Structure Of Ion Channels Research Articles

Structural studies of ion channels: achievements, problems and perspectives

The superfamily of membrane proteins known as P-loop channels encompasses potassium, sodium, and calcium channels, as well as TRP channels and ionotropic glutamate receptors. An increasing number of crystal and cryo-EM structures are uncovering both general and specific features of these channels. Fundamental folding principles, the arrangement of structural segments, key residues that influence ionic selectivity, gating, and binding sites for toxins and medically relevant ligands have now been firmly established. The advent of AlphaFold2 (AF2) models represents another significant step in computationally predicting protein structures. Comparison of experimental P-loop channel structures with their corresponding AF2 models shows consistent folding patterns in experimentally resolved regions. Despite this remarkable progress, many crucial structural details, particularly important for predicting the outcomes of mutations and designing new medically relevant ligands, remain unresolved. Certain methodological challenges currently hinder the direct assessment of such details. Until the next methodological breakthrough occurs, a promising approach to analyzing ion channel structures in greater depth involves integrating various experimental and theoretical methods.

Voltage-Gated Ion Channels: Structure, Pharmacology and Photopharmacology.

Voltage-gated ion channels are transmembrane proteins responsible for the generation and propagation of action potentials in excitable cells. Over the last decade, advancements have enabled the elucidation of crystal structures of ion channels. This progress in structural understanding, particularly in identifying the binding sites of local anesthetics, opens avenues for the design of novel compounds capable of modulating ion conduction. However, many traditional drugs lack selectivity and come with adverse side effects. The emergence of photopharmacology has provided an orthogonal way of controlling the activity of compounds, enabling the regulation of ion conduction with light. In this review, we explore the central pore region of voltage-gated sodium and potassium channels, providing insights from both structural and pharmacological perspectives. We discuss the different binding modes of synthetic compounds that can physically occlude the pore and, therefore, block ion conduction. Moreover, we examine recent advances in the photopharmacology of voltage-gated ion channels, introducing molecular approaches aimed at controlling their activity by using photosensitive drugs.

Exploring the influence of pore shape on conductance and permeation

There are increasing numbers of ion channel structures featuring heteromeric subunit assembly, exemplified by synaptic α1βB glycine and α4β2 nicotinic receptors. These structures exhibit inherent pore asymmetry, but the relevance of this to function is unknown. Furthermore, molecular dynamics simulations performed on symmetrical homomeric channels often lead to thermal distortion whereby conformations of the resulting ensemble are also asymmetrical. When functionally annotating ion channels, researchers often rely on minimal constrictions determined via radius-profile calculations performed with computer programs, such as HOLE or CHAP, coupled with an assessment of pore hydrophobicity. However, such tools typically employ spherical probe particles, limiting their ability to accurately capture pore asymmetry. Here, we introduce an algorithm that employs ellipsoidal probe particles, enabling a more comprehensive representation of the pore geometry. Our analysis reveals that the use of nonspherical ellipsoids for pore characterization provides a more accurate and easily interpretable depiction of conductance. To quantify the implications of pore asymmetry on conductance, we systematically investigated carbon nanotubes with varying degrees of pore asymmetry as model systems. The conductance through these channels shows surprising effects that would otherwise not be predicted with spherical probes. The results have broad implications not only for the functional annotation of biological ion channels but also for the design of synthetic channel systems for use in areas such as water filtration. Furthermore, we make use of the more accurate characterization of channel pores to refine a physical conductance model to obtain a heuristic estimate for single-channel conductance. The code is freely available, obtainable as pip-installable python package and provided as a web service.

