Two TMS K⁺ and water channel (conducts K⁺ (K_D = 8 mM); blocked by Na⁺ (190 mM) (Renart et al., 2006) and tetrabutylammonium (Iwamoto et al., 2006)). Ion permeation occurs by ion-ion contacts in single file fashion through the selectivity filter (Köpfer et al. 2014). A narrow pore lined with four arrays of carbonyl groups is responsible for ion selectivity, whereas a conformational change of the four inner transmembrane helices (TMS2) is involved in gating (Baker et al. 2007). Two gates have been identified; one is located at the inner bundle crossing and is activated by H⁺ while the second gate is in the selectivity filter (Rauh et al. 2017). The C-terminal domain mediates pH modulation (Hirano et al., 2011; Pau et al., 2007). KcsA exhibits a global twisting motion upon gating (Shimizu et al., 2008).  Activity is influenced by the phase of the lipid bilayer (Seeger et al. 2010), and occupancy of nonannular lipid binding sites increases the stability of the tetrameric complex (Triano et al. 2010).  The open conformation of KcsA can disturb the bilayer integrity and catalyze the flipping of phospholipids (Nakao et al. 2014).  This protein is identical to the KcsA orthologue (P0A333) in Streptomyces coelicolor.  The stability of the pre domain in KcsA is stabilized by GCN4 (Yuchi et al. 2008).  The potential role of pore hydration in channel gating has been evaluated (Blasic et al. 2015).  Having multiple K⁺ ions bound simultaneously is required for selective K⁺ conduction, and a reduction in the number of bound K⁺ ions destroys the multi-ion selectivity mechanism utilized by K⁺ channels (Medovoy et al. 2016).  The channel accomodates K⁺ and H₂O molecules alternately in a K⁺-H₂O-K⁺-H₂O  series through the channel (Kratochvil et al. 2016). Insertion of KcsA is spontaneous and directional as the cytosolic part of the protein does not translocate across the membrane barrier. Charged residues, not hydrophobic residues, are crucial for insertion of the unfolded protein into the membrane via electrostatic interactions between membrane and protein.  A two-step mechanism was proposed. An initial electrostatic attraction between membrane and protein represents the first step prior to insertion of hydrophobic residues into the hydrocarbon core of the membrane (Altrichter et al. 2016). Bend, splay, and twist distinguish KcsA gate opening, filter opening, and filter-gate coupling, respectively (Mitchell and Leibler 2017). Details of the water permeability have been presented. Water flow through KcsA is halved by 200 mM K⁺ in the aqueous solution, which indicates an effective K⁺ dissociation constant in that range for a singly occupied channel. (Hoomann et al. 2013). A parameterized MARTINI program can be used to predict the hinging motions of the protein (Li et al. 2019). Activation of KcsA is initiated by proton binding to the pH gate upon an intracellular drop in pH which prompts a conformational switch, leading to a loss of affinity for potassium ions at the selectivity filter and therefore to channel inactivation (Rivera-Torres et al. 2016). An alteration in the conformational equilibrium of the intracellular K⁺-gate is one of the fundamental mechanisms underlying the dysfunctions of K⁺ channels caused by disease-related mutations (Iwahashi et al. 2020). Folding and misfolding of KcsA monomers during assembly and tetramerization has been examined (Song et al. 2021). The flexible C-terminus stabilizes KcsA tetramers at a neutral pH with decreased stabilization at acidic pH (Howarth and McDermott 2022). Under equilibrium conditions, in the absence of a transmembrane voltage, both water and K⁺ occupy the selectivity filter of the KcsA channel in the closed conductive state (Ryan et al. 2023).

Voltage-sensitive Na⁺ channel, Na_V1.7 (Cox et al., 2006). The human orthologue, SCN3A or Nav1.3, when mutated causes cryptogenic pediatric partial epilepsy (Holland et al., 2008; Zaman et al. 2020). Batrachotoxin (BTX) is a steroidal alkaloid neurotoxin that activates NaV channels through interacting with transmembrane domain-I-segment 6 (IS6) of these channels. Ginsenoside inhibits BTX binding (Lee et al. 2008). VGSCs are heterotrimeric complexes consisting of a single pore-forming alpha subunit joined by two beta subunits, a noncovalently linked beta1 or beta3 and a covalently linked beta2 or beta4 subunit (Hull and Isom 2017).  The binding mode and functional components of the analgesic-antitumour peptide from Buthus martensii Karsch to human voltage-gated sodium channel 1.7 have been characterized (Zhao et al. 2019). Dvorak et al. 2021 developed allosteric modulators of ion channels by targeting their PPI interfaces, particularly in the C-terminal domain of the Nav, with auxiliary proteins. Fenestrations are key functional regions of Nav that modulate drug binding, lipid binding, and influence gating behaviors (Gamal El-Din and Lenaeus 2022). Compartment-specific localizations and trafficking mechanisms for VGSCs are regulated separately to modulate membrane excitability in the brain (Liu et al. 2022). Naview is a library for drawing and annotating voltage-gated sodium channel membrane diagrams (Afonso et al. 2022). Deltamethrin (DLT) is a type-II pyrethroid ester insecticide used in agricultural and domestic applications as well as in public health. Exposure to DLT produced a differential and dose-dependent stimulation of peak Na+ currents, Conversely, tefluthrin (Tef), a type-I pyrethroid insecticide, accentuates I(Na) with a slowing in inactivation time course of the current (Lin et al. 2022). MicroRNA-335-5p suppresses voltage-gated sodium channel expression and may be a target for seizure control (Heiland et al. 2023).  Voltage-gated sodium channels are enhancing factors in the metastasis of metastatic prostate cancer cells (Yildirim-Kahriman 2023). Decreasing microtubule detyrosination modulates Nav1.5 subcellular distribution and restores sodium current in Mdx cardiomyocytes (Nasilli et al. 2024).

Type 2 Na⁺ channel, SCN2A or NaV1.2, of 2,005 aas and 24 TMSs.  Mutations give rise to epileptic encephalophathy, Ohtahara syndrome (Nakamura et al. 2013). They may also give rise to autism (ASD) (Tavassoli et al. 2014). This protein is orthologous to the rat Na⁺ channel, TC# 1.A.1.10.1 and very similar to the type 1 Na⁺ channel (1.A.1.10.7). Na_V1.2 has a single pore-forming alpha-subunit and two transmembrane beta-subunits. Expressed primarily in the brain, Na_V1.2 is critical for initiation and propagation of action potentials. Milliseconds after the pore opens, sodium influx is terminated by inactivation processes mediated by regulatory proteins including calmodulin (CaM). Both calcium-free (apo) CaM and calcium-saturated CaM bind tightly to an IQ motif in the C-terminal tail of the alpha-subunit. Thermodynamic studies and solution structure (2KXW) of a C-domain fragment of apo 13C,15N- CaM (CaMC) bound to an unlabeled peptide with the sequence of the rat NaV1.2 IQ motif showed that apo CaMC (a) was necessary and sufficient for binding, and (b) bound more favorably than calcium-saturated CaMC. CaMN  apparently does not influence apo CaM binding to NaV1.2IQp (Mahling et al. 2017). The phenotypic spectrum of SCN2A-related epilepsy is broad, ranging from benign epilepsy in neonate and infancy to severe epileptic encephalopathy. Oxcarbazepine and valproate are the most effective drugs in epilepsy patients with SCN2A variants. Sodium channel blockers often worsen seizures in patients with seizure onset beyond 1 year of age. Abnormal brain MRI findings and de novo variations are often related to poor prognosis. Most SCN2A variants located in transmembrane regions were related to patients with developmental delay (Zeng et al. 2022). The beta4-subunit and PRRT2 form a push-pull system that finely tunes the membrane expression and function of Na_V channels and the intrinsic neuronal excitability (Valente et al. 2022). Icariin can be used to treat epilepsy by inhibiting neuroinflammation via promoting microglial polarization to the M2 phenotype (Wang et al. 2023).

Sodium channel Nav1.4-beta complex of 1820 and 209 aas, respectively.  Voltage-gated sodium (Nav) channels initiate and propagate action potentials. Yan et al. 2017 presented the cryo-EM structure of EeNav1.4, the Nav channel from electric eel, in complex with the beta1 subunit at 4.0 Å resolution. The immunoglobulin domain of beta1 docks onto the extracellular L5I and L6IV loops of EeNav1.4 via extensive polar interactions, and the single transmembrane helix interacts with the third voltage-sensing domain (VSDIII). The VSDs exhibit ""up"" conformations, while the intracellular gate of the pore domain is kept open by a digitonin-like molecule. Structural comparison with closed NavPaS shows that the outward transfer of gating charges is coupled to the iris-like pore domain dilation through intricate force transmissions involving multiple channel segments. The IFM fast inactivation motif on the III-IV linker is plugged into the corner enclosed by the outer S4-S5 and inner S6 segments in repeats III and IV, suggesting a potential allosteric blocking mechanism for fast inactivation (Yan et al. 2017). The PDB# for the complex is 5XSY, and that for the two subunits are 5XSY_A and 5XSY_B. Domain 4 TMS 6 of Nav1.4 plays a key role in channel gating regulation, and is targeted by the neurotoxin, veratridine (VTD) (Niitsu et al. 2018).

Voltage-gated sodium channel of 1836 aas and 24 TMSs, PaFPC1. The 3-d structure has been determined (Shen et al. 2017).  It mediates the voltage-dependent sodium ion permeability in excitable membranes.

Ca²⁺-regulated heart Na⁺ channel, Na_v1.5, SCN5A or INa channel of 2016 aas. The COOH terminus functions in the control of channel inactivation and in pathologies caused by inherited mutations that disrupt it (Glaaser et al., 2006); regulated by ProTx-II Toxin (Smith et al. 2007), telethonin, the titin cap protein (167aas; secreted protein; O15273) (Mazzone et al., 2008), and the Mog1 protein, a central component of the channel complex (Wu et al., 2008). Na_v1.5, the principal Na⁺ channel in the heart, possesses an ankyrin binding site, and direct interaction with ankyrin-G is required for the expression of Na_v1.5 at the cardiomyocyte cell surface (Bennett and Healy, 2008; Lowe et al., 2008). Mutations cause type 3 long QT syndrome and type 1 Brugada syndrome, two distinct heritable arrhythmia syndromes (Mazzone et al., 2008; Kapplinger et al. 2010; Wang et al. 2015). SCN5A mutations causing arrhythmic dilated cardiomyopathy, commonly localized to the voltage-sensing mechanism, and giving rise to gating pore currents (currents that go through the voltage sensor) have been identified (McNair et al., 2011; Moreau et al., 2014).  Patients with Brugada syndrome are prone to develop ventricular tachyarrhythmias that may lead to syncope, cardiac arrest or sudden cardiac death (Sheikh and Ranjan 2014) and (Kapplinger et al. 2015). Mutations causing disease have been identified (Qureshi et al. 2015). These give rise to arrhythias and cardiomyopathies (Moreau et al. 2015).  Mutations that cause relative resistance to slow inactivation have been identified (Chancey et al. 2007).  Green tea catechins are potential anti-arrhythmics because of the significant effect of Epigallocatechin-3-Gallate (E3G) on  cardiac sodium channelopathies that display a hyperexcitability phenotype (Boukhabza et al. 2016). A mutatioin, R367G, causes the familial cardiac conductioin disease (Yu et al. 2017). The C-terminal domain of calmodulin (CaM) binds to an IQ motif in the C-terminal tail of the alpha-subunit of all NaV isoforms, and contributes to calcium-dependent pore-gating in some (Isbell et al. 2018). Ventricular fibrillation in patients with Brugada syndrome (BrS) is often initiated by premature ventricular contractions, and the presence of SCN5A mutations increases the risk upon exposure to sodium channel blockers in patients with or without baseline type-1 ECG (Amin et al. 2018). A mutation (R367G) is associated with familial cardiac conduction disease (Yu et al. 2017). Among ranolazine, flecainide, and mexiletine, only mexiletine restored inactivation kinetics of the currents of the mutant protein, A1656D (Kim et al. 2019).  Epigallocatechin-3-gallate (EGCG) is protective against cardiovascular disorders due in part to its action on multiple molecular pathways and transmembrane proteins, including the cardiac Nav1.5 channels (Amarouch et al. 2020). An SCN1B variant affects both cardiac-type (NaV1.5) and brain-type (NaV1.1) sodium currents and contributes to complex concomitant brain and cardiac disorders (Martinez-Moreno et al. 2020). Mice null for Scn1b, which encodes NaV beta1 and beta1b subunits, have defects in neuronal development and excitability, spontaneous generalized seizures, cardiac arrhythmias, and early mortality (Martinez-Moreno et al., 2020; Martinez-Moreno et al. 2020). The structural basis of cytoplasmic NaV1.5 and NaV1.4 regulation has been reviewed (Nathan et al. 2021). Fibroblast growth factor 21 ameliorates NaV1.5 and Kir2.1 channel dysregulation in human AC16 cardiomyocytes (Li et al. 2021). The interaction of Nav1.5 with MOG1 (RANGRF), a Ran guanine nucleotide release factor and chaparone, provides a possible molecular mechanism for Brugada syndrome (Xiong et al. 2021). Arrhythmic phenotypes are a defining feature of dilated cardiomyopathy-associated SCN5A variants (Peters et al. 2021). A SCN5A genetic variant, Y739D, is associated with Brugada syndrome (Zaytseva et al. 2022). Melatonin treatment causes an increase of conduction via enhancement of sodium channel protein expression and increases of sodium current in the ventricular myocytes (Durkina et al. 2022). Quantification of Nav1.5 expression has been published (Adams et al. 2022). Cardiac sodium channel complexes play a role in arrhythmia, and the structural and functional roles of the beta1 and beta3 subunits have been determined (Salvage et al. 2022). Brugada Syndrome (BrS) treatment is electrocardiography with ST-segment elevation in the direct precordial derivations. The clinical presentation of the disease is highly variable. Patients can remain completely asymptomatic, but they can also develop episodes of syncope, atrial fibrillation (AF), sinus node dysfunction (SNF), conduction disorders, asystole, and ventricular fibrillation (VF). This disease is caused by mutations in the genes responsible for the potential action of cardiac cells. The most commonly involved gene is SCN5A, which controls the structure and function of the heart's sodium channel (Brugada 2023).

