Structure Of Potassium Channel Research Articles

Exploring Undergraduate Biochemistry Students' Gesture Production Through an Embodied Framework.

Interpreting three-dimensional models of biological macromolecules is a key skill in biochemistry, closely tied to students' visuospatial abilities. As students interact with these models and explain biochemical concepts, they often use gesture to complement verbal descriptions. Here, we utilize an embodied cognition-based approach to characterize undergraduate students' gesture production as they described and interpreted an augmented reality (AR) model of potassium channel structure and function. Our analysis uncovered two emergent patterns of gesture production employed by students, as well as common sets of gestures linked across categories of biochemistry content. Additionally, we present three cases that highlight changes in gesture production following interaction with a 3D AR visualization. Together, these observations highlight the importance of attending to gesture in learner-centered pedagogies in undergraduate biochemistry education.

Voltage-gated sodium channels (Na<sub>V</sub>) in GtoPdb v.2023.1

Sodium channels are voltage-gated sodium-selective ion channels present in the membrane of most excitable cells. Sodium channels comprise of one pore-forming α subunit, which may be associated with either one or two β subunits [179]. α-Subunits consist of four homologous domains (I-IV), each containing six transmembrane segments (S1-S6) and a pore-forming loop. The positively charged fourth transmembrane segment (S4) acts as a voltage sensor and is involved in channel gating. The crystal structure of the bacterial NavAb channel has revealed a number of novel structural features compared to earlier potassium channel structures including a short selectivity filter with ion selectivity determined by interactions with glutamate side chains [278]. Interestingly, the pore region is penetrated by fatty acyl chains that extend into the central cavity which may allow the entry of small, hydrophobic pore-blocking drugs [278]. Auxiliary β1, β2, β3 and β4 subunits consist of a large extracellular N-terminal domain, a single transmembrane segment and a shorter cytoplasmic domain.The nomenclature for sodium channels was proposed by Goldin et al., (2000) [146] and approved by the NC-IUPHAR Subcommittee on sodium channels (Catterall et al., 2005, [53]).

Ion Conduction Mechanisms in Potassium Channels Revealed by Permeation Cycles.

Potassium channels are responsible for the selective yet efficient permeation of potassium ions across cell membranes. Despite many available high-resolution structures of potassium channels, those conformations inform only on static information on the ion permeation processes. Here, we use molecular dynamics simulations and Markov state models to obtain dynamical details of ion permeation. The permeation cycles, expressed in terms of selectivity filter occupancy and representing ion permeation events, are illustrated. We show that the direct knock-on permeation represents the dominant permeation mechanism over a wide range of potassium concentrations, temperatures, and membrane voltages for the pore of MthK. Direct knock-on is also observed in other potassium channels with a highly conserved selectivity filter, demonstrating the robustness of the permeation mechanism. Lastly, we investigate the charge strength dependence of permeation cycles. Our results shed light on the underlying permeation details, which are valuable in studying conduction mechanisms in potassium channels.

DIMPLE: deep insertion, deletion, and missense mutation libraries for exploring protein variation in evolution, disease, and biology

Insertions and deletions (indels) enable evolution and cause disease. Due to technical challenges, indels are left out of most mutational scans, limiting our understanding of them in disease, biology, and evolution. We develop a low cost and bias method, DIMPLE, for systematically generating deletions, insertions, and missense mutations in genes, which we test on a range of targets, including Kir2.1. We use DIMPLE to study how indels impact potassium channel structure, disease, and evolution. We find deletions are most disruptive overall, beta sheets are most sensitive to indels, and flexible loops are sensitive to deletions yet tolerate insertions.

Ball-and-Chain Inactivation in Potassium Channels.

