Channels ◽  
Sensing Membrane ◽  
Positively Charged ◽  
Gating Charges

Positively-charged amino acids respond to membrane potential changes to drive voltage sensor movement in voltage-gated ion channels, but determining the displacements of voltage sensor gating charges has proven difficult. We optically tracked the movement of the two most extracellular charged residues (R1, R2) in the Shaker potassium channel voltage sensor using a fluorescent positively-charged bimane derivative (qBBr) that is strongly quenched by tryptophan. By individually mutating residues to tryptophan within the putative pathway of gating charges, we observed that the charge motion during activation is a rotation and a tilted translation that differs between R1 and R2. Tryptophan-induced quenching of qBBr also indicates that a crucial residue of the hydrophobic plug is linked to the Cole-Moore shift through its interaction with R1. Finally, we show that this approach extends to additional voltage-sensing membrane proteins using the Ciona intestinalis voltage sensitive phosphatase (CiVSP) (Murata et al., 2005a).

scholarly journals Structure and physiological function of the human KCNQ1 channel voltage sensor intermediate state

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Keenan C Taylor ◽  
Po Wei Kang ◽  
Panpan Hou ◽  
Nien-Du Yang ◽  
Georg Kuenze ◽  
...  
Keyword(s):  
Potassium Channel ◽  
Intermediate State ◽  
Three Dimensional ◽  
Voltage Sensor ◽  
Dimensional Structure ◽  
Delayed Rectifier ◽  
Voltage Gated ◽  
Delayed Rectifier Current ◽  
Activated State ◽  
Voltage Gated Ion Channels

Voltage-gated ion channels feature voltage sensor domains (VSDs) that exist in three distinct conformations during activation: resting, intermediate, and activated. Experimental determination of the structure of a potassium channel VSD in the intermediate state has previously proven elusive. Here, we report and validate the experimental three-dimensional structure of the human KCNQ1 voltage-gated potassium channel VSD in the intermediate state. We also used mutagenesis and electrophysiology in Xenopus laevisoocytes to functionally map the determinants of S4 helix motion during voltage-dependent transition from the intermediate to the activated state. Finally, the physiological relevance of the intermediate state KCNQ1 conductance is demonstrated using voltage-clamp fluorometry. This work illuminates the structure of the VSD intermediate state and demonstrates that intermediate state conductivity contributes to the unusual versatility of KCNQ1, which can function either as the slow delayed rectifier current (IKs) of the cardiac action potential or as a constitutively active epithelial leak current.

scholarly journals Transfer of Voltage Independence from a Rat Olfactory Channel to the Drosophila Ether-à-go-go K+ Channel

1997 ◽  
Vol 109 (3) ◽  
pp. 301-311 ◽  
Author(s):  
Chih-Yung Tang ◽  
Diane M. Papazian
Keyword(s):  
Cyclic Nucleotide ◽  
Voltage Sensor ◽  
K Channel ◽  
Voltage Range ◽  
Voltage Gated ◽  
Cyclic Nucleotide Gated Channels ◽  
Voltage Dependent ◽  
Relative Stabilization ◽  
Voltage Gated Ion Channels ◽  
S4 Segment

The S4 segment is an important part of the voltage sensor in voltage-gated ion channels. Cyclic nucleotide-gated channels, which are members of the superfamily of voltage-gated channels, have little inherent sensitivity to voltage despite the presence of an S4 segment. We made chimeras between a voltage-independent rat olfactory channel (rolf) and the voltage-dependent ether-à-go-go K+ channel (eag) to determine the basis of their divergent gating properties. We found that the rolf S4 segment can support a voltage-dependent mechanism of activation in eag, suggesting that rolf has a potentially functional voltage sensor that is silent during gating. In addition, we found that the S3-S4 loop of rolf increases the relative stability of the open conformation of eag, effectively converting eag into a voltage-independent channel. A single charged residue in the loop makes a significant contribution to the relative stabilization of the open state in eag. Our data suggest that cyclic nucleotide-gated channels such as rolf contain a voltage sensor which, in the physiological voltage range, is stabilized in an activated conformation that is permissive for pore opening.

scholarly journals An electrostatic potassium channel opener targeting the final voltage sensor transition