The skeletal muscle Na⁺ channel, Na_V1.4 of 1836 aas and 24 TMSs. Mutations in charged residues in the S4 segment cause hypokalemic periodic paralysis (HypoPP)) due to sustained sarcolemmal depolarization (Struyk and Cannon 2007; Sokolov et al., 2007; Groome et al. 2014). Also causes myotonia; regulated by calmodulin which binds to the C-terminus of Na_v1.4 (Biswas et al., 2008). Na_V1.4 gating pores are permeable to guanidine as well as Na⁺ and H⁺ (Sokolov et al., 2010). The R669H mutation allows transmembrane permeation of protons, but not larger cations, similar to the conductance displayed by histidine substitution at Shaker K⁺ channel S4 sites (Struyk and Cannon 2007).  The mechanism of inactivation involves transient interactions between intracellular domains resulting in direct pore occlusion by the IFM motif and concomitant extracellular interactions with the beta1 subunit (Sánchez-Solano et al. 2016). Potassium-sensitive hypokalaemic and normokalaemic periodic paralysis are inherited skeletal muscle diseases in humans, characterized by episodes of flaccid muscle weakness. They are caused by single mutations in positively charged residues ('gating charges') in the S4 transmembrane segment of the voltage sensor of the voltage-gated sodium channel Nav1.4 or the calcium channel Cav1.1. Mutations of the outermost gating charges (R1 and R2) cause hypokalaemic periodic paralysis by creating a pathogenic gating pore in the voltage sensor through which cations leak in the resting state. Mutations of the third gating charge (R3) cause normokalaemic periodic paralysis owing to cation leak in both activated and inactivated states (Jiang et al. 2018). The neurotoxic cone snail peptide μ-GIIIA specifically blocks skeletal muscle voltage-gated sodium (Na_V1.4) channels (Leipold et al. 2017). the cryo-electron microscopy structure of the human Na_v1.4-β1 complex at 3.2-Å resolution. Accurate model building was made for the pore domain, the voltage-sensing domains, and the β1 subunit (Pan et al. 2018) provided insight into the molecular basis for Na⁺ permeation and kinetic asymmetry of the four repeats. Structural analysis of reported functional residues and disease mutations corroborates an allosteric blocking mechanism for fast inactivation of Na_v channels. the S4-S5L of the DI, DII and DIII domains allosterically modulate the activation gate and stabilize its open state (Malak et al. 2020). The structural basis of cytoplasmic NaV1.5 and NaV1.4 regulation has been reviewed (Nathan et al. 2021). Mutations in SCN4A give rise to a variety of pathological conditions (Sun et al. 2021). Hypokalemic periodic paralysis (HypoPP) is a rare autosomal dominant disease caused by mutations in either calcium or sodium transmembrane voltage-gated ion channels in the ER of skeletal muscle (Calise et al. 2023).

Voltage-sensitive Na⁺ channel, type 9, α-subunit, Nav1.7 or SCN9A (orthologous to 1.A.1.10.1). Loss of function, resulting from point mutations, results in a channelopathy called Congenital Insensitivity to Pain (CIP) (He et al. 2018), that causes the congenital inability to experience pain (Cregg et al., 2010; Kleopa, 2011). An S241T mutation causes inherited erythromelalgia IEM; erythermalgia, an autosomal dominant neuropathy characterized by burning pain in the extremities in response to mild warmth (due to altered gating) (Lampert et al., 2006; Drenth and Waxman, 2007). Gain-of-function mutations in the Na(v)1.7 channel lead to DRG neuron hyperexcitability associated with severe pain, whereas loss of the Na(v)1.7 channel in patients leads to indifference to pain (Dib-Hajj et al., 2007). Blocked by 1-benzazepin-2-one (Kd = 1.6 nM) (Williams et al., 2007). Mutations in the Nav1.7 Na channel α-subunit give rise to familial pain syndromes called chronic non-paoxysmal neuropathic pain (Catterall et al., 2008; Fischer and Waxman, 2010; Dabby et al. 2011 ). It interacts with the sodium channel beta3 (Scn3b), rather than the beta1 subunit, as well as the collapsing-response mediator protein (Crmp2) through which the analgesic drug lacosamide regulates Nav1.7 current (Kanellopoulos et al. 2018). The R1488 variant is totally inactive (He et al. 2018). Nav1.7 is inhibited by knottins (see TC# 8.B.19.2) (Agwa et al. 2018). Nav1.7 interacts with the following proteins: syn3b (TC# 8.a.17.1.2; the β3 subunit), Crmp2, Syt2 (Q8N9I0) and Tmed10 (P49755), and it also regulates opioid receptor efficacy (Kanellopoulos et al. 2018). Mutations in TRPA1 and Nav1.7 to  insensitivity to pain-promoting algogens such as capsaicin, acid, and allyl isothiocyanate (AITC), have been documented (Eigenbrod et al. 2019). Na_v1.7 is associated with endometrial cancer (Liu et al. 2019) and fever-associated seizures or epilepsy (FASE) (Ding et al. 2019). Nav1.7 and Nav1.8 peripheral nerve sodium channels are modulated by protein kinases A and C (Vijayaragavan et al. 2004). Sodium channel Na_V1.7 and potassium channel K_V7.2 promote and oppose excitability in nociceptors, respectively. Inflammation differentially controls transport of depolarizing Nav versus hyperpolarizing Kv channels to drive rat nociceptor activity (Higerd-Rusli et al. 2023). The structural basis for severe pain, caused by mutations in the S4-S5 linkers of voltage-gated sodium channel Na_V1.7, have been revealed (Wisedchaisri et al. 2023).

Neuronal nonselective cation channel, NALCN (forms background leak conductance and controls neuronal excitability; Lu et al., 2007).  It is also found in the pancreatic β-cell (Swayne et al. 2010). NALCN serves as a variable sensor that responds to calcium or sodium ion flux, depending on whether the total cellular current density is generated more from calcium-selective or sodium-selective channels (Senatore and Spafford 2013). It functions in a complex with Unc80 (3258 aas; Q8N2C7) and Unc79 (2635 aas; Q9P2D8) (Bramswig et al. 2018). Heterozygous de novo NALCN missense variants in the S5/S6 pore-forming segments lead to congenital contractures of the limbs and face, hypotonia, and developmental delay (Bramswig et al. 2018). Overexpression of the NALCN gene ablates allyl isothiocyanate-promoting pain reception by nociceptors (Eigenbrod et al. 2019).

The phosphoinositide (PI(3,5)P₂)-activated Na⁺ two pore channel-2, TPC2, in endosomes and lysosomes (Wang et al. 2012). Previously thought, incorectly, according to Wang et al. 2012, to be a nicotinic acid adenine dinucleotide phosphate (NAADP)-dependent two pore Ca²⁺ channel.  TPC2, like TPC1, has a 12 TMS topology (two channel units) (Hooper et al., 2011). The two domains of human TPCs can insert into the membrane independently (Churamani et al., 2012).  Cang et al. (2013), showed that TPC1 and TPC2 together form an ATP-sensitive two-pore Na⁺ channel that senses the metabolic state of the cell.  The channel complex detects nutrient status, becomes constitutively open upon nutrient removal, and controls the lysosome's membrane potential, pH stability, and amino acid homeostasis.  Essential for Ebola virus (EBOV) host entry. Several inhibitors of TPC2 that act in the nM (tetrandrine) or μM (verapamil; Ned19) range block channel activity, prevent Ebola Virus from escaping cell vesicles and may be used to treat the disease (Sakurai et al. 2015).  TPC2 may transport both Na⁺ and Ca²⁺ (Sakurai et al. 2015). Lipid-gated monovalent ion fluxes, mediated by TPC1 and TPC2 in mice, regulate endocytic traffic and support immune surveillance. This is in part achieved by catalyzing Na⁺ export from visicles derived from the plasma membrane by phagocytosis or pinocytosis, causing contraction and allowing the maintenance of a uniform cell volume (Freeman et al. 2020). This system is important for melanocyte function (Wiriyasermkul et al. 2020).  Convergent activation of two-pore channels mediated by the NAADP-binding proteins JPT2 and LSM12 has been reported (Gunaratne et al. 2023).  The lysosomal two-pore channels 2 (TPC2) and IP3 receptors (IP3Rs) located in the endoplasmic reticulum may be coupled (Yuan et al. 2024).

Muscle plasmalemma, voltage-gated, L-type dihydropyridine receptor Ca²⁺ channel, α-1 subunit (DHPR) (Ba²⁺ > Ca²⁺), Cav1.1, CACNA1S,CACH1 CDCN1, CACNL1A3 of 1873 aas in the human orthologue.  Distinc voltage sensor domains control voltage sensitivity and kinetics of current activation (Tuluc et al. 2016). Rapid changes in the transmembrane potential are detected by the voltage-gated Ca²⁺ channel, dihydropyridine receptor (DHPR), embedded in the sarcolemma. DHPR transmits the contractile signal to another Ca²⁺ channel, the ryanodine receptor (RyR1), embedded in the membrane of the sarcoplasmic reticulum (SR), which releases a large amounts of Ca²⁺ from the SR that initiate muscle contraction (Shishmarev 2020).

The phosphoinositide (PI(3,5)P₂)-activated Na⁺ two pore channel-1, TPC1 of endosomes and lysosomes (Wang et al. 2012). Previously thought, incorectly, according to Wang et al. (2012), to be an NAADP-activated two pore voltage-dependent calcium channel protein.  However, Cang et al. (2013), showed that TPC1 and TPC2 (TC# 1.A.1.11.19) together form an ATP-sensitive two-pore Na⁺ channel that senses the metabolic state of the cell.  The channel complex detects nutrient status, becomes constitutively open upon nutrient removal, and controls the lysosome's membrane potential, pH stability, and amino acid homeostasis.  May be regulated by the HCLS-associated X-1 (HAX-1) protein (Lam et al. 2013). The cryoEM 3-D structure has been ellucidated (She et al. 2018). This voltage-dependent, phosphatidylinositol 3,5-bisphosphate (PtdIns(3,5)P₂)-activated Na⁺ channel was solved in both the apo closed state and ligand-bound open state. The channel has a coin-slot-shaped ion pathway in the filter that defines the selectivity of mammalian TPCs. Only the voltage-sensing domain from the second 6-TMS domain confers voltage dependence while endolysosome-specific PtdIns(3,5)P₂ binds to the first 6-TMS domain and activates the channel under conditions of depolarizing membrane potential. Structural comparisons between the apo and PtdIns(3,5)P2-bound structures show the interplay between voltage and ligand activation. These MmTPC1 structures reveal lipid binding and regulation in a 6-TMS voltage-gated channel (She et al. 2018).