Sodium Bicarbonate Decreases Alcohol Consumption in Mice

pH is an important biological factor in the brain as H+ alters the structure and function of many ion channels, transporters and receptors, thus affecting relevant synaptic activities and complex firing patterns. pH may change activities in the neural circuits responsible for motivation and reward, such that drug reward values or modulatory properties are modified to change drinking frequency and amount. In this study, we tested whether alcohol consumption can be affected by systemic pH in mice. C57BL/6J mice were given free access to 0.2 M NaHCO3 for 6 days, which resulted in a 0.2 pH unit increase compared to water-fed control mice ( p < 0.05; n = 6/group). Mice were then subjected to voluntary alcohol consumption in the two-bottle free choice procedure with one bottle containing water and the other containing 5-20% alcohol (5% increment for 4 days each). NaHCO3-fed mice consumed similar amounts of 5-15% alcohol as pair-fed mice but decreased the consumption of 20% alcohol (p < 0.05). Consistent with this result, NaHCO3-fed mice had decreased alcohol preference (i.e., alcohol intake/total fluid intake) at 20%. These changes were observed in both males and females. In separate experiments, mice were subjected to the two-bottle free choice procedure using water and 20% alcohol, with both bottles containing 0.05-0.4 M NaHCO3. The alcohol consumption was significantly decreased in the presence of 0.1 M or higher NaHCO3. The minimum effective dose of NaHCO3 was 0.08 g/kg body weight, calculated from the amount of alcohol mice consumed. Determined by the diarrhea scoring system, mice produced a normal state of feces after receiving 0.4 M NaHCO3 for 2 days, thus indicating negligible abdominal discomfort at this concentration. In other experiments, mice were given NaHCO3 by oral gavage and 1 hour later subjected to 20% alcohol in the drinking in the dark procedure. The alcohol consumption was significantly decreased in NaHCO3-fed mice, compared to pair-fed mice. In conclusion, our study shows that mildly raising blood pH by NaHCO3 decreases alcohol consumption in mice. Given that pH change has been considered as a consequence of alcohol metabolism, our data provide first evidence on the pH as a factor affecting alcohol consumption. This work was supported by the grant from the National Institute of Alcohol Abuse and Alcoholism (R21AA028606). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

TRPA5 encodes a thermosensitive ankyrin ion channel receptor in a triatomine insect

As ectotherms, insects need heat-sensitive receptors to monitor environmental temperatures and facilitate thermoregulation. We show that TRPA5, a class of ankyrin transient receptor potential (TRP) channels absent in dipteran genomes, may function as insect heat receptors. In the triatomine bug Rhodnius prolixus (order: Hemiptera), a vector of Chagas disease, the channel RpTRPA5B displays a uniquely high thermosensitivity, with biophysical determinants including a large channel activation enthalpy change (72kcal/mol), a high temperature coefficient (Q10= 25), and invitro temperature-induced currents from 53°C to 68°C (T0.5= 58.6°C), similar to noxious TRPV receptors in mammals. Monomeric and tetrameric ion channel structure predictions show reliable parallels with fruit fly dTRPA1, with structural uniqueness in ankyrin repeat domains, the channel selectivity filter, and potential TRP functional modulator regions. Overall, the finding of a member of TRPA5 as a temperature-activated receptor illustrates the diversity of insect molecular heat detectors.

Structural modeling of ion channels using AlphaFold2, RoseTTAFold2, and ESMFold

ABSTRACT Ion channels play key roles in human physiology and are important targets in drug discovery. The atomic-scale structures of ion channels provide invaluable insights into a fundamental understanding of the molecular mechanisms of channel gating and modulation. Recent breakthroughs in deep learning-based computational methods, such as AlphaFold, RoseTTAFold, and ESMFold have transformed research in protein structure prediction and design. We review the application of AlphaFold, RoseTTAFold, and ESMFold to structural modeling of ion channels using representative voltage-gated ion channels, including human voltage-gated sodium (NaV) channel - NaV1.8, human voltage-gated calcium (CaV) channel – CaV1.1, and human voltage-gated potassium (KV) channel – KV1.3. We compared AlphaFold, RoseTTAFold, and ESMFold structural models of NaV1.8, CaV1.1, and KV1.3 with corresponding cryo-EM structures to assess details of their similarities and differences. Our findings shed light on the strengths and limitations of the current state-of-the-art deep learning-based computational methods for modeling ion channel structures, offering valuable insights to guide their future applications for ion channel research.

Structural Insights and Influence of Terahertz Waves in Midinfrared Region on Kv1.2 Channel Selectivity Filter.

Potassium ion channels are the structural basis for excitation transmission, heartbeat, and other biological processes. The selectivity filter is a critical structural component of potassium ion channels, whose structure is crucial to realizing their function. As biomolecules vibrate and rotate at frequencies in the terahertz band, potassium ion channels are sensitive to terahertz waves. Therefore, it is worthwhile to investigate how the terahertz wave influences the selectivity filter of the potassium channels. In this study, we investigate the structure of the selectivity filter of Kv1.2 potassium ion channels using molecular dynamics simulations. The effect of an electric field on the channel has been examined at four different resonant frequencies of the carbonyl group in SF: 36.75 37.06, 37.68, and 38.2 THz. As indicated by the results, 376GLY appears to be the critical residue in the selectivity filter of the Kv1.2 channel. Its dihedral angle torsion is detrimental to the channel structural stability and the transmembrane movement of potassium ions. 36.75 THz is the resonance frequency of the carbonyl group of 376GLY. Among all four frequencies explored, the applied terahertz electric field of this frequency has the most significant impact on the channel structure, negatively impacting the channel stability and reducing the ion permeability by 20.2% compared to the absence of fields. In this study, we simulate that terahertz waves in the mid-infrared frequency region can significantly alter the structure and function of potassium ion channels and that the effects of terahertz waves differ greatly based on frequency.

