scholarly journals Intermediate state trapping of a voltage sensor

2012 ◽  
Vol 140 (6) ◽  
pp. 635-652 ◽  
Author(s):  
Jérôme J. Lacroix ◽  
Stephan A. Pless ◽  
Luca Maragliano ◽  
Fabiana V. Campos ◽  
Jason D. Galpin ◽  
...  
Keyword(s):  
Ion Channels ◽  
Conformational Changes ◽  
Intermediate State ◽  
Molecular Mechanisms ◽  
Voltage Sensor ◽  
Functional Coupling ◽  
Kv Channel ◽  
New Information ◽  
Helical Turn ◽  
Voltage Gated Ion Channels

Voltage sensor domains (VSDs) regulate ion channels and enzymes by undergoing conformational changes depending on membrane electrical signals. The molecular mechanisms underlying the VSD transitions are not fully understood. Here, we show that some mutations of I241 in the S1 segment of the Shaker Kv channel positively shift the voltage dependence of the VSD movement and alter the functional coupling between VSD and pore domains. Among the I241 mutants, I241W immobilized the VSD movement during activation and deactivation, approximately halfway between the resting and active states, and drastically shifted the voltage activation of the ionic conductance. This phenotype, which is consistent with a stabilization of an intermediate VSD conformation by the I241W mutation, was diminished by the charge-conserving R2K mutation but not by the charge-neutralizing R2Q mutation. Interestingly, most of these effects were reproduced by the F244W mutation located one helical turn above I241. Electrophysiology recordings using nonnatural indole derivatives ruled out the involvement of cation-Π interactions for the effects of the Trp inserted at positions I241 and F244 on the channel’s conductance, but showed that the indole nitrogen was important for the I241W phenotype. Insight into the molecular mechanisms responsible for the stabilization of the intermediate state were investigated by creating in silico the mutations I241W, I241W/R2K, and F244W in intermediate conformations obtained from a computational VSD transition pathway determined using the string method. The experimental results and computational analysis suggest that the phenotype of I241W may originate in the formation of a hydrogen bond between the indole nitrogen atom and the backbone carbonyl of R2. This work provides new information on intermediate states in voltage-gated ion channels with an approach that produces minimum chemical perturbation.

scholarly journals Cryo-EM structure of the KvAP channel reveals a non-domain-swapped voltage sensor topology

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Xiao Tao ◽  
Roderick MacKinnon
Keyword(s):  
Ion Channels ◽  
Conformational Changes ◽  
Voltage Sensor ◽  
Structural Domains ◽  
Structural Class ◽  
Voltage Sensors ◽  
Voltage Gated ◽  
Voltage Dependent ◽  
Voltage Gated Ion Channels ◽  
Gated Ion Channels

Conductance in voltage-gated ion channels is regulated by membrane voltage through structural domains known as voltage sensors. A single structural class of voltage sensor domain exists, but two different modes of voltage sensor attachment to the pore occur in nature: domain-swapped and non-domain-swapped. Since the more thoroughly studied Kv1-7, Nav and Cav channels have domain-swapped voltage sensors, much less is known about non-domain-swapped voltage-gated ion channels. In this paper, using cryo-EM, we show that KvAP from Aeropyrum pernix has non-domain-swapped voltage sensors as well as other unusual features. The new structure, together with previous functional data, suggests that KvAP and the Shaker channel, to which KvAP is most often compared, probably undergo rather different voltage-dependent conformational changes when they open.

The Croonian Lecture, 1983 - Voltage-gated ion channels in the nerve membrane

1983 ◽  
Vol 220 (1218) ◽  
pp. 1-30 ◽  
Keyword(s):  
Ion Channels ◽  
Conformational Changes ◽  
Molecular Mechanisms ◽  
Essential Feature ◽  
Ionic Currents ◽  
Fluctuation Analysis ◽  
Gating Current ◽  
Nervous Impulse ◽  
Voltage Gated Ion Channels ◽  
Kinetics Of

In the Croonian Lecture for 1957, Sir Alan Hodgkin described the role of the channels selective for sodium and potassium ions in the conduction of the nervous impulse. An essential feature of these channels is the manner in which the complex kinetics of their opening and closing is controlled by the electric field across the membrane, and the purpose of the present lecture is to consider the advances that have been made in the past 25 years towards an understanding of the underlying molecular mechanisms. One such advance has been the successful recording, independently of the ionic currents, of the small asymmetry current known as the gating current, that accompanies the conformational changes that take place in the sodium channels. A quantitative analysis of the characteristics of the gating current suggests that activation is brought about by two more or less independent processes operating in parallel, to one of which the slower mechanism of inactivation is coupled sequentially. However, it is clear that a complete picture of the gating system will only be arrived at by combining evidence of this kind with that provided by other new lines of approach such as studies of single ion channels in various tissues by means of fluctuation analysis and the patch-clamping technique, and a reinvestigation of the kinetics of activation of the potassium channels.