Two pore Ca²⁺ > Na⁺, Li⁺ or K⁺ (non-selective for these three monovalen caions) channel protein of 733 aas and 12 TMSs, TPC1 (Guo et al. 2017). The crystal structure of this vacuolar two-pore channel, a homodimer, has been solved (Guo et al. 2015) (Kintzer and Stroud 2016). Activation requires both voltage and cytosolic Ca²⁺. Ca²⁺ binding to the cytosolic EF-hand domain triggers conformational changes coupled to the pair of pore-lining inner helices from the first 6-TMS domains, whereas membrane potential only activates the second voltage-sensing domain, the conformational changes of which are coupled to the pair of inner helices from the second 6-TMS domains. Luminal Ca²⁺ or Ba²⁺ modulates voltage activation by stabilizing the second voltage-sensing domain in the resting state and shift voltage activation towards more positive potentials. The basis for understanding ion permeation, channel activation, the location of voltage-sensing domains and regulatory ion-binding sites is partially explained by the 3-d structure (Kintzer and Stroud 2016). Only the second Shaker domain senses voltage (Jaślan et al. 2016). It has a selectivity filter that is passable by hydrated divalent cations (Demidchik et al. 2018). Dickinson et al. 2022 determined structures at different stages along its activation coordinate. These structures of activation intermediates, when compared with the resting-state structure, portray a mechanism in which the voltage-sensing domain undergoes dilation and in-membrane plane rotation about the gating charge-bearing helix, followed by charge translocation across the charge transfer seal. These structures, in concert with patch-clamp electrophysiology, showed that residues in the pore mouth sense inhibitory Ca²⁺ and are allosterically coupled to the voltage sensor. These conformational changes provide insight into the mechanism of voltage-sensor domain activation in which activation occurs vectorially over a series of elementary steps (Dickinson et al. 2022).  Inhibition of the Akt/PKB kinase increases Nav1.6-mediated currents and neuronal excitability in CA1 hippocampal pyramidal neurons (Marosi et al. 2022).

Voltage-dependent P/Q-type Ca²⁺ channel subunit α1A, CACNA1A (CACH4; CACN3; CACNL1A4) of 2,505 aas. The CACNA1A gene is widely expressed throughout the CNS. The encoding protein is 90% identical to 1.A.1.11.8.  Associated with four neurological phenotypes: familial and sporadic hemiplegic migraine type 1 (FHM1, SHM1), episodic ataxia type 2 (EA2), spinocerebellar ataxia type 6 (SCA6) and epileptic encephalopathy with nerve atrophy (Reinson et al. 2016). A gain of function mutation gave symptoms of congenital ataxia, abnormal eye movements and developmental delay with severe attacks of hemiplegic migraine (García Segarra et al. 2014). Mutations can cause F/SHM with high penitrance (Prontera et al. 2018). CACNA1A variants lead to a wide spectrum of neurological disorders including epileptic or non-epileptic paroxysmal events, cerebellar ataxia, and developmental delay. The variants are either gain of function GOF) or loss of function (LOF) mutations (Zhang et al. 2020). CACNA1A pathogenic variants have been linked to several neurological disorders including severe early onset developmental encephalopathies and cerebellar atrophy. Y1384 variants exhibit differential splice variant-specific effects on recovery from inactivation (Gandini et al. 2021). Patients with CACNA1A mutational variants located in the transmembrane region may be at high risk of status epilepticus (Niu et al. 2022). Patients with ataxia in the absence of epilepsy can be caused by a CACNA1A mutationand respond to pyridoxine (Du et al. 2017). lamotrigine can be used to treat patients with refractory epilepsy due to calcium channel mutations (Hu et al. 2022; De Romanis and Sopranzi 2018). Eupatilin depresses glutamate exocytosis from cerebrocortical synaptosomes by decreasing P/Q-type Ca²⁺ channels and synapsin I phosphorylation and alleviates glutamate excitotoxicity caused by kainic acid by preventing glutamatergic alterations in the mamalian cortex. Thus, eupatilin is a potential therapeutic agent in the treatment of brain impairment associated with glutamate excitotoxicity (Lu et al. 2022).

Voltage-sensitive calcium channel (VSCC), CA_V1.3, encoded by the CACNA1D gene, of 2161 aas and 24 TMSs (Singh et al. 2008). Ca_V1.3-R990H channels conduct omega-currents at hyperpolarizing potentials, but not upon membrane depolarization compared with wild-type channels (Monteleone et al. 2017). A CACNA1D de novo mutation causes a severe neurodevelopmental disorder (Hofer et al. 2020).

Pore-forming, alpha-1S subunit of the voltage-gated calcium channel, of 1873 aas and 24 TMSs, Cav1.1; CACNA1S; CACN1; CACH1; CACNL1A3, that gives rise to L-type calcium currents in skeletal muscle. Calcium channels containing the alpha-1S subunit play an important role in excitation-contraction coupling in skeletal muscle via their interaction with RYR1, which triggers Ca²⁺ release from the sarcplasmic reticulum and ultimately results in muscle contraction. Long-lasting (L-type) calcium channels belong to the 'high-voltage activated' (HVA) group (Jiang et al. 2018). The 3-d structure of a bacterial homologue has been solved (Jiang et al. 2018). Mutations in arginly residues in the TMS4 voltage lead to increased leak currently that may be responsible for hypokalaemic periodic paralysis (Kubota et al. 2020). Mutations in the voltage sensor domain of CaV1.1, the alpha1S subunit of the L-type calcium channel in skeletal muscle cause hypokalemic periodic paralysis (HypoPP), and these mutations give rise to gating pore currents (Wu et al. 2021). The voltage-gated T-type calcium channel is modulated by kinases and phosphatases (Sharma et al. 2023). Advances in Ca_V1.1 gating, dealing with permeation and voltage-sensing mechanisms, have been reviewed (Bibollet et al. 2023). It is possible to prevent calcium leak associated with short-coupled polymorphic ventricular tachycardia in patient-derived cardiomyocytes (Sleiman et al. 2023). Far-infrared ameliorates Pb-induced renal toxicity via voltage-gated calcium channel-mediated calcium influx (Ko et al. 2023). Verapamil mitigates chloride and calcium bi-channelopathy in a myotonic dystrophy mouse model (Cisco et al. 2024).

The voltage-dependent L-type Ca²⁺ channel α-subunit-1C (L-type Ca_v1.2), CACNA1C (CACH2, CACN2, CACNL1A1, CCHL1A1) of 2221 aa. Mutations cause Timothy's syndrome, a disorder associated with autism (Splawski et al., 2006). The C-terminus of Ca_v1.2 encodes a transcription factor (Gomez-Ospina et al., 2006). Ca_v1.2 associates with the α-2, δ-1, β and γ subunits (Yang et al., 2011). The CRAC channel activator STIM1 binds and inhibits L-type voltage-gated calcium channel, Ca_v1.2 (Park et al., 2010).  This channel appears to function as the molecular switch for synaptic transmission (Atlas 2013). Intramembrane signalling occurs with syntaxin 1A for catecholamine release in chromaffin cells (Bachnoff et al. 2013).  miR-153 intron RNA is a negative regulator of both insulin and dopamine secretion through its effect on Cacna1c expression, suggesting that IA-2beta and miR-153 have opposite functional effects on the secretory pathway (Xu et al. 2015).   Co-localizes with Syntaxin-1A in nano clusters at the plasma membrane (Sajman et al. 2017). It is a high voltage-activated Ca²⁺ channel in contrast to Ca_v3.3 which is a low voltage-activated Ca²⁺ channel (Sanchez-Sandoval et al. 2018). Nifedipine blocks and potentiates this and other L-type VIC Ca²⁺ channels (Wang et al. 2018).  Cav1.2 is upregulated when STIM1 is deficient (Pascual-Caro et al. 2018). CaV1.2 regulates chondrogenesis during limb development (Atsuta et al. 2019). CACNA1C may be a prognostic predictor of survival in ovarian cancer (Chang and Dong 2021). Kinase and phosphatase modulation of T-type Ca²⁺ channel (TTCC) isoforms Cav3.1, Cav3.2, and Cav3.3, are mostly described for roles unrelated to cellular excitability (Sharma et al. 2023), and potential modulations that are yet to be explored are also discussed. Palmitoylation of the pore-forming subunit of Ca(v)1.2 controls channel voltage sensitivity and calcium transients in cardiac myocytes (Kuo et al. 2023).  A novel binding site between the voltage-dependent calcium channel Ca_V1.2 subunit and the Ca_Vβ2 subunit has been discovered using a new analysis method for protein-protein interactions (Murakami et al. 2023). CACNA1C is one of the top risk genes for schizophrenia; A novel 17-variant block across introns 36-45 of CACNA1C was significantly associated with schizophrenia; a novel 17-variant block across introns 36-45 of CACNA1C was responsible (Guo et al. 2023). A novel binding site has been found between the voltage-dependent calcium channel Ca_V1.2 subunit and Ca_Vβ2 subunit (Murakami et al. 2023).

6TMS K⁺ channel (Munsey et al. 2002; Kuo et al., 2003).  The kch gene, the only known potassium channel gene in E. coli, has the property to express both full-length Kch and its cytosolic domain (RCK) due to a methionine at position 240. The RCK domains form an octameric ring structure and regulate the gating of the potassium channels after having bound certain ligands. Several different gating ring structures have been reported for the soluble RCK domains.  The octameric structure of Kch may be composed of two tetrameric full-length proteins through RCK interaction (Kuang et al. 2013).  The RCK domains face the solution, and an RCK octameric gating ring arrangement does not form under certain conditions (Kuang et al. 2015).

2 TMS ( P-loop) Ca²⁺-gated K⁺ channel, MthK (see Jiang et al., 2002 for the crystal structure, and Parfenova et al., 2006 for mutations affecting open probability). For the studies of ion permeation and Ca²⁺ blockage, see Derebe et al., 2011. (structures: 3LDD_A and 2OGU_A.). Voltage-dependent K⁺ channels including MthK which lacks a canonical voltage sensor can undergo a gating process known as C-type inactivation, which involves entry into a nonconducting state through conformational changes near the channel's selectivity filter (Thomson and Rothberg, 2010). C-type inactivation may involve movements of transmembrane voltage sensor domains. In the absence of Ca²⁺, a single structure in a closed state was observed by cryoEM that was highly flexible with large rocking motions of the gating ring and bending of pore-lining helices (Fan et al. 2020). In Ca²⁺-bound conditions, several open-inactivated conformations were present with the different channel conformations being distinguished by rocking of the gating rings with respect to the transmembrane region. In all conformations displaying open channel pores, the N-terminus of one subunit of the channel tetramer sticks into the pore and plugs it. Deletion of this N terminus led to non-inactivating channels with structures of open states without a pore plug, indicating that this N-terminal peptide is responsible for a ball-and-chain inactivation mechanism (Fan et al. 2020). Lipid-protein interactions influence the conformational equilibrium between two states of the channel that differ according to whether a TMS has a kink. Two key residues in the kink region mediate crosstalk between two gates at the selectivity filter and the central cavity, respectively. Opening of one gate eventually leads to closure of the other (Gu and de Groot 2020). Activation of MthK is exquisitely regulated by temperature (Jiang et al. 2020).