Structure of human TRPV2 ion channel informs differences in activation properties

Structure of human TRPV2 ion channel informs differences in activation properties

Lipid nanodisc scaffold and size alter the structure of a pentameric ligand-gated ion channel

Lipid nanodiscs have become a standard tool for studying membrane proteins, including using single particle cryo-electron microscopy (cryo-EM). We find that reconstituting the pentameric ligand-gated ion channel (pLGIC), Erwinia ligand-gated ion channel (ELIC), in different nanodiscs produces distinct structures by cryo-EM. The effect of the nanodisc on ELIC structure extends to the extracellular domain and agonist binding site. Additionally, molecular dynamic simulations indicate that nanodiscs of different size impact ELIC structure and that the nanodisc scaffold directly interacts with ELIC. These findings suggest that the nanodisc plays a crucial role in determining the structure of pLGICs, and that reconstitution of ion channels in larger nanodiscs may better approximate a lipid membrane environment.

Electrophysiological evaluation of the effect of peptide toxins on voltage-gated ion channels: a scoping review on theoretical and methodological aspects with focus on the Central and South American experience.

The effect of peptide toxins on voltage-gated ion channels can be reliably assessed using electrophysiological assays, such as the patch-clamp technique. However, much of the toxinological research done in Central and South America aims at purifying and characterizing biochemical properties of the toxins of vegetal or animal origin, lacking electrophysiological approaches. This may happen due to technical and infrastructure limitations or because researchers are unfamiliar with the techniques and cellular models that can be used to gain information about the effect of a molecule on ion channels. Given the potential interest of many research groups in the highly biodiverse region of Central and South America, we reviewed the most relevant conceptual and methodological developments required to implement the evaluation of the effect of peptide toxins on mammalian voltage-gated ion channels using patch-clamp. For that, we searched MEDLINE/PubMed and SciELO databases with different combinations of these descriptors: "electrophysiology", "patch-clamp techniques", "Ca2+ channels", "K+ channels", "cnidarian venoms", "cone snail venoms", "scorpion venoms", "spider venoms", "snake venoms", "cardiac myocytes", "dorsal root ganglia", and summarized the literature as a scoping review. First, we present the basics and recent advances in mammalian voltage-gated ion channel's structure and function and update the most important animal sources of channel-modulating toxins (e.g. cnidarian and cone snails, scorpions, spiders, and snakes), highlighting the properties of toxins electrophysiologically characterized in Central and South America. Finally, we describe the local experience in implementing the patch-clamp technique using two models of excitable cells, as well as the participation in characterizing new modulators of ion channels derived from the venom of a local spider, a toxins' source less studied with electrophysiological techniques. Fostering the implementation of electrophysiological methods in more laboratories in the region will strengthen our capabilities in many fields, such as toxinology, toxicology, pharmacology, natural products, biophysics, biomedicine, and bioengineering.

Bioinspired ion channel receptor based on hygroelectricity for precontact sensing of living organism

Tactile sensors play an important role in human-machine interaction (HMI). Compared to contact tactile sensing, which leaves physical hardware vulnerable to wear and tear, proximity sensing is better at reacting to remote events before physical contact. The apteronotus albifrons possess ion channel receptors for remote surroundings perception. Inspired by the relevant ion channel structure and self-powered operation mode, we designed a new proximity sensor with ion rectification characteristics and self-powered capability. This bio-inspired ion channel receptor exploits the hygroelectric effect to convert the humidity information into a series of current signals when the living organism approaches, and it is insensitive to non-aquatic non-organisms. The sensor offers high sensitivity (2.3 mm−1), a suitable range (0–10 mm) for close object detection, fast response (0.3 s), and fast recovery (2.5 s). The unique combination of bio-sensitivity, non-contact detection characteristics, and humidity-based power generation capabilities enriches the functionality of future HMI electronics. As a proof of concept, the sensor has been successfully applied in different scenarios such as human health management, early warning systems, non-contact switches to prevent virus transmission, object recognition, and finger trajectory detection.