Carefully orchestrated opening and closing of ion channels control the diffusion of ions across cell membranes, generating the electrical signals required for fast transmission of information throughout the nervous system. Inactivation is a parsimonious means for channels to restrict ion conduction without the need to remove the activating stimulus. Voltage-gated channel inactivation plays crucial physiological roles, such as controlling action potential duration and firing frequency in neurons. The ball-and-chain moniker applies to a type of inactivation proposed first for sodium channels and later shown to be a universal mechanism. Still, structural evidence for this mechanism remained elusive until recently. We review the ball-and-chain inactivation research starting from its introduction as a crucial component of sodium conductance during electrical signaling in the classical Hodgkin and Huxley studies, through the discovery of its simple intuitive mechanism in potassium channels during the molecular cloning era, to the eventual elucidation of a potassium channel structure in a ball-and-chain inactivated state.

Early Steps in C-Type Inactivation of the hERG Potassium Channel.

Fast C-type inactivation confers distinctive functional properties to the hERG potassium channel, and its association to inherited and acquired cardiac arrythmias makes the study of the inactivation mechanism of hERG at the atomic detail of paramount importance. At present, two models have been proposed to describe C-type inactivation in K+-channels. Experimental data and computational work on the bacterial KcsA channel support the hypothesis that C-type inactivation results from a closure of the selectivity filter that sterically impedes ion conduction. Alternatively, recent experimental structures of a mutated Shaker channel revealed a widening of the extracellular portion of the selectivity filter, which might diminish conductance by interfering with the mechanism of ion permeation. Here, we performed molecular dynamics simulations of the wild-type hERG, a non-inactivating mutant (hERG-N629D), and a mutant that inactivates faster than the wild-type channel (hERG-F627Y) to find out which and if any of the two reported C-type inactivation mechanisms applies to hERG. Closure events of the selectivity filter were not observed in any of the simulated trajectories but instead, the extracellular section of the selectivity filter deviated from the canonical conductive structure of potassium channels. The degree of widening of the potassium binding sites at the extracellular entrance of the channel was directly related to the degree of inactivation with hERG-F627Y > wild-type hERG > hERG-N629D. These findings support the hypothesis that C-type inactivation in hERG entails a widening of the extracellular entrance of the channel rather than a closure of the selectivity filter.

Predicting the functional impact of KCNQ1 variants with artificial neural networks.

Recent advances in experimental and computational protein structure determination have provided access to high-quality structures for most human proteins and mutants thereof. However, linking changes in structure in protein mutants to functional impact remains an active area of method development. If successful, such methods can ultimately assist physicians in taking appropriate treatment decisions. This work presents three artificial neural network (ANN)-based predictive models that classify four key functional parameters of KCNQ1 variants as normal or dysfunctional using PSSM-based evolutionary and/or biophysical descriptors. Recent advances in predicting protein structure and variant properties with artificial intelligence (AI) rely heavily on the availability of evolutionary features and thus fail to directly assess the biophysical underpinnings of a change in structure and/or function. The central goal of this work was to develop an ANN model based on structure and physiochemical properties of KCNQ1 potassium channels that performs comparably or better than algorithms using only on PSSM-based evolutionary features. These biophysical features highlight the structure-function relationships that govern protein stability, function, and regulation. The input sensitivity algorithm incorporates the roles of hydrophobicity, polarizability, and functional densities on key functional parameters of the KCNQ1 channel. Inclusion of the biophysical features outperforms exclusive use of PSSM-based evolutionary features in predicting activation voltage dependence and deactivation time. As AI is increasingly applied to problems in biology, biophysical understanding will be critical with respect to 'explainable AI', i.e., understanding the relation of sequence, structure, and function of proteins. Our model is available at www.kcnq1predict.org.