Voltage-activated, Ca²⁺  channel blocker-inhibited, Na⁺ channel, NaChBac (Ren et al., 2001; Zhao et al., 2004; Nurani et al, 2008; Charalambous and Wallace, 2011). Arginine residues in the S4 segment play a role in voltage-sensing (Chahine et al. 2004). Transmembrane and extramembrane regions contribute to thermal stability (Powl et al., 2012). Deprotonation of arginines in S4 is involved in NaChBac gating (Paldi, 2012). Hinge-bending motions in the pore domain of NaChBac have been reported (Barber et al., 2012).  The C-terminal coiled-coli stabilizes subunit interactions (Mio et al. 2010).  Within the 4 TMS voltage sensor, coupling between residues in S1 and S4 determines its resting conformation (Paldi and Gurevitz 2010). The conserved asparagine was changed to aspartate, N225D, and this substitution shifted the voltage-dependence of inactivation by 25 mV to more hyperpolarized potentials. The mutant also displays greater thermostability (O'Reilly et al. 2017).  Possibly, the side-chain amido group of asn₂₂₅ forms one or more hydrogen bonds with different channel elements, and  these interactions are important for normal channel function. The T1-tetramerization domain of Kv1.2 (TC# 1.A.1.2.10) rescues expression and preserves the function of a truncated form of the NaChBac sodium channel (D'Avanzo et al. 2022). The structure of NaChBac embedded in liposomes has been solved by cryo electron tomography (Chang et al. 2023). The small channel has most of its residues embedded in the membrane, and these are flexible, determining the channel dimensions.

Voltage-gated Na⁺ channel, Na_vCh or Na_vAb. The 3d-structure is known (3ROW; Payandeh et al., 2011; 4MW3A-D; 4+ selectivity through partial dehydration of Na⁺ via its direct interaction with conserved glutamate side chains). The pore is preferentially occupied by two ions, which can switch between different configurations by crossing low free-energy barriers (Furini and Domene, 2012). Jiang et al. 2018 presented high-resolution structures of NavAb with the analogous gating-charge mutations that have similar functional effects as in the human channels that cause hypokalaemic and normokalaemic periodic paralysis.  Wisedchaisri et al. 2019 presented a cryo-EM structure of the resting state and a complete voltage-dependent gating mechanism via the voltage sensor (VS). The S4 segment of the VS is drawn intracellularly, with three gating charges passing through the transmembrane electric field. This movement forms an elbow connecting S4 to the S4-S5 linker, tightens the collar around the S6 activation gate, and prevents its opening. This structure supports the classical "sliding helix" mechanism of voltage sensing and provides a complete gating mechanism for voltage sensor function, pore opening, and activation-gate closure based on high-resolution structures of a single sodium channel protein (Wisedchaisri et al. 2019).

(see also TC#s 1.A.1.10.4 and 1.A.1.11.32).

The transport reaction catalyzed by Na_vCh is:

Na⁺ (in) ⇌ Na⁺ (out)

Bacterial voltage-gated sodium channel, Nav.  3-d crystal structures of vaious conformations are known (4P_3A A-D; 4PA7_A-D; 4P9P_A-D. etc.)  (McCusker et al. 2012).  It has its internal cavity accessible to the cytoplasmic surface as a result of a bend/rotation about a central residue in the carboxy-terminal TMS that opens the gate to allow entry of hydrated sodium ions. The molecular dynamics of ion transport through the open conformation has been analyzed (Ulmschneider et al. 2013).  The C-terminal four helix coiled coil bundle domain couples inactivation with channel opening, depedent on the negatively charged linker region (Bagnéris et al. 2013). A NaVSp1-specific S4-S5 linker peptide induced both an increase in NaVSp1 current density and a negative shift in the activation curve, consistent with the S4-S5 linker stabilizing the open state (Malak et al. 2020).

Tetrameric 6 TMS subunit Na⁺ channel protein, Na_V.  Two low resolution cyroEM structures revealed two conformations reconstituted in lipid bilayers (Tsai et al. 2013). Despite a voltage sensor arrangement identical with that in the activated form, Tsai et al. 2013 observed two distinct pore domain structures: a prominent form with a relatively open inner gate, and a closed inner-gate conformation similar to the first prokaryotic Na_v structure. Structural differences, together with mutational and electrophysiological analyses, indicated that widening of the inner gate was dependent on interactions among the S4-S5 linker, the N-terminal part of S5 and its adjoining part in S6, and on interhelical repulsion by a negatively charged C-terminal region subsequent to S6 (Tsai et al. 2013).

6 TMS voltage-gated K⁺ channel, KCNQ2 or Kv7.2.  Mutations cause benign familial neonatal convulsions (BNFC; epilepsy; Maljevic et al. 2016; Soldovieri et al. 2019).  It forms homotetramers or heterotetramers with KCNQ3/Kv7.3) (Soldovieri et al., 2006; Uehara et al., 2008)). Like all other Kv7.2 channels, it is activated by phosphatidyl inositol-4,5-bisphosphate and hence can be regulated by various neurotransmitters and hormones (Telezhkin et al. 2013).  Gating pore currents that go through the gating pores in TMSs1-4 (the voltage sensor) may give rise to peripheral nerve hyperexcitability (Moreau et al. 2014). Retigabine and ICA73, two anti-epileptic drugs, act via distinct mechanisms due to interactions with specific residues that underlie subtype specificity of KCNQ channel openers (Wang et al. 2016). A  tight spatial and functional relationship between the DAT/GLT-1 transporters and the Kv7.2/7.3 potassium channel immediately readjusts the membrane potential of the neuron, probably to limit the neurotransmitter-mediated neuronal depolarization (Bartolomé-Martín et al. 2019). E-2-dodecenal from cilantro (Coriandrum sativum) is a potent activator and anticonvulsant that binds with an affinity of 60 nM to TMS5 in several KCNQ channels including KCNQ2 and 3 (Manville and Abbott 2019). The activities of Kv7 channels are modulated by polyunsaturated fatty acids (Larsson et al. 2020). Anticancer effects of FS48 from salivary glands of Xenopsylla cheopis via its blockage of voltage-gated K⁺ channels has been demonstrated (Xiong et al. 2023). The drug, ezogabine restoresnormal activity ,decreasing depressive symptoms in major depressive disorder patients (Costi et al. 2021). Both L- and D-isomers of S-nitrosocysteine (CSNO) can bind to the intracellular domain of voltage-gated potassium channels in vitro. CSNO binding inhibits these channels in the carotid body, leading to increased minute ventilation in vivo (Krasinkiewicz et al. 2023).

6 TMS voltage-gated K⁺ channel, KCNQ3 or K_v7.3.  Mutations cause benign familial neonatal convulsions (BNFC; epilepsy; Maljevic et al. 2016). Forms homotetramers or heterotetramers with KCNQ2 (Soldovieri et al., 2006; Uehara et al., 2008).  Retigabine and ICA73, two anti-epileptic drugs, act via distinct mechanisms due to interactions with specific residues that underlie subtype specificity of KCNQ channel openers (Wang et al. 2016). Gabapentin at low concentrations is a activator of KCNQ3, KCNQ2/3 and KCNQ5 but not KCNQ2 or KCNQ4 (Manville and Abbott 2018). At high concentrations it can be inhibitory.  A  tight spatial and functional relationship between the DAT/GLT-1 transporters and the Kv7.2/7.3 potassium channel immediately readjusts the membrane potential of the neuron, probably to limit the neurotransmitter-mediated neuronal depolarization (Bartolomé-Martín et al. 2019). E-2-dodecenal from cilantro (Coriandrum sativum) is a potent activator and anticonvulsant that binds with an affinity of 60 nM to TMS5 in several KCNQ channels including KCNQ2 and 3 (Manville and Abbott 2019). Pathogenic variants in KCNQ2 and KCNQ3, paralogous genes encoding Kv7.2 and Kv7.3 voltage-gated K⁺ channel subunits, are responsible for early-onset developmental/epileptic disorders characterized by heterogeneous clinical phenotypes ranging from benign familial neonatal epilepsy (BFNE) to early-onset developmental and epileptic encephalopathy (DEE). KCNQ2 variants account for the majority of pedigrees with BFNE, and KCNQ3 variants are responsible for a much smaller subgroup (Miceli et al. 2020). The M240R variant mainly affects the voltage sensitivity, in contrast to previously analyzed BFNE Kv7.3 variants that reduce current density (Miceli et al. 2020).

6 TMS cell volume sensitive, voltage-gated K⁺ channel, KCNQ4 or Kv7.4 (mutations cause DFNA2, an autosomal dominant form of progressive hearing loss) (forms homomers or heteromers with KCNQ3) (localized to the basal membrane of cochlear outer hair cells and in several nuclei of the central auditory pathway in the brainstem). Four splice variants form heterotetramers; each subunit has different voltage and calmodulin-sensitivities (Xu et al., 2007).  Autosomal dominant mutant forms leading to progressive hearing loss (DFNA2) have been characterized (Kim et al. 2011). Phosphatidylinositol 4,5-bisphosphate (PIP2) and polyunsaturated fatty acids (PUFAs) impact ion channel function (Taylor and Sanders 2016). This channel may be present in mitochondria (Parrasia et al. 2019). Polyunsaturated fatty acids are modulators of KV7 channels (Larsson et al. 2020). The pathogenicity classification of KCNQ4 missense variants in clinical genetic testing has been described (Zheng et al. 2022). KCNQ4 potassium channel subunit deletion leads to exaggerated acoustic startle reflex in mice (Maamrah et al. 2023).

K⁺ voltage-gated channel, LQT-like subfamily; Kv7.1; KvLQT1. KCNQ1 (regulated by KCNE peptides (TC# 8.A.58) affect voltage sensor equilibrium (Rocheleau and Kobertz, 2007). Almost 300 mutations of KCNQ1 have been identified in patients, and most are linked to the long QT syndrome (LQT1), some in the voltage sensor (Peroz et al., 2008; Eldstrom et al. 2010; Qureshi et al. 2013; Ikrar et al. 2008). KCNQ1-KCNE1 complexes may interact intermittently with the actin cytoskeleton via the C-terminal region (Mashanov et al., 2010). The stoichiometry of the KCNQ1 - KCNE1 complex is flexible, with up to four KCNE1 subunits associating with the four KCNQ1 subunits of the channel (Nakajo et al., 2010). A familial mutation in the voltage-sensor of the KCNQ1 channel results in a cardiac phenotype (Henrion et al., 2012). KCNQ1 regulates insulin secretion in the MIN6 beta-cell line (Yamagata et al., 2011; Gofman et al., 2012).  Electrostatic interactions of S4 arginines with E1 and S2 contribute to gating movements of S4, but coupling requires the lipid phosphatidylinositol 4,5-bisphosphate (PIP2) as voltage-sensing domain activation failed to open the pore in the absence of PIP2 (Zaydman et al. 2013). The D242N mutation causes impaired action potential adaptation to exercise and an increase in heart rate. Moreover, the D242 amino acyl position is involved in the KCNE1-mediated regulation of the voltage-dependence of activation of the K_V7.1 channel (Moreno et al. 2017). The KCNQ1 channel interacts with MinK (KCNE1) to cause pore constriction, generating the slow delayed rectifier (IKs) current in the heart (Jalily Hasani et al. 2018). KCNQ1 rescues TMC1 plasma membrane expression but not mechanosensitive channel activity (Harkcom et al. 2019). Activation of the neuronal Kv7/KCNQ/M-current represents an attractive therapeutic strategy for treatment of hyperexcitability-related neuropsychiatric disorders such as epilepsy, pain, and depression, and channel openers for treatment of antiepilepsy have been developed (Zhang et al. 2019). The relationship between mutation locations in KCNQ1, which is a major gene in long QT syndrome (LQTS), and phenotype has been analyzed and used for risk stratification (Yagi et al. 2018). The proximal C-terminal regions of KCNQ1 and KCNE1 participate in a physical and functional interaction during channel opening that is sensitive to perturbation (Chen et al. 2019). Retigabine analogs are activators of Kv7 channels (Ostacolo et al. 2020). People with borderline QTc prolongations were carriers of KCNQ1 mutations in TMSs 2 and 5, leading to haploinsufficiency, and they are potentially at risk of developing drug-induced arrhythmia (Gouas et al. 2004). Collision induced unfolding differentiates functional variants of the KCNQ1 voltage sensor domain (Fantin et al. 2020). The activated KCNQ1 channel promotes a fibrogenic response in hereditary gingival fibromatosis via clustering and activation of Ras (Gao et al. 2020). QT syndrome (LQTS) increases the risk of life-threatening arrhythmia in young individuals with structurally normal hearts. It may involve sixteen genes such as the KCNQ1, KCNH2, and SCN5A (Lin et al. 2020). The human KCNQ1 voltage sensing domain (VSD) has been studied in lipodisq nanoparticles by electron paramagnetic resonance (EPR) spectroscopy (Sahu et al. 2020). Structural mechanisms for the activation of the human cardiac KCNQ1 channel by electro-mechanical coupling enhancers have been reviewed (Ma et al. 2022). The pathogenicity of KCNQ1 variants  using zebrafish as a model has been reviewed (Cui et al. 2023). Phosphatidyl-inositol-4,5-bisphosphate (PIP₂) is required for coupling between the voltage sensor and the pore of the potassium voltage-gated K_V7 channel family, especially the K_V7.1 channel. Modulation of the I(KS) channel by PIP₂ requires two binding sites per monomer (Kongmeneck et al. 2023).  Divergent regulation of the KCNQ1/E1 channel can be accomplished by targeted recruitment of protein kinase A to distinct sites on the channel complex (Zou et al. 2023).  Rare missense variants with a clear phenotype of Long QT Syndrome, type 1 (LQTS) have a high likelihood to be present within the pore and adjacent TMSs (S5-Pore-S6) (Novelli et al. 2023).  LHFPL5 is a key element in force transmission from the tip link to the hair cell mechanotransducer channel (Beurg et al. 2024).