Abstract 13049: Electrophysiological Mechanism of Potassium Channel KCNQ1 p.V162M Mutation Induced Atrial Tachycardia and Epilepsy

Objective: Arrhythmia and epilepsy are the causes of syncope and sudden death. Both of them are associated with abnormal function of potassium ion channels. Delayed rectified potassium current (IKs) is generated by combining the α subunit of potassium ion channel encoded by KCNQ1 gene with the β subunit encoded by KCNE1 gene. It plays an important role in electrical excitation and K+ circulation of nerve cells. In this study, a patient with atrial arrhythmia and epilepsy with a history of syncope was identified by genetic testing as having a mutation in the KCNQ1 p.V162M gene. The purpose of this study was to investigate the effect of KCNQ1 p.V162M mutation on the structure and function of potassium ion channels. Methods: The mutant plasmid KCNQ1-p.V162M was constructed for cell transfection. The expression of protein and mRNA were analyzed by western blotting and real-time quantitative PCR. The IKs of the two groups were detected by whole-cell patch clamp technique. Results: Whole exon gene sequencing revealed that the patient carried the KCNQ1 p.V162M mutation. The results of molecular biology experiment showed that mRNA, KCNQ1 total protein and membrane protein expressions in mutant group were significantly lower than those in wild group. Whole-cell patch clamp results showed that the peak current density and tail current of IKs in the mutant group were significantly decreased compared with those in the wild group, indicating that the mutation of KCNQ1 p.V162M could lead to decreased function of potassium ion channels. Conclusions: KCNQ1 p.V162M mutation can lead to decreased expression of KCNQ1 protein and decreased function of potassium ion channel, which may be the potential mechanism of atrial tachycardia and epilepsy induced by this mutation.

Molecular tuning of sea anemone stinging.

Jellyfish and sea anemones fire single-use, venom-covered barbs to immobilize prey or predators. We previously showed that the anemone Nematostella vectensis uses a specialized voltage-gated calcium (CaV) channel to trigger stinging in response to synergistic prey-derived chemicals and touch (Weir et al., 2020). Here, we use experiments and theory to find that stinging behavior is suited to distinct ecological niches. We find that the burrowing anemone Nematostella uses uniquely strong CaV inactivation for precise control of predatory stinging. In contrast, the related anemone Exaiptasia diaphana inhabits exposed environments to support photosynthetic endosymbionts. Consistent with its niche, Exaiptasia indiscriminately stings for defense and expresses a CaV splice variant that confers weak inactivation. Chimeric analyses reveal that CaVβ subunit adaptations regulate inactivation, suggesting an evolutionary tuning mechanism for stinging behavior. These findings demonstrate how functional specialization of ion channel structure contributes to distinct organismal behavior.

Molecular tuning of sea anemone stinging

Jellyfish and sea anemones fire single-use, venom-covered barbs to immobilize prey or predators. We previously showed that the anemone Nematostella vectensis uses a specialized voltage-gated calcium (CaV) channel to trigger stinging in response to synergistic prey-derived chemicals and touch (Weir et al., 2020). Here, we use experiments and theory to find that stinging behavior is suited to distinct ecological niches. We find that the burrowing anemone Nematostella uses uniquely strong CaV inactivation for precise control of predatory stinging. In contrast, the related anemone Exaiptasia diaphana inhabits exposed environments to support photosynthetic endosymbionts. Consistent with its niche, Exaiptasia indiscriminately stings for defense and expresses a CaV splice variant that confers weak inactivation. Chimeric analyses reveal that CaVβ subunit adaptations regulate inactivation, suggesting an evolutionary tuning mechanism for stinging behavior. These findings demonstrate how functional specialization of ion channel structure contributes to distinct organismal behavior.

Design and One-Pot Ultrasound Synthesis of Inorganic Base-Promoted Fluorescent Ligand-Gated Ion Channel Fused Arylpyrazole Sulfonamide Skeletons to Enhance Phloem Mobility and Insecticidal Activity as GABA and nACh Receptors Inhibitors.

