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

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).

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

10.1101/2020.04.23.058818 ◽

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).

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

eLife ◽

10.7554/elife.53901 ◽

2020 ◽

Vol 9 ◽

Cited By ~ 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.

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Biaryl sulfonamide motifs up- or down-regulate ion channel activity by activating voltage sensors

The Journal of General Physiology ◽

10.1085/jgp.201711942 ◽

2018 ◽

Vol 150 (8) ◽

pp. 1215-1230 ◽

Cited By ~ 4

Author(s):

Sara I. Liin ◽

Per-Eric Lund ◽

Johan E. Larsson ◽

Johan Brask ◽

Björn Wallner ◽

...

Keyword(s):

Calcium Channels ◽

Action Potential ◽

Ion Channel ◽

Potassium Channel ◽

Voltage Sensor ◽

Pharmaceutical Drugs ◽

Voltage Gated ◽

Channel Opening ◽

Ion Conducting ◽

Voltage Gated Ion Channels

Voltage-gated ion channels are key molecules for the generation of cellular electrical excitability. Many pharmaceutical drugs target these channels by blocking their ion-conducting pore, but in many cases, channel-opening compounds would be more beneficial. Here, to search for new channel-opening compounds, we screen 18,000 compounds with high-throughput patch-clamp technology and find several potassium-channel openers that share a distinct biaryl-sulfonamide motif. Our data suggest that the negatively charged variants of these compounds bind to the top of the voltage-sensor domain, between transmembrane segments 3 and 4, to open the channel. Although we show here that biaryl-sulfonamide compounds open a potassium channel, they have also been reported to block sodium and calcium channels. However, because they inactivate voltage-gated sodium channels by promoting activation of one voltage sensor, we suggest that, despite different effects on the channel gates, the biaryl-sulfonamide motif is a general ion-channel activator motif. Because these compounds block action potential–generating sodium and calcium channels and open an action potential–dampening potassium channel, they should have a high propensity to reduce excitability. This opens up the possibility to build new excitability-reducing pharmaceutical drugs from the biaryl-sulfonamide scaffold.

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Structural model of a voltage-gated potassium channel based on spectroscopic data

Biochemical Society Transactions ◽

10.1042/bst0290589 ◽

2001 ◽

Vol 29 (4) ◽

pp. 589-593 ◽

Cited By ~ 2

Author(s):

P. I. Haris

Keyword(s):

Potassium Channel ◽

Structural Model ◽

Structural Data ◽

Theoretical Models ◽

Shaker Potassium Channel ◽

Voltage Gated ◽

Folded Structure ◽

Membrane Spanning ◽

Voltage Gated Ion Channels ◽

Pore Domain

It is estimated that membrane proteins comprise as much as 30% of most genomes. Yet our knowledge of membrane-protein folding is still in its infancy. Consequently, there is a great need for developing approaches that can further advance our understanding of how peptides and proteins interact with membranes and thereby attain their folded structure. An approach that we have been exploring involves dissecting voltage-gated ion channels into simple peptide domains for the purpose of determining their structure in different media using physical techniques. We have synthesized peptides corresponding to the six membrane-spanning segments, as well as the pore domain, of the Shaker channel and characterized their secondary structures. From these studies we have developed a model for the transmembrane structure of the Shaker potassium channel that is constructed from α-helices. The hard structural data obtained from these studies lends support to the recent theoretical models of this channel protein that have been developed by others.

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Gating-induced large aqueous volumetric remodeling and aspartate tolerance in the voltage sensor domain of Shaker K+ channels

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1806578115 ◽

2018 ◽

Vol 115 (32) ◽

pp. 8203-8208 ◽

Cited By ~ 6

Author(s):

Ignacio Díaz-Franulic ◽

Vivian González-Pérez ◽

Hans Moldenhauer ◽

Nieves Navarro-Quezada ◽

David Naranjo

Keyword(s):

Voltage Sensor ◽

Side Chains ◽

Aqueous Interfaces ◽

Voltage Gated ◽

Low Dielectric ◽

Positive Side ◽

Voltage Gated Ion Channels ◽

Positively Charged ◽

As If

Neurons encode electrical signals with critically tuned voltage-gated ion channels and enzymes. Dedicated voltage sensor domains (VSDs) in these membrane proteins activate coordinately with an unresolved structural change. Such change conveys the transmembrane translocation of four positively charged arginine side chains, the voltage-sensing residues (VSRs; R1–R4). Countercharges and lipid phosphohead groups likely stabilize these VSRs within the low-dielectric core of the protein. However, the role of hydration, a sign-independent charge stabilizer, remains unclear. We replaced all VSRs and their neighboring residues with negatively charged aspartates in a voltage-gated potassium channel. The ensuing mild functional effects indicate that hydration is also important in VSR stabilization. The voltage dependency of the VSR aspartate variants approached the expected arithmetic summation of charges at VSR positions, as if negative and positive side chains faced similar pathways. In contrast, aspartates introduced between R2 and R3 did not affect voltage dependence as if the side chains moved outside the electric field or together with it, undergoing a large displacement and volumetric remodeling. Accordingly, VSR performed osmotic work at both internal and external aqueous interfaces. Individual VSR contributions to volumetric works approached arithmetical additivity but were largely dissimilar. While R1 and R4 displaced small volumes, R2 and R3 volumetric works were massive and vectorially opposed, favoring large aqueous remodeling during VSD activation. These diverse volumetric works are, at least for R2 and R3, not compatible with VSR translocation across a unique stationary charge transfer center. Instead, VSRs may follow separated pathways across a fluctuating low-dielectric septum.

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