2011 ◽  
Vol 137 (6) ◽  
pp. 563-577 ◽  
Author(s):  
Sara I. Börjesson ◽  
Fredrik Elinder
Keyword(s):  
Ion Channels ◽  
Epileptic Seizures ◽  
Voltage Sensor ◽  
K Channel ◽  
Voltage Gated ◽  
Pharmacological Mechanism ◽  
Channel Opening ◽  
Channel Surface ◽  
Future Drug ◽  
Voltage Gated Ion Channels

Free polyunsaturated fatty acids (PUFAs) modulate the voltage dependence of voltage-gated ion channels. As an important consequence thereof, PUFAs can suppress epileptic seizures and cardiac arrhythmia. However, molecular details for the interaction between PUFA and ion channels are not well understood. In this study, we have localized the site of action for PUFAs on the voltage-gated Shaker K channel by introducing positive charges on the channel surface, which potentiated the PUFA effect. Furthermore, we found that PUFA mainly affects the final voltage sensor movement, which is closely linked to channel opening, and that specific charges at the extracellular end of the voltage sensor are critical for the PUFA effect. Because different voltage-gated K channels have different charge profiles, this implies channel-specific PUFA effects. The identified site and the pharmacological mechanism will potentially be very useful in future drug design of small-molecule compounds specifically targeting neuronal and cardiac excitability.

scholarly journals Protein Rearrangements Underlying Slow Inactivation of the Shaker K+ Channel

1998 ◽  
Vol 112 (4) ◽  
pp. 377-389 ◽  
Author(s):  
E. Loots ◽  
E.Y. Isacoff
Keyword(s):  
Voltage Sensor ◽  
K Channel ◽  
Slow Inactivation ◽  
Protein Motions ◽  
Voltage Dependent ◽  
Conformational Rearrangements ◽  
P Type ◽  
Gating Process ◽  
Insight Into ◽  
Local Protein

Voltage-dependent ion channels transduce changes in the membrane electric field into protein rearrangements that gate their transmembrane ion permeation pathways. While certain molecular elements of the voltage sensor and gates have been identified, little is known about either the nature of their conformational rearrangements or about how the voltage sensor is coupled to the gates. We used voltage clamp fluorometry to examine the voltage sensor (S4) and pore region (P-region) protein motions that underlie the slow inactivation of the Shaker K+ channel. Fluorescent probes in both the P-region and S4 changed emission intensity in parallel with the onset and recovery of slow inactivation, indicative of local protein rearrangements in this gating process. Two sequential rearrangements were observed, with channels first entering the P-type, and then the C-type inactivated state. These forms of inactivation appear to be mediated by a single gate, with P-type inactivation closing the gate and C-type inactivation stabilizing the gate's closed conformation. Such a stabilization was due, at least in part, to a slow rearrangement around S4 that stabilizes S4 in its activated transmembrane position. The fluorescence reports of S4 and P-region fluorophore are consistent with an increased interaction of the voltage sensor and inactivation gate upon gate closure, offering insight into how the voltage-sensing apparatus is coupled to a channel gate.

scholarly journals Extracellular protons accelerate hERG channel deactivation by destabilizing voltage sensor relaxation

2018 ◽  
Vol 151 (2) ◽  
pp. 231-246 ◽  
Author(s):  
Yu Patrick Shi ◽  
Samrat Thouta ◽  
Yen May Cheng ◽  
Tom W. Claydon
Keyword(s):  
Ph Dependence ◽  
Voltage Sensor ◽  
Herg Channel ◽  
K Channel ◽  
Delayed Rectifier ◽  
Mode Shift ◽  
Deactivation Kinetics ◽  
Shift Behavior ◽  
Activated State ◽  
Herg Channels

hERG channels underlie the delayed-rectifier K+ channel current (IKr), which is crucial for membrane repolarization and therefore termination of the cardiac action potential. hERG channels display unusually slow deactivation gating, which contributes to a resurgent current upon repolarization and may protect against post-depolarization–induced arrhythmias. hERG channels also exhibit robust mode shift behavior, which reflects the energetic separation of activation and deactivation pathways due to voltage sensor relaxation into a stable activated state. The mechanism of relaxation is unknown and likely contributes to slow hERG channel deactivation. Here, we use extracellular acidification to probe the structural determinants of voltage sensor relaxation and its influence on the deactivation gating pathway. Using gating current recordings and voltage clamp fluorimetry measurements of voltage sensor domain dynamics, we show that voltage sensor relaxation is destabilized at pH 6.5, causing an ∼20-mV shift in the voltage dependence of deactivation. We show that the pH dependence of the resultant loss of mode shift behavior is similar to that of the deactivation kinetics acceleration, suggesting that voltage sensor relaxation correlates with slower pore gate closure. Neutralization of D509 in S3 also destabilizes the relaxed state of the voltage sensor, mimicking the effect of protons, suggesting that acidic residues on S3, which act as countercharges to S4 basic residues, are involved in stabilizing the relaxed state and slowing deactivation kinetics. Our findings identify the mechanistic determinants of voltage sensor relaxation and define the long-sought mechanism by which protons accelerate hERG deactivation.