scholarly journals Helix breaking transition in the S4 of HCN channel is critical for hyperpolarization-dependent gating

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Marina A Kasimova ◽  
Debanjan Tewari ◽  
John B Cowgill ◽  
Willy Carrasquel Ursuleaz ◽  
Jenna L Lin ◽  
...  
Keyword(s):  
Ion Channels ◽  
Voltage Sensor ◽  
Inward Rectification ◽  
Hcn Channels ◽  
Hcn Channel ◽  
Functional Studies ◽  
Gating Charge ◽  
Helical Turn ◽  
Voltage Gated Ion Channels ◽  
Dynamics Simulations

In contrast to most voltage-gated ion channels, hyperpolarization- and cAMP gated (HCN) ion channels open on hyperpolarization. Structure-function studies show that the voltage-sensor of HCN channels are unique but the mechanisms that determine gating polarity remain poorly understood. All-atom molecular dynamics simulations (~20 μs) of HCN1 channel under hyperpolarization reveals an initial downward movement of the S4 voltage-sensor but following the transfer of last gating charge, the S4 breaks into two sub-helices with the lower sub-helix becoming parallel to the membrane. Functional studies on bipolar channels show that the gating polarity strongly correlates with helical turn propensity of the substituents at the breakpoint. Remarkably, in a proto-HCN background, the replacement of breakpoint serine with a bulky hydrophobic amino acid is sufficient to completely flip the gating polarity from inward to outward-rectifying. Our studies reveal an unexpected mechanism of inward rectification involving a linker sub-helix emerging from HCN S4 during hyperpolarization.

scholarly journals Interfacial gating triad is crucial for electromechanical transduction in voltage-activated potassium channels

2014 ◽  
Vol 144 (5) ◽  
pp. 457-467 ◽  
Author(s):  
Sandipan Chowdhury ◽  
Benjamin M. Haehnel ◽  
Baron Chanda
Keyword(s):  
Potassium Channels ◽  
Conformational Changes ◽  
Electromechanical Coupling ◽  
Structural Integrity ◽  
Voltage Sensor ◽  
Amino Acid Residues ◽  
Kv Channel ◽  
Electromechanical Transduction ◽  
Voltage Dependent ◽  
Graph Theoretical Approach

Voltage-dependent potassium channels play a crucial role in electrical excitability and cellular signaling by regulating potassium ion flux across membranes. Movement of charged residues in the voltage-sensing domain leads to a series of conformational changes that culminate in channel opening in response to changes in membrane potential. However, the molecular machinery that relays these conformational changes from voltage sensor to the pore is not well understood. Here we use generalized interaction-energy analysis (GIA) to estimate the strength of site-specific interactions between amino acid residues putatively involved in the electromechanical coupling of the voltage sensor and pore in the outwardly rectifying KV channel. We identified candidate interactors at the interface between the S4–S5 linker and the pore domain using a structure-guided graph theoretical approach that revealed clusters of conserved and closely packed residues. One such cluster, located at the intracellular intersubunit interface, comprises three residues (arginine 394, glutamate 395, and tyrosine 485) that interact with each other. The calculated interaction energies were 3–5 kcal, which is especially notable given that the net free-energy change during activation of the Shaker KV channel is ∼14 kcal. We find that this triad is delicately maintained by balance of interactions that are responsible for structural integrity of the intersubunit interface while maintaining sufficient flexibility at a critical gating hinge for optimal transmission of force to the pore gate.

scholarly journals A new mechanism of voltage-dependent gating exposed by KV10.1 channels interrupted between voltage sensor and pore

2017 ◽  
Vol 149 (5) ◽  
pp. 577-593 ◽  
Author(s):  
Adam P. Tomczak ◽  
Jorge Fernández-Trillo ◽  
Shashank Bharill ◽  
Ferenc Papp ◽  
Gyorgy Panyi ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Voltage Sensor ◽  
Ion Flow ◽  
Gating Model ◽  
Cryo Electron Microscopy ◽  
Channel Gate ◽  
Voltage Dependent ◽  
Voltage Gated Ion Channels ◽  
Voltage Sensing Domain ◽  
Pore Domain

Voltage-gated ion channels couple transmembrane potential changes to ion flow. Conformational changes in the voltage-sensing domain (VSD) of the channel are thought to be transmitted to the pore domain (PD) through an α-helical linker between them (S4–S5 linker). However, our recent work on channels disrupted in the S4–S5 linker has challenged this interpretation for the KCNH family. Furthermore, a recent single-particle cryo-electron microscopy structure of KV10.1 revealed that the S4–S5 linker is a short loop in this KCNH family member, confirming the need for an alternative gating model. Here we use “split” channels made by expression of VSD and PD as separate fragments to investigate the mechanism of gating in KV10.1. We find that disruption of the covalent connection within the S4 helix compromises the ability of channels to close at negative voltage, whereas disconnecting the S4–S5 linker from S5 slows down activation and deactivation kinetics. Surprisingly, voltage-clamp fluorometry and MTS accessibility assays show that the motion of the S4 voltage sensor is virtually unaffected when VSD and PD are not covalently bound. Finally, experiments using constitutively open PD mutants suggest that the presence of the VSD is structurally important for the conducting conformation of the pore. Collectively, our observations offer partial support to the gating model that assumes that an inward motion of the C-terminal S4 helix, rather than the S4–S5 linker, closes the channel gate, while also suggesting that control of the pore by the voltage sensor involves more than one mechanism.