An epilepsy-associated KV1.2 charge-transfer-center mutation impairs KV1.2 and KV1.4 trafficking

We report on a heterozygous KCNA2 variant in a child with epilepsy. KCNA2 encodes KV1.2 subunits, which form homotetrameric potassium channels and participate in heterotetrameric channel complexes with other KV1-family subunits, regulating neuronal excitability. The mutation causes substitution F233S at the KV1.2 charge transfer center of the voltage-sensing domain. Immunocytochemical trafficking assays showed that KV1.2(F233S) subunits are trafficking deficient and reduce the surface expression of wild-type KV1.2 and KV1.4: a dominant-negative phenotype extending beyond KCNA2, likely profoundly perturbing electrical signaling. Yet some KV1.2(F233S) trafficking was rescued by wild-type KV1.2 and KV1.4 subunits, likely in permissible heterotetrameric stoichiometries: electrophysiological studies utilizing applied transcriptomics and concatemer constructs support that up to one or two KV1.2(F233S) subunits can participate in trafficking-capable heterotetramers with wild-type KV1.2 or KV1.4, respectively, and that both early and late events along the biosynthesis and secretion pathway impair trafficking. These studies suggested that F233S causes a depolarizing shift of ∼48 mV on KV1.2 voltage dependence. Optical tracking of the KV1.2(F233S) voltage-sensing domain (rescued by wild-type KV1.2 or KV1.4) revealed that it operates with modestly perturbed voltage dependence and retains pore coupling, evidenced by off-charge immobilization. The equivalent mutation in the Shaker K+ channel (F290S) was reported to modestly affect trafficking and strongly affect function: an ∼80-mV depolarizing shift, disrupted voltage sensor activation and pore coupling. Our work exposes the multigenic, molecular etiology of a variant associated with epilepsy and reveals that charge-transfer-center disruption has different effects in KV1.2 and Shaker, the archetypes for potassium channel structure and function.

Voltage-gated sodium channels (Na<sub>V</sub>) in GtoPdb v.2021.3

Sodium channels are voltage-gated sodium-selective ion channels present in the membrane of most excitable cells. Sodium channels comprise of one pore-forming α subunit, which may be associated with either one or two β subunits [177]. α-Subunits consist of four homologous domains (I-IV), each containing six transmembrane segments (S1-S6) and a pore-forming loop. The positively charged fourth transmembrane segment (S4) acts as a voltage sensor and is involved in channel gating. The crystal structure of the bacterial NavAb channel has revealed a number of novel structural features compared to earlier potassium channel structures including a short selectivity filter with ion selectivity determined by interactions with glutamate side chains [274]. Interestingly, the pore region is penetrated by fatty acyl chains that extend into the central cavity which may allow the entry of small, hydrophobic pore-blocking drugs [274]. Auxiliary β1, β2, β3 and β4 subunits consist of a large extracellular N-terminal domain, a single transmembrane segment and a shorter cytoplasmic domain.The nomenclature for sodium channels was proposed by Goldin et al., (2000) [144] and approved by the NC-IUPHAR Subcommittee on sodium channels (Catterall et al., 2005, [52]).

Tetraoctylammonium, a Long Chain Quaternary Ammonium Blocker, Promotes a Noncollapsed, Resting-Like Inactivated State in KcsA.

Alkylammonium salts have been used extensively to study the structure and function of potassium channels. Here, we use the hydrophobic tetraoctylammonium (TOA+) to shed light on the structure of the inactivated state of KcsA, a tetrameric prokaryotic potassium channel that serves as a model to its homologous eukaryotic counterparts. By the combined use of a thermal denaturation assay and the analysis of homo-Förster resonance energy transfer in a mutant channel containing a single tryptophan (W67) per subunit, we found that TOA+ binds the channel cavity with high affinity, either with the inner gate open or closed. Moreover, TOA+ bound at the cavity allosterically shifts the equilibrium of the channel’s selectivity filter conformation from conductive to an inactivated-like form. The inactivated TOA+–KcsA complex exhibits a loss in the affinity towards permeant K+ at pH 7.0, when the channel is in its closed state, but maintains the two sets of K+ binding sites and the W67–W67 intersubunit distances characteristic of the selectivity filter in the channel resting state. Thus, the TOA+–bound state differs clearly from the collapsed channel state described by X-ray crystallography and claimed to represent the inactivated form of KcsA.