The small conductance Ca²⁺-activated K⁺ channel, SkCa2, Sk2 or Kcnn2 (not inhibited by arachidonate) (activated by three small organic molecules, the 1-EBIO and N5309 channel enhancers and the DCEBIO channel modulation (Pedarzani et al., 2005)).  It is inhibited by protonation of outer pore histidine residues (Goodchild et al., 2009). The same is true for SK3 (K(Ca) 2.3 (Q9UGI6)). Regulates endothelial vascular function (Sonkusare et al., 2012).  Distinct subcellular mechanisms enhance the surface membrane expression by its interacting proteins, α-actinin 2 (TC# 8.A.66.1.3) and filamin A (TC# 8.A.66.1.4) (Zhang et al. 2016). SK channel activators can compensate for age-related changes of the autorhythmic functions of the cerebellum (Karelina et al. 2017). SK2 proteins are more abundant in Purkinje cells than in the ventricular myocytes of normal rabbit ventricles (Reher et al. 2017). Apamin inhibits and isoproterenol activates this and other SK (KCNN) channels, and activation by isoproterenol is sex-dependent (Chen et al. 2018).  Diverse interactions between KCa and TRP channels integrate cytoplasmic Ca²⁺, oxidative, and electrical signaling affecting cardiovascular physiology and pathology (Behringer and Hakim 2019). This channel may be present in mitochondria (Parrasia et al. 2019). A non-neuronal hSK3 isoform has a dominant-negative effect on hSK3 currents (Wittekindt et al. 2004). Medicinal plant products can interact with SKCa (Rajabian et al. 2022). Varients may cause conformational changes that alter the ability of the protein to modulate ion channel activities (d'Apolito et al. 2023).

The intermediate conductance, Ca²⁺-activated K⁺ channel, IKCa, Kcnn4, SK4, Sk4, Smik, Ik1 hIK1, IKCa or KCa3.1, also called the Gardos channel, of 543 aas and 6 TMSs. It is inhibited by 1 μM arachidonate which binds in the pore (Hamilton et al., 2003)). Nucleoside diphosphate kinase B (NDPK-B) activates KCa3.1 via histidine phosphorylation, resulting in receptor-stimulated Ca²⁺ flux and T cell activation (Di et al., 2010). It regulates endothelial vascular function (Sonkusare et al., 2012).  Tissue-specific expression of splice variants of the orthologous rat KCNN4 protein have been reported (Barmeyer et al. 2010).  Residues involved in gating have been identified (Garneau et al. 2014). It is also present in the inner mitochondrial membrane where increases of mitochondrial matrix [Ca²⁺] cause mtKCa3.1 opening, thus linking inner membrane K⁺ permeability and transmembrane potential to Ca²⁺ signalling (De Marchi et al. 2009). KCa3.1 (IKCa) channels are expressed in CA1 hippocampal pyramidal cells and contribute to the slow afterhyperpolarization that regulates spike accommodation (Turner et al. 2016). SK channel activators can compensate for age-related changes of the autorhythmic functions of the cerebellum (Karelina et al. 2017). The activation mechanism has been revealed by the cryoEM structure of the SK4-calmodulin complex (Lee and MacKinnon 2018).  It is responsible for hyperpolarization in some tumor cells (Lazzari-Dean et al. 2019). Mutations are linked to dehydrated hereditary stomatocytosis (xerocytosis) (Andolfo et al. 2015). This channel is present in mitochondria (Parrasia et al. 2019). KCNN4 promotes the progression of lung adenocarcinoma by activating the AKT and ERK signaling pathways (Xu et al. 2021). KCa3.1 channels in human microglia link extracellular ATP-evoked Ca²⁺ transients to changes in membrane conductance with an inflammation-dependent mechanism, and suggests that during brain inflammation, the KCa3.1-mediated microglial response to purinergic signaling may be reduced (Palomba et al. 2021). Both IK(Ca) and BK(Ca) regulate cell volume in human glioblastoma cells (Michelucci et al. 2023). Lysosomal Ca²⁺ release is sustained by ER→lysosome Ca²⁺ refilling and K⁺ efflux through the KCa3.1 channel in inflammasome activation and metabolic inflammation (Kang et al. 2024).

The archaeal voltage-regulated K channel, KvAP (Ruta et al., 2003). X-ray and solution structures are available. The latter shows phospholipid interactions with the isolated voltage sensor domain (Butterwick and MacKinnon 2010; Li et al. 2014). The gating-charge arginine in TMS4 of the voltage sensor forms part of the helical hairpin "paddle", and it moves 15-20 Å through the membrane to open the pore (Ruta et al., 2005). The orientation and depth of insertion of the voltage-sensing S4 helix has been determined (Doherty et al., 2010). A synthetic S6 segment derived from the KvAP channel self-assembles, permeabilizes lipid vesicles, and exhibits ion channel activity in bilayer lipid membrane (Verma et al., 2011).  Thus the gating mechanism combines structural rearrangements and electric-field remodeling ( Li et al. 2014).  KvAP has been reconstituted in Giant Unilamellar Vesicles (GUVs) (Garten et al. 2015).  TMS4 (S4) which senses voltage also promotes membrane insertion of the voltage-sensor domain (Mishima et al. 2016). KvAP has a configuration consistent with a water channel, possibly underlying the conductance of protons, and other cations, through voltage-sensor domains (Freites et al. 2006). The structural dynamics of the paddle motif loop in the activated conformation of the KvAP voltage sensor have been studied from biophysical standpoints (Das et al. 2019). The S4 alpha-helix, which is straight in the experimental crystal structure solved under depolarized conditions (Vm approximately 0), breaks into two segments when the cell membrane is hyperpolarized (Vm << 0) and reversibly forms a single straight helix following depolarization (Vm = 0) ((Bignucolo and Bernèche 2020). The outermost segment of S4 translates along the normal to the membrane, bringing new perspective to previously paradoxical accessibility experiments that were initially thought to imply the displacement of the whole VSD across the membrane. The breakage of S4 under (hyper)polarization could be a general feature of Kv channels with a non-swapped topology. The surface charge of the membrane does not significantly affect the topology and structural dynamics of the sensor loop in membranes (Das and Raghuraman 2021). The dynamic variability of the sensor loop is preserved in both zwitterionic (POPC) and anionic (POPC/POPG) lipid membranes. The lifetime distribution analysis for the NBD-labelled residues by the maximum entropy method (MEM) demonstrates that, in contrast to micelles, the membrane environment not only reduces the relative discrete population of sensor loop conformations, but also broadens the lifetime distribution peaks. The local curvature of cellular membranes acts as a driving force for the targeting of membrane-associated proteins to specific membrane domains, as well as a sorting mechanism for transmembrane proteins, as demonstrated for KvAP (Kluge et al. 2022). 

Voltage-gated K⁺ channel, chain A, Shaker-related, K_v1.2 or KCNA2 (Crystal structure known, Long et al., 2007; Chen et al. 2010). It functions with the auxiliary subunit, Ivβ1.2; 8.A.5.1.1) (Peters et al. 2009).  Delemotte et al. (2010) described the effects of sensor domain mutations on molecular dynamics of K_v1.2.  The Sigma 1 receptor (Q99720; Sigma non-opioid intracellular receptor 1) interacts with K_v1.2 to shape neuronal and behavioral responses to cocaine (Kourrich et al. 2013).  Amino acid substitutions cause Shaker to become heat-sensing (opens with increasing temperature as for TrpV1) or cold-sensing (opens with decreasing temperature as for TrpM8) (Chowdhury et al. 2014).  The Shaker Kv channel was truncated after the 4th transmembrane helix S4 (Shaker-iVSD) which showed altered gating kinetics and formed a cation-selective ion channel with a strong preference for protons (Zhao and Blunck 2016).  Direct axon-to-myelin linkage by abundant K_V1/Cx29 (TC# 1.A.24.1.12) channel interactions in rodent axons supports the idea of an electrically active role for myelin in increasing both the saltatory conduction velocity and the maximal propagation frequency in mammalian myelinated axons (Rash et al. 2016). A cryoEM structure (3 - 4 Å resolution; paddle chimeric channel; closed form) in nanodiscs has been determined (Matthies et al. 2018). Possible gating mechanisms have been discussed (Kariev and Green 2018; Infield et al. 2018). Pathogenic variants in KCNA2, encoding the voltage-gated potassium channel Kv1.2, have been identified as the cause for an evolving spectrum of neurological disorders. Affected individuals show early-onset developmental and epileptic encephalopathy, intellectual disability, and movement disorders resulting from cerebellar dysfunction (Döring et al. 2021). In addition, individuals with a milder course of epilepsy, complicated hereditary spastic paraplegia, and episodic ataxia have been reported. Biophysical properties of a delayed rectifier K⁺ current can contribute to its role ingenerating spontaneous myogenic activity (Hu et al. 2021). The local curvature of cellular membranes acts as a driving force for the targeting of membrane-associated proteins to specific membrane domains, as well as a sorting mechanism for transmembrane proteins, as demonstrated for the chimeric channel, Kv1.2/2.1; KvChim induces a strong positive membrane curvature (Kluge et al. 2022). 2-Aminoethoxydiphenyl borate (2-APB) has inhibitory effects on three K_V1 channels, Kv1.2, Kv1.3 and Kv1.4 (Zhao et al. 2023). Voltage-gated K⁺ channels have two distinct gates that regulate ion flux: the activation gate (A-gate) formed by the bundle crossing of the S6 transmembrane helices and the slow inactivation gate in the selectivity filter. These two gates are bidirectionally coupled. Szanto et al. 2023 suggested that the coupling between the A-gate and the slow inactivation gate is mediated by rearrangements in the S6 segment. S6 rearrangements are consistent with a rigid rod-like rotation of S6 around its longitudinal axis upon inactivation.