scholarly journals Trapping of a Methanesulfonanilide by Closure of the Herg Potassium Channel Activation Gate

2000 ◽  
Vol 115 (3) ◽  
pp. 229-240 ◽  
Author(s):  
John S. Mitcheson ◽  
Jun Chen ◽  
Michael C. Sanguinetti
Keyword(s):  
Single Channel ◽  
K Channel ◽  
Membrane Hyperpolarization ◽  
K Channels ◽  
Large Size ◽  
Channel Opening ◽  
Activated State ◽  
Activation Gate ◽  
Herg Channels ◽  
Positively Charged

Deactivation of voltage-gated potassium (K+) channels can slow or prevent the recovery from block by charged organic compounds, a phenomenon attributed to trapping of the compound within the inner vestibule by closure of the activation gate. Unbinding and exit from the channel vestibule of a positively charged organic compound should be favored by membrane hyperpolarization if not impeded by the closed gate. MK-499, a methanesulfonanilide compound, is a potent blocker (IC50 = 32 nM) of HERG K+ channels. This bulky compound (7 × 20 Å) is positively charged at physiological pH. Recovery from block of HERG channels by MK-499 and other methanesulfonanilides is extremely slow (Carmeliet 1992; Ficker et al. 1998), suggesting a trapping mechanism. We used a mutant HERG (D540K) channel expressed in Xenopus oocytes to test the trapping hypothesis. D540K HERG has the unusual property of opening in response to hyperpolarization, in addition to relatively normal gating and channel opening in response to depolarization (Sanguinetti and Xu 1999). The hyperpolarization-activated state of HERG was characterized by long bursts of single channel reopening. Channel reopening allowed recovery from block by 2 μM MK-499 to occur with time constants of 10.5 and 52.7 s at −160 mV. In contrast, wild-type HERG channels opened only briefly after membrane hyperpolarization, and thus did not permit recovery from block by MK-499. These findings provide direct evidence that the mechanism of slow recovery from HERG channel block by methanesulfonanilides is due to trapping of the compound in the inner vestibule by closure of the activation gate. The ability of HERG channels to trap MK-499, despite its large size, suggests that the vestibule of this channel is larger than the well studied Shaker K+ channel.

scholarly journals Drug-induced ion channel opening tuned by the voltage sensor charge profile

2014 ◽  
Vol 143 (2) ◽  
pp. 173-182 ◽  
Author(s):  
Nina E. Ottosson ◽  
Sara I. Liin ◽  
Fredrik Elinder
Keyword(s):  
Small Molecule ◽  
Voltage Sensor ◽  
K Channel ◽  
Detailed Knowledge ◽  
Drug Induced ◽  
New Class ◽  
Channel Opening ◽  
Future Drug ◽  
Small Molecule Compound ◽  
Voltage Gated Ion Channels

Polyunsaturated fatty acids modulate the voltage dependence of several voltage-gated ion channels, thereby being potent modifiers of cellular excitability. Detailed knowledge of this molecular mechanism can be used in designing a new class of small-molecule compounds against hyperexcitability diseases. Here, we show that arginines on one side of the helical K-channel voltage sensor S4 increased the sensitivity to docosahexaenoic acid (DHA), whereas arginines on the opposing side decreased this sensitivity. Glutamates had opposite effects. In addition, a positively charged DHA-like molecule, arachidonyl amine, had opposite effects to the negatively charged DHA. This suggests that S4 rotates to open the channel and that DHA electrostatically affects this rotation. A channel with arginines in positions 356, 359, and 362 was extremely sensitive to DHA: 70 µM DHA at pH 9.0 increased the current >500 times at negative voltages compared with wild type (WT). The small-molecule compound pimaric acid, a novel Shaker channel opener, opened the WT channel. The 356R/359R/362R channel drastically increased this effect, suggesting it to be instrumental in future drug screening.

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