Thermodynamics of ion binding and occupancy in potassium channels.

Potassium channels modulate various cellular functions through efficient and selective conduction of K+ ions. The mechanism of ion conduction in potassium channels has recently emerged as a topic of debate. Crystal structures of potassium channels show four K+ ions bound to adjacent binding sites in the selectivity filter, while chemical intuition and molecular modeling suggest that the direct ion contacts are unstable. Molecular dynamics (MD) simulations have been instrumental in the study of conduction and gating mechanisms of ion channels. Based on MD simulations, two hypotheses have been proposed, in which the four-ion configuration is an artifact due to either averaged structures or low temperature in crystallographic experiments. The two hypotheses have been supported or challenged by different experiments. Here, MD simulations with polarizable force fields validated by ab initio calculations were used to investigate the ion binding thermodynamics. Contrary to previous beliefs, the four-ion configuration was predicted to be thermodynamically stable after accounting for the complex electrostatic interactions and dielectric screening. Polarization plays a critical role in the thermodynamic stabilities. As a result, the ion conduction likely operates through a simple single-vacancy and water-free mechanism. The simulations explained crystal structures, ion binding experiments and recent controversial mutagenesis experiments. This work provides a clear view of the mechanism underlying the efficient ion conduction and demonstrates the importance of polarization in ion channel simulations.

Barium blockade of the KcsA channel in open and closed conformation datasets.

Barium is a potent blocker of the KcsA potassium channel. A strategy using x-ray crystallography and molecular dynamics (MD) simulation has been used to understand this phenomenon as described in Rohaim et al. [1]. Wild type KcsA is purified to homogeneity and crystallized in low and high K+ conditions. Crystals are grown using the hanging drop vapor diffusion method. To examine barium binding in the selectivity filter of KcsA, the crystals are systemically soaked in various concentrations of barium chloride solution. X-ray crystallography datasets are collected at the Advanced Photon Source. A total of 10 datasets are collected for various barium ion concentrations. Diffraction data are processed using the crystallography pipeline software RAPID. The crystal structures are solved by molecular replacement methods. The structure models are visualized using COOT and refined using REFMAC. Anomalous map coefficients are calculated using the phenix.maps tool in the PHENIX software suite. The datasets are deposited in the Protein Data Bank. The data provides a detailed picture of barium ion interaction with potassium channels. Structural analysis of the KcsA channel reveals two distinct configurations, open- and closed- state. Further MD simulation analysis suggests an energetically favorable binding mechanism for barium ion in the selectivity filter. The data could be used to interpret functional experiments related to barium blockade for potassium channels. Also, it is valuable for comparison and cross validation with other relevant potassium channel structures.

Generation of induced pluripotent stem cell GZHMCi001-A and GZHMCi001-B derived from peripheral blood mononuclear cells of epileptic patients with KCNC1 mutation.

Myoclonus Epilepsy and Ataxia due to Potassium channel mutation (MEAK) is a rare epilepsy caused by changes in the structure and function of potassium channels due to mutations in the potassium voltage-gated channel subfamily C member 1 (KCNC1) gene. MEAK is one of the progressive myoclonus epilepsy (PME), and there are few studies on MEAK pathogenesis and targeted drugs. Here, we used peripheral blood from MEAK patients with KCNC1 (c.959G>A) gene mutation to establish induced pluripotent stem cells (iPSC). The iPSC of KCNC1 mutation established by us is a powerful tool for related research.