Voltage-gated K⁺ channel, Kv1.1 or KCNA1. It is palmitoylated, modulating voltage sensing (Gubitosi-Klug et al. 2005). It is regulated by syntaxin (TC family 8.A.91) through dual action on channel surface expression and conductance (Feinshreiber et al., 2009). Defects cause episodic ataxia type 1 (EA1), an autosomal dominant K⁺ channelopathy accompanied by short attacks of cerebellar ataxia and dysarthria (D'Adamo et al. 2014; Yuan et al. 2020). Direct axon-to-myelin linkage by abundant K_V1/Cx29 channel interactions in rodent axons supports the idea of an electrically active role for myelin in increasing both the saltatory conduction velocity and the maximal propagation frequency in mammalian myelinated axons (Rash et al. 2016). Kv1.1 is present in bull sperm where it is necessary for normal sperm progressive motility, percent capacitated spermatozoa (B-pattern) and the acrosome reaction (Gupta et al. 2018). Gating induces large aqueous volumetric remodeling (Díaz-Franulic et al. 2018). Paulhus et al. 2020 have reviewed the pathology of mutants in this protein and showed that epilepsy or seizure-related variants tend to cluster in the S1/S2 transmembrane domains and in the pore region of Kv1.1, whereas EA1-associated variants occur along the whole length of the protein, but variants at the C-terminus are more likely to suffer from seizures and neurodevelopmental disorders (Yuan et al. 2020). Mutation in KCNA1 has been identified that impairs voltage sensitivity (Imbrici et al. 2021). Altering expression of the genes encoding Kv1.1, Piezo2, and TRPA1 regulate the response of mechanosensitive muscle nociceptors (Nagaraja et al. 2021). Genetic variants have expanded the functional, molecular, and pathological diversity of KCNA1 channelopathies (Paulhus and Glasscock 2023). Carbamazepine suppresses the impaired startle response and brain hyperexcitability in kcna1a(-/-) zebrafish but had no effect on the seizure frequency in Kcna1(-/-) mice, suggesting that this EA1 zebrafish model might better translate to humans than rodents (Dogra et al. 2023).

The voltage-gated K⁺ channel subfamily D member 3, KCND3 or K_v4.3.  Mutations cause spinocerebellar ataxia type 19 (Duarri et al. 2012). Positively charged residues in S4 contribute to channel inactivation and recovery (Skerritt and Campbell 2007). The crystal structure of Kv4.3 with its regulatory subunit, Kchip1, has been solved (2NZ0) (Wang et al. 2007).

Margatoxin-sensitive voltage-gated K⁺ channel, Kv1.3 (in plasma and mitochondrial membranes of T lymphocytes) (Szabò et al., 2005). K_v1.3 associates with the sequence similar (>80%) K_v1.5 protein in macrophage forming heteromers that like K_v1.3 homomers are r-margatoxin sensitive (Vicente et al., 2006). However, the heteromers have different biophysical and pharmacological properties. The K_v1.3 mitochondrial potassium channel is involved in apoptotic signalling in lymphocytes (Gulbins et al., 2010). Interactions between the C-terminus  of K_v1.5 and K_vβ regulate pyridine nucleotide-dependent changes in channel gating (Tipparaju et al., 2012).  Intracellular trafficking of the K_V1.3 K⁺ channel is regulated by the pro-domain of a matrix metalloprotease (Nguyen et al. 2013).  Direct binding of caveolin regulates K_v1 channels and allows association with lipid rafts (Pérez-Verdaguer et al. 2016). Addtionally, Na_vBeta1 interacts with the voltage sensing domain (VSD) of K_v1.3 through W172 in the transmembrane segment to modify the gating process (Kubota et al. 2017). During insertion of Kv1.3, the extended N-terminus of the second α-helix, S2, inside the ribosomal tunnel is converted into a helix in a transition that depends on the nascent peptide sequence at specific tunnel locations (Tu and Deutsch 2017). The microRNA, mmumiR449a, reduced the mRNA expression levels of transient receptor potential cation channel subfamily A member 1 (TRPA1), and calcium activated potassium channel subunit alpha1 (KCNMA1) and increased the level of transmembrane phosphatase with tension homology (TPTE) in the DRG cells (Lu et al. 2017). This channel is regulation by sterols (Balajthy et al. 2017). Loss of function causes atrial fibrillation, a rhythm disorder characterized by chaotic electrical activity of cardiac atria (Olson et al. 2006). The N-terminus and S1 of Kv1.5 can attract and coassemble with the rest of the channel (i.e. Frag(304-613)) to form a functional channel independently of the S1-S2 linkage (Lamothe et al. 2018). This channel may be present in mitochondria (Parrasia et al. 2019). Kv1.3 plays an essential role in the immune response mediated by leukocytes and is functional at both the plasma membrane and the inner mitochondrial membrane. Plasma membrane Kv1.3 mediates cellular activation and proliferation, whereas mitochondrial Kv1.3 participates in cell survival and apoptosis (Capera et al. 2022). Kv1.3 uses the TIM23 complex to translocate to the inner mitochondrial membrane. This mechanism is unconventional because the channel is a multimembrane spanning protein without a defined N-terminal presequence. Transmembrane domains cooperatively mediate Kv1.3 mitochondrial targeting involving the cytosolic HSP70/HSP90 chaperone complex as a key regulator of the process (Capera et al. 2022).

Voltage-gated K⁺ channel, Shaker. Shaker and Shab K⁺ channels are blocked by quinidine (Gomez-Lagunas, 2010). Also regulated by unsaturated fatty acids (Börjesson and Elinder, 2011).  TMSs 3 and 4 comprise the voltage sensor paddle (Xu et al. 2013).  Partially responsible for action potential repolarization during synaptic transmission (Ford and Davis 2014). Shaker K⁺ channels can be mutated in S4 to create an analogous "omega" pore (Held et al. 2018). The NMR structure of the isolated Shaker voltage-sensing domain in LPPG micelles has been reported (Chen et al. 2019). Substituting the first S4 arginine with a smaller amino acid opens a high-conductance pathway for solution cations in the Shaker K⁺ channel at rest. The cationic current does not flow through the central K⁺ pore and is influenced by mutation of a conserved residue in S2, suggesting that it flows through a protein pathway within the voltage-sensing domain (Tombola et al. 2005). The current can be carried by guanidinium ions, suggesting that this is the pathway for transmembrane arginine permeation. Tombola et al. 2005 proposed that when S4 moves, it ratchets between conformations in which one arginine after another occupies and occludes to ions in the narrowest part of this pathway. Specific resin acids activate and open voltage-gated channels  dependent on its exact binding dynamics (Silverå Ejneby et al. 2021). Charge-voltage curves of a Shaker potassium channel are not hysteretic at steady state (Cowgill and Chanda 2023). shaker is a critical sleep regulator in Drosophila (Cirelli et al. 2005).

K⁺ voltage-gated ether-a-go-go-related channel, H-ERG (KCNH2; Erg; HErg; Erg1, Kv11.1) subunit K_v11.1 (long QT syndrome type 2) (Gong et al., 2006; Chartrand et al. 2010; McBride et al. 2013). Selective expression of HERG and Kv2 channels influences proliferation of uterine cancer cells (Suzuki and Takimoto 2004). H-ERG forms a heteromeric K⁺ channel regulating cardiac repolarization, neuronal firing frequency and neoplastic cell growth (Szabó et al., 2011). Oligomerization is due to N-terminal interactions between two splice variants, hERG1a and hERG1b (Phartiyal et al., 2007). Structure function relationships of ERG channel activation and inhibition have been reviewed (Durdagi et al., 2010). Interactions between the N-terminal domain and the transmembrane core modulate hERG K channel gating (Fernández-Trillo et al., 2011). The marine algal toxin azaspiracid is an open state blocker (Twiner et al., 2012). Verapamil blocks channel activity by binding to Y₆₅₂ and F₆₅₆ in TMS S₆ (Duan et al. 2007).  Hydrophobic interactions between the voltage sensor and the channel domain mediate inactivation (Perry et al. 2013), but voltage sensing by the S4 segment can be transduced to the channel gate in the absence of physical continuity between the two modules (Lörinczi et al. 2015).  Mutations give rise to long QT syndrome (Osterbur et al. 2015).  Polyphenols such as caffeic acid, phenylethyl ester (CAPE) and curcumin inhibit by modification of gating, not by blocking the pore (Choi et al. 2013). Potassium ions can inhibit tumorigenesis through inducing apoptosis of hepatoma cells by upregulating potassium ion transport channel proteins HERG and VDAC1 (Xia et al. 2016).  Incorrectly folded hERG can be degraded by Bag1-stimulated Trc-8-dependent proteolysis (Hantouche et al. 2016). The S1 helix regulates channel activity. Thus, S1 region mutations reduce both the action potential repolarizing current passed by Kv11.1 channels in cardiac myocytes, as well as the current passed in response to premature depolarizations that normally helps protect against the formation of ectopic beats (Phan et al. 2017). Interactions of beta1 integrins with hERG1 channels in cancer cells stimulate distinct signaling pathways that depended on the conformational state of hERG1 (Becchetti et al. 2017). ERG1 is sensitive to the alkaloid, ginsenoside 20(S) Rg3 which alters the gating of hERG1 channels by interacting with and stabilizing the voltage sensor domain in an activated state (Gardner et al. 2017). Channels split at the S4-S5 linker, at the intracellular S2-S3 loop, and at the extracellular S3-S4 loop are fully functional channel proteins (de la Peña et al. 2018). IKr is the rapidly activating component of the delayed rectifier potassium current, the ion current largely responsible for the repolarization of the cardiac action potential. Inherited forms of long QT syndrome (LQTS) in humans are linked to functional modifications in the Kv11.1 (hERG) ion channel and potentially life threatening arrhythmias. hERG1b affects the generation of the cardiac Ikr and plays an important role in cardiac electrophysiology (Perissinotti et al. 2018). X-ray crystallography and cryoEM have revealed features of the "nonswapped" transmembrane architecture, an "intrinsic ligand," and small hydrophobic pockets off a pore cavity. Drug block and inactivation mechanisms are discussed (Robertson and Morais-Cabral 2019). It forms a complex with β-integrin (TC#9.B.87.1.25) and NHE1 (TC# 2.A.36.1.13) (Iorio et al. 2020). Cardiotoxicity is caused mainly by the inhibition of human ether-a-go-go related gene (hERG) channel protein which leads to a life-threatening condition known as cardiac arrhythmia and is due to probable collapse of the pore. (Koulgi et al. 2021). Transmembrane hERG channel currents have been measured based on solvent-free lipid bilayer microarrays (Miyata et al. 2021). A computational method for identifying an optimal combination of existing drugs to repair the action potentials of SQT1 ventricular myocytes has been published (Jæger et al. 2021). Ginsenoside Rg3 may be a promising cardioprotective agent against vandetanib-induced QT interval prolongation through targeting hERG channels (Zhang et al. 2021). Insight has been obtained into the potassium currents of hERG (Guidelli 2023). Two novel KCNH2 mutations contribute to long QT syndrome (Owusu-Mensah et al. 2024). Channel activity is affected by moxifloxacin, terfenadine, arsenic, pentamidine, erythromycin, and sotalol (Goineau et al. 2024).

The KCNH1 K+ channel protein of 989 aas and 6 TMSs.  Tian et al. 2023 expanded the phenotypic spectrum of KCNH1 and explored the correlations between epilepsy and molecular sub-regional locations.  They found two novel missense variants of KCNH1 in three individuals with isolated FS/epilepsy. Variants caused a spectrum of epileptic disorders ranging from a benign form of genetic isolated epilepsy/FS to intractable form of epileptic encephalopathy. The genotypes and variant locations helped explain the phenotypic variation of patients with KCNH1 variants (Tian et al. 2023).

K⁺ voltage-gated channel, rEAG1; K_v10.1; rat ether a go-go channel 1 (962 aas). Blocked by Cs⁺, Ba²⁺ and quinidine (Schwarzer et al., 2008). Cysteines control the N- and C-linker-dependent gating of KCNH1 potassium channels  (Sahoo et al., 2012).  The 3-d structure has been determined at 3.8 Å resolution using single-particle cryo-EM with calmodulin bound. The structure suggests a novel mechanism of voltage-dependent gating. Calmodulin binding closes the potassium pore (Whicher and MacKinnon 2016). Eag1 has three intracellular domains: PAS, C-linker, and CNBHD. Whicher and MacKinnon 2019 demonstrated that the Eag1 intracellular domains are not required for voltage-dependent gating but likely interact with the VS to modulate gating. Specific interactions between the PAS, CNBHD, and VS domains modulate voltage-dependent gating, and VS movement destabilizes these interactions to promote channel opening. Mutations affecting these interactions render Eag1 insensitive to calmodulin inhibition (Whicher and MacKinnon 2019). The structure of the calmodulin insensitive mutant in a pre-open conformation suggests that channel opening may occur through a rotation of the intracellular domains, and calmodulin may prevent this rotation by stabilizing interactions between the VS and the other intracellular domains. Intracellular domains likely play a similar modulatory role in voltage-dependent gating of the related Kv11-12 channels. The human ortholog, EAG or EAG-1, is 989 aas long and is 95% identical to the rat protein.  In ether-a-go-go K⁺ channels, voltage-dependent activation is modulated by ion binding to a site located in an extracellular-facing crevice between transmembrane segments S2 and S3 in the voltage sensor. Silverman et al. 2004 found that acidic residues, D278 in S2 and D327 in S3, are able to coordinate a variety of divalent cations, including Mg²⁺, Mn²⁺, and Ni²⁺, which have qualitatively similar functional effects, but different half-maximal effective concentrations. EAG (ether-a-go-go) voltage-dependent K⁺ channels with similarities and Differences in the structural organization and gating (Barros et al. 2020).