scholarly journals Pulsed electric fields create pores in the voltage sensors of voltage-gated ion channels

2019 ◽  
Author(s):  
L. Rems ◽  
M. A. Kasimova ◽  
I. Testa ◽  
L. Delemotte
Keyword(s):  
Ion Channels ◽  
Membrane Proteins ◽  
Membrane Permeability ◽  
Electric Fields ◽  
Molecular Mechanisms ◽  
Pulsed Electric Fields ◽  
Pulse Treatment ◽  
Voltage Gated ◽  
Voltage Gated Ion Channels ◽  
Gated Ion Channels

AbstractPulsed electric fields are increasingly used in medicine to transiently increase the cell membrane permeability via electroporation, in order to deliver therapeutic molecules into the cell. One type of events that contributes to this increase in membrane permeability is the formation of pores in the membrane lipid bilayer. However, electrophysiological measurements suggest that membrane proteins are affected as well, particularly voltage-gated ion channels (VGICs). The molecular mechanisms by which the electric field could affects these molecules remain unidentified. In this study we used molecular dynamics (MD) simulations to unravel the molecular events that take place in different VGICs when exposing them to electric fields mimicking electroporation conditions. We show that electric fields induce pores in the voltage-sensor domains (VSDs) of different VGICs, and that these pores form more easily in some channels than in others. We demonstrate that poration is more likely in VSDs that are more hydrated and are electrostatically more favorable for the entry of ions. We further show that pores in VSDs can expand into so-called complex pores, which become stabilized by lipid head-groups. Our results suggest that such complex pores are considerably more stable than conventional lipid pores and their formation can lead to severe unfolding of VSDs from the channel. We anticipate that such VSDs become dysfunctional and unable to respond to changes in transmembrane voltage, which is in agreement with previous electrophyiological measurements showing a decrease in the voltage-dependent transmembrane ionic currents following pulse treatment. Finally, we discuss the possibility of activation of VGICs by submicrosecond-duration pulses. Overall our study reveals a new mechanism of electroporation through membranes containing voltage-gated ion channels.Statement of SignificancePulsed electric fields are often used for treatment of excitable cells, e.g., for gene delivery into skeletal muscles, ablation of the heart muscle or brain tumors. Voltage-gated ion channels (VGICs) underlie generation and propagation of action potentials in these cells, and consequently are essential for their proper function. Our study reveals the molecular mechanisms by which pulsed electric fields directly affect VGICs and addresses questions that have been previously opened by electrophysiologists. We analyze VGICs’ characteristics, which make them prone for electroporation, including hydration and electrostatic properties. This analysis is easily transferable to other membrane proteins thus opening directions for future investigations. Finally, we propose a mechanism for long-lived membrane permeability following pulse treatment, which to date remains poorly understood.

scholarly journals PIP2 mediates functional coupling and pharmacology of neuronal KCNQ channels

2017 ◽  
Vol 114 (45) ◽  
pp. E9702-E9711 ◽  
Author(s):  
Robin Y. Kim ◽  
Stephan A. Pless ◽  
Harley T. Kurata
Keyword(s):  
Conformational Changes ◽  
Neuronal Excitability ◽  
Voltage Dependence ◽  
Voltage Sensor ◽  
Functional Coupling ◽  
Voltage Sensing ◽  
Kcnq Channels ◽  
C Terminus ◽  
Hyperpolarizing Shift ◽  
State Ionic

Retigabine (RTG) is a first-in-class antiepileptic drug that suppresses neuronal excitability through the activation of voltage-gated KCNQ2–5 potassium channels. Retigabine binds to the pore-forming domain, causing a hyperpolarizing shift in the voltage dependence of channel activation. To elucidate how the retigabine binding site is coupled to changes in voltage sensing, we used voltage-clamp fluorometry to track conformational changes of the KCNQ3 voltage-sensing domains (VSDs) in response to voltage, retigabine, and PIP2. Steady-state ionic conductance and voltage sensor fluorescence closely overlap under basal PIP2 conditions. Retigabine stabilizes the conducting conformation of the pore and the activated voltage sensor conformation, leading to dramatic deceleration of current and fluorescence deactivation, but these effects are attenuated upon disruption of channel:PIP2 interactions. These findings reveal an important role for PIP2 in coupling retigabine binding to altered VSD function. We identify a polybasic motif in the proximal C terminus of retigabine-sensitive KCNQ channels that contributes to VSD–pore coupling via PIP2, and thereby influences the unique gating effects of retigabine.

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.

Sign in / Sign up

Export Citation Format

Share Document