Anionic Lipids Modulate Structure and Function of Epilepsy-Causing Voltage-Gated Potassium Channel

Anionic Lipids Modulate Structure and Function of Epilepsy-Causing Voltage-Gated Potassium Channel

Structure of Lymphocyte Potassium Channel K <sub>v</sub>1.3 and Modulation by Cell-Penetrating Immunomodulatory Plant Defensin

Author(s): Ong, Seow Theng; Liu, Sanling; Bajaj, Saumya; Zhao, Yue; Tanner, Mark R; Chang, Shih Chieh; Zhang, Yong; Krishnarjuna, Bankala; Ng, Xuan Rui; Morales, Rodrigo AV; Ming, Wei Chen; Luo, Dahai; Patel, Dharmeshkumar; Yasmin, Sabina; Ng, Jeremy Jun Heng; Nguyen, Hai M; Abbas, El Sahili; Lescar, Julien; Patil, Rahul; Charman, Susan A; Robins, Edward; Goggi, Julian; Peng, Wen Tan; Sadasivam, Pragalath; Ramasamy, Boominathan; SV, Hartimath; Dhawan, Vikas; Bednenko, Janna; Colussi, Paul; Wulff, Heike; Zhang, Longhua; Pennington, Michael; Kuyucak, Serdar; Norton, Raymond S; Beeton, Christine; Tian, Changlin; Chandy, George

Voltage-gated sodium channels (version 2019.4) in the IUPHAR/BPS Guide to Pharmacology Database

Sodium channels are voltage-gated sodium-selective ion channels present in the membrane of most excitable cells. Sodium channels comprise of one pore-forming α subunit, which may be associated with either one or two β subunits [176]. α-Subunits consist of four homologous domains (I–IV), each containing six transmembrane segments (S1–S6) and a pore-forming loop. The positively charged fourth transmembrane segment (S4) acts as a voltage sensor and is involved in channel gating. The crystal structure of the bacterial NavAb channel has revealed a number of novel structural features compared to earlier potassium channel structures including a short selectivity filter with ion selectivity determined by interactions with glutamate side chains [268]. Interestingly, the pore region is penetrated by fatty acyl chains that extend into the central cavity which may allow the entry of small, hydrophobic pore-blocking drugs [268]. Auxiliary β1, β2, β3 and β4 subunits consist of a large extracellular N-terminal domain, a single transmembrane segment and a shorter cytoplasmic domain.The nomenclature for sodium channels was proposed by Goldin et al., (2000) [143] and approved by the NC-IUPHAR Subcommittee on sodium channels (Catterall et al., 2005, [51]).

How RCK domains regulate gating of K+ channels.

Potassium channels play a crucial role in the physiology of all living organisms. They maintain the membrane potential and are involved in electrical signaling, pH homeostasis, cell-cell communication and survival under osmotic stress. Many prokaryotic potassium channels and members of the eukaryotic Slo channels are regulated by tethered cytoplasmic domains or associated soluble proteins, which belong to the family of regulator of potassium conductance (RCK). RCK domains and subunits form octameric rings, which control ion gating. For years, a common regulatory mechanism was suggested: ligand-induced conformational changes in the octameric ring would pull open a gate in the pore via flexible linkers. Consistently, ligand-dependent conformational changes were described for various RCK gating rings. Yet, recent structural and functional data of complete ion channels uncovered that the following signal transduction to the pore domains is divers. The different RCK-regulated ion channels show remarkably heterogeneous mechanisms with neither the connection from the RCK domain to the pore nor the gate being conserved. Some channels even lack the flexible linkers, while in others the gate cannot easily be assigned. In this review we compare available structures of RCK-gated potassium channels, highlight the similarities and differences of channel gating, and delineate existing inconsistencies.