Cyclic nucleotide-binding, voltage-gated, Mg²⁺-dependent, CaMKII-regulated K⁺ channel, Eag.  Eag recruits CASK (TC# 9.B.106.3.2) to the plasma membrane; forms a heterotetramer (Liu et al. 2010). Phosphorylation is catalyzed by CaMKII (TC# 8.A.104.1.11)

K⁺- and Na⁺-conducting NaK channel, NaK2K of 97 aas and 2 TMSs. The 3-D structure has been solved with Na⁺ and K⁺bound (Shi et al., 2006).  It exhibits tight structural and dynamic coupling between the selectivity filter and the channel scaffold (Brettmann et al. 2015). A hydrophobic residue at the bottom of the selectivity filter, Phe92, appears in dual conformations. One of the two conformations of Phe92 restricts the diameter of the exit pore around the selectivity filter, limiting ion flow through the channel, while the other conformation of Phe92 provides a larger-diameter exit pore from the selectivity filter. Thus, Phe92 acts as a hydrophobic gate (Langan et al. 2020).

The 6TMS bacterial cyclic nucleotide-regulated, voltage independent channel, MlotiK1 or MloK1 (Clayton et al., 2008). Gating involves large rearrangements of the cyclic nucleotide-binding domains (Mari et al., 2011).  The electron crystalographic structure is available (PDB 4CHW) revealing ligand-induced structural changes (Schünke et al. 2011; Scherer et al. 2014; Kowal et al. 2014). Such changes may be lipid dependent (McCoy et al. 2014). High-speed atomic force microscopy has been used to measure millisecond to microsecond dynamics (Heath and Scheuring 2019).

Calcium-, magnesium- and voltage-activated K⁺ channel, Slo1 (Kcma1; KCNMA, KCNMA1), a BK channel, of 1236 aas and 6 N-terminal TMSs. Its activation dampens the excitatory events that elevate the cytosolic Ca²⁺ concentration and/or depolarize the cell membrane. It therefore contributes to repolarization of the membrane potential, and it plays a key role in controlling excitability in a number of systems. Ethanol and carbon monoxide-bound heme increase channel activation while heme inhibits channel activation (Tang et al. 2003). The molecular structures of the human Slo1 channel in complex with beta4 has been solved revealing four beta4 subunits, each containing two transmembrane helices, encircling Slo1, contacting it through helical interactions inside the membrane. On the extracellular side, beta4 forms a tetrameric crown over the pore. Structures with high and low Ca²⁺ concentrations show that identical gating conformations occur in the absence and presence of beta4, implying that beta4 serves to modulate the relative stabilities of 'pre-existing' conformations rather than creating new ones (Tao and MacKinnon 2019). BK channels show increased activities in Angelman syndrome due to genetic defects in the ubiquitin protein ligase E3A (UBE3A) gene (Sun et al. 2019). It is a large-conductance potassium (BK) channel that can be synergistically and independently activated by membrane voltage and intracellular Ca²⁺. The only covalent connection between the cytosolic Ca²⁺-sensing domain and the TM pore and voltage sensing domains is a 15-residue 'C-linker' which plays a direct role in mediating allosteric coupling between BK domains (Yazdani et al. 2020).  Site specific deacylation by the alpha/beta acyl-hydrolase domain-containing protein 17A, ABHD17a (Q96GS6, 310 aas), controls BK channel splice variant activity (McClafferty et al. 2020). Compared with the structure of isolated hSlo1 Ca²⁺ sensing gating rings, two opposing subunits in hBK unfurled, resulting in a wider opening towards the transmembrane region of hBK. In the pore gate domain, two opposing subunits moved downwards relative to the two other subunits (Tonggu and Wang 2022). A gating lever, mediated by S4/S5 segment interactions within the transmembrane domain, rotates to engage and stabilize the open conformation of the S6 inner pore helix upon V sensor activation (Sun and Horrigan 2022). An indirect pathway, mediated by the carboxyl-terminal cytosolic domain (CTD) and C-linker connects the CTD to S6, and stabilizes the closed conformation when V sensors are at rest (Sun and Horrigan 2022). Co-dependent regulation of p-BRAF (TC# 8.A.23.1.48) and the potassium channel KCNMA1 levels drives glioma progression (Xie et al. 2023). Potassium channelopathies associated with epilepsy-related syndromes and directions for therapeutic interventionhave been reviewed (Gribkoff and Winquist 2023). The influx of Ca^2+, mediated by the hypotonic-induced activation of mechanosensitive channels, is a key step for opening both the BK(Ca) and the IK(Ca) channels. The influx of Ca²⁺, mediated by the hypotonic-induced activation of mechanosensitive channels, is a key step for opening both the BK(Ca) and the IK(Ca) (TC# 1.A.1.16.2) channels (Michelucci et al. 2023).  Disease-associated KCNMA1 variants decrease circadian clock robustness in channelopathy mouse models (Dinsdale et al. 2023).  High-resolution structures illuminate key principles underlying voltage and LRRC26 regulation of Slo1 channels (Kallure et al. 2023).

Slo2.2 sodium-activated potassium channel subfamily T member 1 of 1217 aas and 6 putative N-terminal TMSs plus a P-loop.  Cryo electron microscopic structures of the chicken orthologue at 4.5 Å resolution has been solved, revealing a large cytoplasmic gating ring in which resides the Na⁺-binding site and a transmembrane domain that closely resembles voltage-gated K⁺ channels (Hite et al. 2015).

K⁺ channel, AKT1; may form heteromeric channels with KC1 (TC # 1.A.1.4.9) (Geiger et al., 2009).  Required for seed development and postgermination growth in low potassium (Pyo et al. 2010).  Functions optimally with intermediate potassium concentrations (~1 mM) (Nieves-Cordones et al. 2014). In barley, it may play a role in drought resistance (Cai et al. 2019).  HAK/KUP/KT8, GrAKT2.1 and GrAKT1.1 potassium channels may function in response to abiotic stress in Gossypium raimondii (Azeem et al. 2021). Plants obtain nutrients from the soil via transmembrane transporters and channels in their root hairs, from which ions radially transport in toward the xylem for distribution across the plant body. Dickinson et al. 2021 determined structures of the hyperpolarization-activated channel, AKT1, from Arabidopsis thaliana, which mediates K⁺ uptake from the soil into plant roots. The structures of AtAKT1, embedded in lipid nanodiscs, show that the channel undergoes a reduction of C4 to C2 symmetry, possibly to regulate its electrical activation.

The voltage-sensitive inward rectifying K⁺ channel, KAT1 (similar to 1.A.1.4.3; activated by protein 14-3-3 (AAF87262)) (Sottocornola et al., 2006). May also transport Na⁺ and Cs⁺ (Nakamura and Gaber, 2009). Forms heterotetrameric channels with KAT2 with a stoichiometry of 2:2 (Lebaudy et al., 2010). The pH-sensor is built of a sensory cloud rather than of single key amino acids (Gonzalez et al., 2011). The transmembrane core region of KAT1 is important for its activity in S. cerevisiae, and this involves not only the pore region but also parts of its voltage-sensor domain (Saito et al. 2017). Electromechanical coupling and gating polarity in KAT1 displays a depolarized voltage sensor, which interacts with a closed pore domain directly via two interfaces and indirectly via an intercalated phospholipid. Direct interaction between the sensor and the C-linker hairpin in the adjacent pore subunit is the primary determinant of gating polarity (Clark et al. 2020). Possibly an inward motion of the S4 sensor helix of 5-7 Å underlies a direct-coupling mechanism, driving a conformational reorientation of the C-linker and ultimately opening the activation gate formed by the S6 intracellular bundle. KAT1, and presumably other hyperpolarization-gated plant CNBD channels, can open from an S4-down VSD conformation homologous to the divalent/proton-inhibited conformation of EAG family K⁺ channels (Zhou et al. 2021).

Orthologue K⁺/Na⁺ pacemaker channel, Hcn4 (Scicchitano et al., 2012).  Hyperpolarization-activated cyclic nucleotide-regulated HCN channels underlie the Na⁺-K⁺ permeable IH pacemaker current. As with other voltage-gated members of the 6-transmembrane KV channel superfamily, opening of HCN channels involves dilation of a helical bundle formed by the intracellular ends of S6, but this is promoted by inward, not outward, displacement of S4. Direct agonist binding to a ring of cyclic nucleotide-binding sites, one of which lies immediately distal to each S6 helix, imparts cAMP sensitivity to HCN channel opening. At depolarized potentials, HCN channels are further modulated by intracellular Mg²⁺ which blocks the open channel pore and blunts the inhibitory effect of outward K⁺ flux. Lyashchenko et al. 2014 showed that cAMP binding to the gating ring enhances not only channel opening but also the kinetics of Mg²⁺ block.  Mutations in HCN4 cause sick sinus and the Brugada syndrome, cardiac abnormalities. HCN4 is associated with famiial sinus bradycardia (Boulton et al. 2017). Activation of Hcn4 by cAMP has been reviewed (Porro et al. 2020). The HCN1-4 channel family is responsible for the hyperpolarization-activated cation current If/Ih that controls automaticity in cardiac and neuronal pacemaker cells. Saponaro et al. 2021 presented cryo-EM structures of HCN4 in the presence or absence of bound cAMP, displaying the pore domain in closed and open conformations. Analysis of cAMP-bound and -unbound structures shed light on how ligand-induced transitions in the channel cytosolic portion mediate the effect of cAMP on channel gating and highlighted the regulatory role of a Mg²⁺ coordination site formed between the C-linker and the S4-S5 linker. Comparison of open/closed pore states shows that the cytosolic gate opens through concerted movements of the S5 and S6 transmembrane helices. Furthermore, in combination with molecular dynamics analyses, the open pore structures provide insights into the mechanisms of K⁺/Na⁺ permeation (Saponaro et al. 2021). 

Hyperpolarization-activated cyclic nucleotide-gated (HCN) inward current-carrying cationic channel, I(f), (HCN2/HCN4) (Ye and Nerbonne, 2009).  Functional interactions between the HCN2 TM region and C-terminal region govern multiple CNB fold-mediated mechanisms, implying that the molecular mechanisms of autoinhibition, open-state trapping, and Quick-Activation include participation of TM region structures (Page et al. 2020). Rhythmic activity in pacemaker cells, as in the sino-atrial node in the heart, depends on the activation of HCN channels. As in depolarization-activated K⁺ channels, the fourth transmembrane segment S4 functions as the voltage sensor in hyperpolarization-activated HCN channels (Wu et al. 2021). S4 in HCN channels moves in two steps in response to hyperpolarizations, and the second S4 step correlates with gate opening (Wu et al. 2021). It is a nuclear hormone receptor that binds estrogens with an affinity similar to that of ESR1/ER-alpha, and activates expression of reporter genes containing estrogen response elements (ERE) in an estrogen-dependent manner (Koyama et al. 2010). It may lack ligand binding ability and has no or only very low ERE binding activity, resulting in the loss of ligand-dependent transactivation ability. Male moujse ejaculation drives sexual satiety and selectively activates Esr2neurons in the BNSTpr of both sexes (Zhou et al. 2023).  Changes in binding affinity, rather than changes in cAMP concentration, can modulate HCN channels (Porro et al. 2024).