Ligand-gated ion channels are essential in living organisms, and sulfonamides have antibacterial effects and can be readily coordinated with metal ions with good biological activity. A series of fluorescent ligand-gated ion channel fused arylpyrazole sulfonamide skeletons (APSnM) were synthesized based on a one-pot ultrasound strategy promoted by an inorganic base. APSnM had a high fluorescence quantum yield and a large Stokes shift in ethanol solvent. The ligand bonded ions took on a different color from the ligand and can be used as a probe to detect their own residue on plant surfaces. Their hydrophobic parameters and the fluorescence distribution in Chinese cabbage leaves indicated that APSnM significantly increased the phloem mobility of the plant. The insecticidal activity of APS3Na was higher (LC50 = 7.2423 μg/mL) than that of fipronil (15.2312 μg/mL) against Plutella xylostella, and the mechanism of high insecticidal activity of APS3Na was simulated by molecular docking, which confirmed its strong interactions with the GABA and nACh receptors of Plutella xylostella. Analysis of the crystal structure of these ligand-gated ion channels further confirmed the consistency of their structure and biological activity.

Elucidation of Ion Channel Structure and Mechanism of Action of Amphotericin B, an Antifungal Agent

Elucidation of Ion Channel Structure and Mechanism of Action of Amphotericin B, an Antifungal Agent

Cell-cycle arrest at the G1/S boundary enhances transient voltage-gated ion channel expression in human and insect cells

Heterologous expression of recombinant ion channel subunits in cell lines is often limited by the presence of a low number of channels at the cell surface level. Here, we introduce a combination of two techniques: viral expression using the baculovirus system plus cell-cycle arrest at the G1/S boundary using either thymidine or hydroxyurea. This method achieved a manifold increase in the peak current density of expressed ion channels compared with the classical liposome-mediated transfection methods. The enhanced ionic current was accompanied by an increase in the density of gating charges, confirming that the increased yield of protein and ionic current reflects the functional localization of channels in the plasma membrane. This modified method of viral expression coordinated with the cell cycle arrest will pave the way to better decipher the structure and function of ion channels and their association with ion channelopathies.

Ultrasonic neuromodulation mediated by mechanosensitive ion channels: current and future.

Ultrasound neuromodulation technology is a promising neuromodulation approach, with the advantages of noninvasiveness, high-resolution, deep penetration and good targeting, which aid in circumventing the side effects of drugs and invasive therapeutic interventions. Ultrasound can cause mechanical effects, activate mechanosensitive ion channels and alter neuronal excitability, producing biological effects. The structural determination of mechanosensitive ion channels will greatly contribute to our understanding of the molecular mechanisms underlying mechanosensory transduction. However, the underlying biological mechanism of ultrasonic neuromodulation remains poorly understood. Hence, this review aims to provide an outline of the properties of ultrasound, the structures of specific mechanosensitive ion channels, and their role in ultrasound neuromodulation.

Christos Pliotas.

"A turning point in my career was solving the structure of a mechanosensitive ion channel in the open state … If I could be granted a superpower, it would be the ability to travel in time because I could meet my science heroes and have the opportunity to learn from the best." Find out more about Christos Pliotas in his Introducing … Profile.

Interplay between VSD, pore and membrane lipids in electromechanical coupling in HCN channels.

Hyperpolarized-activated and Cyclic Nucleotide-gated (HCN) channels are the only members of the voltage-gated ion channel superfamily in mammals that open upon hyperpolarization, conferring them pacemaker properties that are instrumental for rhythmic firing of cardiac and neuronal cells. Activation of their voltage-sensor domains (VSD) upon hyperpolarization occurs through a downward movement of the S4 helix bearing the gating charges, which triggers a break in the alpha-helical hydrogen bonding pattern at the level of a conserved Serine residue. Previous structural and molecular simulation studies had however failed to capture pore opening that should be triggered by VSD activation, presumably because of a low VSD/pore electromechanical coupling efficiency and the limited timescales accessible to such techniques. Here, we have used advanced modeling strategies, including enhanced sampling molecular dynamics simulations exploiting comparisons between non-domain swapped voltage-gated ion channel structures trapped in closed and open states to trigger pore gating and characterize electromechanical coupling in HCN1. We propose that the coupling mechanism involves the reorganization of the interfaces between the VSD helices, in particular S4, and the pore-forming helices S5 and S6, subtly shifting the balance between hydrophobic and hydrophilic interactions in a 'domino effect' during activation and gating in this region. Remarkably, our simulations reveal state-dependent occupancy of lipid molecules at this emergent coupling interface, suggesting a key role of lipids in hyperpolarization-dependent gating. Our model provides a rationale for previous observations and a possible mechanism for regulation of HCN channels by the lipidic components of the membrane.