Генетика мигрени: ассоциативные исследования

Статья посвящена анализу имеющихся на сегодняшний день данных генетических полиморфизмов, излучавшихся при мигрени. При мигрени гены-кандидаты мигрени сгруппированы в четыре функциональных семейства генов: гены нервной системы, гены сердечно-сосудистой системы, гормональные, воспалительные гены. Среди генов нервной системы значимость представляют гены, кодирующие структуру и функциональную активность калиевых каналов. Наибольший интерес представляют гены сердечно-сосудистой системы, которые как повышают риск мигрени, так и увеличивают риск коморбидных кардиоваскулярных заболеваний. Среди генов эндокринной системы роль в патогенезе мигрени играют полиморфизмы гена эстрогенового рецептора 1 типа. Для воспалительных генов циклооксегиназы 2, главного комплекса гистосовместимости HLA-DRB1, лимфотоксина альфа, фактора некроза опухоли альфа и бета найдены положительные ассоциации с мигренью. The article focused on analysis of currently available reports on genetic polymorphisms in migraine. The migraine candidate genes were grouped into four functional families: genes of the nervous system, genes of the cardiovascular system, hormonal genes, and inflammatory genes. Among the nervous system genes, the genes encoding the structure and functional activity of potassium channels are of a special significance. Of the greatest interest are the cardiovascular system genes, which increase the risk of both migraine and comorbid cardiovascular diseases. Among the endocrine system genes, polymorphisms of the type 1 estrogen receptor gene play a role in the pathogenesis of migraine. Positive associations with migraine were found for inflammatory genes of cyclooxygenase 2, the main histocompatibility complex HLA-DRB1, lymphotoxin alpha, and tumor necrosis factors alpha and beta.

Predicting Structural Details of the Sodium Channel Pore Basing on Animal Toxin Studies.

Eukaryotic voltage-gated sodium channels play key roles in physiology and are targets for many toxins and medically important drugs. Physiology, pharmacology, and general architecture of the channels has long been the subject of intensive research in academia and industry. In particular, animal toxins such as tetrodotoxin, saxitoxin, and conotoxins have been used as molecular probes of the channel structure. More recently, X-ray structures of potassium and prokaryotic sodium channels allowed elaborating models of the toxin-channel complexes that integrated data from biophysical, electrophysiological, and mutational studies. Atomic level cryo-EM structures of eukaryotic sodium channels, which became available in 2017, show that the selectivity filter structure and other important features of the pore domain have been correctly predicted. This validates further employments of toxins and other small molecules as sensitive probes of fine structural details of ion channels.

From in silico to in vitro: a trip to reveal flavonoid binding on the Rattus norvegicus Kir6.1 ATP-sensitive inward rectifier potassium channel.

BackgroundATP-sensitive inward rectifier potassium channels (Kir), are a potassium channel family involved in many physiological processes. KATP dysfunctions are observed in several diseases such as hypoglycaemia, hyperinsulinemia, Prinzmetal angina–like symptoms, cardiovascular diseases.MethodsA broader view of the KATP mechanism is needed in order to operate on their regulation, and in this work we clarify the structure of the Rattus norvegicus ATP-sensitive inward rectifier potassium channel 8 (Kir6.1), which has been obtained through a homology modelling procedure. Due to the medical use of flavonoids, a considerable increase in studies on their influence on human health has recently been observed, therefore our aim is to study, through computational methods, the three-dimensional (3D) conformation together with mechanism of action of Kir6.1 with three flavonoids.ResultsComputational analysis by performing molecular dynamics (MD) and docking simulation on rat 3D modelled structure have been completed, in its closed and open conformation state and in complex with Quercetin, 5-Hydroxyflavone and Rutin flavonoids. Our study showed that only Quercetin and 5-Hydroxyflavone were responsible for a significant down-regulation of the Kir6.1 activity, stabilising it in a closed conformation. This hypothesis was supported by in vitro experiments demonstrating that Quercetin and 5-Hydroxyflavone were capable to inhibit KATP currents of rat tail main artery myocytes recorded by the patch-clamp technique.ConclusionCombined methodological approaches, such as molecular modelling, docking and MD simulations of Kir6.1 channel, used to elucidate flavonoids intrinsic mechanism of action, are introduced, revealing a new potential druggable protein site.