Cyclic nucleotide-gated cation channel α3 (CNGA3 or CNG3); photoreceptor cGMP-gated channel α-subunit. Also possibly expressed in inner ear cell cells where it binds to an intracellular C-terminal domain of EMILIN1 (Selvakumar et al., 2012).  Elastic network model analysis of the CNGA3 channel supports a modular model of allosteric gating, according to which protein domains are quasi-independent: they can move independently but are coupled to each other allosterically (Gofman et al. 2014). An intact S4 is required for proper protein folding and/or assembly involving two glycosylation sites in the endoplasmic reticulum membrane (Faillace et al. 2004). It may function with CNGB3 (TC# 1.A.1.5.37; Q9NQW8; 809 aas and 6 TMSs).

Cyclic nucleotide-gated K⁺channel, SthK, of 430 aas, probably with 6 TMSs in a 2 + 2 + 1 + P-loop + 1 TMS arrangement.  The channel is activated by cAMP, not by cGMP, and is highly specific for K⁺ over Na⁺.  It has a C-terminal hydrophilic cAMP-binding domain linked to the 6 TMS channel domain (Brams et al. 2014). An SthK C-linker domain is essential for coupling cyclic nucleotide binding to channel opening (Evans et al. 2020). An agonist-dependent conformational change in which residues of the B'-helix displayed outward movement with respect to the symmetry axis of the channel in the presence of cAMP was observed, but not with the partial agonist, cGMP. This conformational rearrangement was observed both in detergent-solubilized SthK and in channels reconstituted into lipid nanodiscs. In addition to outward movement of the B'-helix, channel activation involves upward translation of the cytoplasmic domain with formation of state-dependent interactions between the C-linker and the transmembrane domain (Evans et al. 2020). Three-deminsional structures are available (7RSY_A-D). SthK is active in a sparsely tethered lipid bilayer membranes (Andersson et al. 2023).

Hyperpolarization-activated and cyclic nucleotide-gated K⁺ channel, HCN (bCNG-1) (P_Na⁺/P_K⁺ ≈ 0.3). The human orthologue (O88703) is 863 aas in length and also catalyzes mixed monovalent cation currents K⁺:Na⁺= 4:1 (Lyashchenko and Tibbs et al., 2008). Biel et al. (2009) presented a detailed review of hyperpolarization-activated cation-channels. They are inhibited by nicotine and epibatidine which bind to the inner pore (Griguoli et al., 2010). They control cardiac and neuronal rhythmicity. HCN channels contain cyclic nucleotide-binding domains (CNBDs) in their C-terminal regions, linked to the pore-forming transmembrane segment with a C-linker. The C-linker couples the conformational changes caused by the direct binding of cyclic nucleotides to the HCN pore opening. Cyclic dinucleotides antagonize the effect of cyclic nucleotides in HCN4 but not in HCN2 channels. Interaction of the C-linker/CNBD with other parts of the channel appears to be necessary for cyclic-dinucleotide binding in HCN4 channels (Hayoz et al. 2017). A conformational trajectory of allosteric gating of the human cone photoreceptor cyclic nucleotide-gated channel has been documented (Hu et al. 2023).  The voltage-sensor rearrangements, directly influenced by membrane lipid domains, can explain the heightened activity of pacemaker HCN channels when localized in cholesterol-poor, disordered lipid domains, leading to membrane hyperexcitability and diseases (Handlin and Dai 2023).  Opioid-induced hyperalgesia and tolerance are driven by HCN ion channels (Han et al. 2024).

TAX-4 cyclic nucleotide-gated cation channel A (CNGA) of 733 aas (Wojtyniak et al. 2013).  Li et al. 2017 determined the 3.5 Å resolution single-particle electron cryo-microscopy structure  in the cyclic guanosine monophosphate (cGMP)-bound open state. The channel has an unusual voltage-sensor-like domain, accounting for its deficient voltage dependence. A carboxy-terminal linker connecting S6 and the cyclic-nucleotide-binding domain interacts directly with both the voltage sensor-like domain and the pore domain, forming a gating ring that couples conformational changes triggered by cyclic nucleotide binding to the gate. The selectivity filter is lined by the carboxylate side chain of a functionally important glutamate and three rings of backbone carbonyls (Li et al. 2017).

spHCN1 is a pacemaker hyperpolarization-activated cyclic nucleotide-gated (HCN) non-selective cation channel of 767 aas and 6 TMSs that opens due to inward movement of the positive charges in the fourth TMS (S4). This channel is similar to a COOH-terminal-deleted HCN1 channel, suggesting that the main functional differences between spHCN and HCN1 channels are due to differences in their COOH termini (Vemana et al. 2004). These channels open after only two S4s have moved, and S4 motion is rate limiting during voltage activation of spHCN channels (Bruening-Wright et al. 2007). HCN channels regulate electrical activity in the heart and brain. Distinct from mammalian isoforms, the sea urchin (spHCN) channel exhibits strong voltage-dependent inactivation in the absence of cAMP (Idikuda et al. 2018). The voltage sensor undergoes a large downward motion during hyperpolarization (Dai et al. 2019). Sea urchin HCN1 and 2 (TC# 1.A.1.5.33) (spHCN) channels undergo inactivation with hyperpolarization which occurs only in the absence of cyclic nucleotide (Dai et al. 2021). Removing cAMP produces a largely rigid-body rotation of the C-linker relative to the transmembrane domain, bringing the A' helix of the C-linker in close proximity to the voltage-sensing S4 helix. In addition, rotation of the C-linker is elicited by hyperpolarization minus cAMP. Thus, in contrast to electromechanical coupling for channel activation - the A' helix serves to couple the S4-helix movement for channel inactivation, which is likely a conserved mechanism for CNBD-family channels (Dai et al. 2021).

Multi-domain cation channel with a C-terminal cyclic nucleotide-binding domain; of 465 aas and 6 TMS, LliK.  Cyclic nucleotide-gated (CNG) and hyperpolarization-activated cyclic nucleotide-regulated (HCN) channels play roles in phototransduction, olfaction, and cardiac pace making.  James et al. 2017 used cryoEM to determine the structure of the intact LliK CNG channel. A short S4-S5 linker connects voltage-sensing and pore domains to produce a non-domain-swapped transmembrane architecture. The conformation of the LliK structure may represent a functional state of this channel family not seen before (James et al. 2017).

HCN1 is a hyperpolarization-activated cyclic nucleotide-gated (HCN) channel of 890 aas and 6 TMSs that opens due to inward movement of the positive charges in the fourth transmembrane domain (S4).  These channels open after only two S4s have moved, and S4 motion is rate limiting during voltage activation of spHCN channels (Bruening-Wright et al. 2007). HCN1 exhibits weak selectivity for potassium over sodium ions.  It's structure (3.5 Å resolution) is known (Lee and MacKinnon 2017).  It contributes to the native pacemaker currents in heart and neurons. It may also mediate responses to sour stimuli. It is inhibited by Cs⁺, zatebradine, capsazepine and ZD7288 (Gill et al. 2004). HCN1 mutational variants include epileptic encephalopathy and common generalized epilepsy. HCN1 has a pivotal function in brain development and control of neuronal excitability (Marini et al. 2018). The interaction with filamin A seems to contribute to localizing HCN1 channels to specific neuronal areas and to modulating channel activity (Gravante et al. 2004). The HCN domain is required for HCN channel cell-surface expression, and it couples voltage- and cAMP-dependent gating mechanisms (Wang et al. 2020). Changes in the local S4 environment provide a voltage-sensing mechanism for mammalian hyperpolarization-activated HCN channels (Bell et al. 2004). Cation leak is an important pathogenic mechanism in HCN1-mediated developmental and epileptic encephalopathy (DEE), and seizures are exacerbated by sodium channel blockers in patients with HCN1 variants that cause cation leak (McKenzie et al. 2023). HCN1 epilepsy is progressing from genetics and mechanisms to precision therapies (Bleakley and Reid 2023).  Opioid-Induced Hyperalgesia and Tolerance Are Driven by HCN Ion Channels (Han et al. 2024).

The cyclic ABP-gated K⁺ channel, SthK of 430 aas and 6 TMSs in a 2 + 2 + 1 + P-loop +1 TMS arrangement. This channel and others have been studied by high-speed atomic force microscopy (HS-AFM) which has made it possible to characterized the conformational dynamics of single unlabeled transmembrane channels and transporters (Heath and Scheuring 2019).

TWIK-1 (KCNK1, HOHO1, KCNO1) inward rectifier K⁺ channel (Enyedi and Czirják, 2010) expressed in the distal nephron segments (Orias et al. 1997).  Lipid tails from both the upper and lower leaflets can partially penetrate into the pore (Aryal et al. 2015). The lipid tails do not sterically occlude the pore, but there is an inverse correlation between the presence of water within the hydrophobic barrier and the number of lipids tails within the lining of the pore (Aryal et al. 2015).

TREK-1 (KCNK2) K⁺ channel subunit (Regulated by group 1 metabotropic glutamate receptors and by diacylglycerols and phosphatidic acids) (Chemin et al., 2003). TREK-1, TREK-2 and TRAAK are all regulated by lysophosphatidic acid, converting these mechano-gated, pH voltage-sensitive channels into leak conductances (Chemin et al., 2005). The mammalian K2P2.1 potassium channel (TREK-1, KCNK2) is highly expressed in excitable tissues, where it plays a key role in the cellular mechanisms of neuroprotection, anaesthesia, pain perception and depression (Cohen et at., 2008). Alternative translation initiation in rat brain yields K2P2.1 potassium channels permeable to sodium (Thomas et al. 2008). The crystal structure of the human 2-pore domain K⁺ channel, K2P1 has been solved (Miller and Long, 2012). Multiple modalities converge on a common gate to control K2P channel function (Bagriantsev et al., 2011).  TREK-1 mediates fast and slow glutamate release in astrocytes upon GPCR activation (Woo et al. 2012). It is a mechanosensitive K⁺ channel, present in rat bladder myocytes, which is activated by swelling and arachidonic acid (Fukasaku et al. 2016). The M2-hinges of TREK-1 and TREK-2 channels control their macroscopic current, subcellular localization and gating (Zhuo et al. 2017). The human ortholog has acc # O95069 and has an additional N-terminal 15 aas. BL-1249, a compound from the fenamate class of nonsteroidal anti-inflammatory drugs, is known to activate K2P2.1(TREK-1), the founding member of the thermo- and mechanosensitive TREK subfamily (Pope et al. 2018). Spadin and arachidonic acid, are known to suppress and activate TREK-1 channels, respectively (Pappa et al. 2020). Membrane phospholipids control gating of the mechanosensitive potassium leak channel, TREK1 (Schmidpeter et al. 2023). A photoswitchable inhibitor of TREK channels controls pain in wild-type intact freely moving animals (Landra-Willm et al. 2023). TREK-1 is an anesthetic-sensitive K⁺ channel (Spencer et al. 2023).  Covalent chemogenetic K_2P channel activators have been developed (Deal et al. 2024).  K_2P potassium channels regulate excitability by affecting the cellular resting membrane potential in the brain, cardiovascular system, immune cells, and sensory organs. They are important in anesthesia, arrhythmia, pain, hypertension, sleep, and migraine headaches.  CATKLAMP (covalent activation of TREK family K⁺ channels to clamp membrane potential) leverages the discovery of a K_2P modulator pocket site that reacts with electrophile-bearing derivatives of a TREK subfamily small-molecule activator, ML335, to activate the channel irreversibly. Deal et al. 2024 showed that CATKLAMP can be used to probe fundamental aspects of K_2P function, as a switch to silence neuronal firing, and is applicable to all TREK subfamily members.

pH-dependent, voltage-insensitive, background potassium channel protein involved in maintaining the membrane potential, KCNK9, K2P9.1 or TASK3 (TASK-3) of 374 aas  (Huang et al. 2011). Terbinafine is a selective activator of TASK3 (Wright et al. 2017). The response of the tandem pore potassium channel TASK-3 to voltage involves gating at the cytoplasmic mouth (Ashmole et al., 2009).  TASK-3 is involved in several physiological and pathological processes including sleep/wake control, cognition and epilepsy (Tian et al. 2019). N-(2-((4-nitro-2-(trifluoromethyl)phenyl)amino)ethyl)benzamide (NPBA) is an activator (Tian et al. 2019). KCC2 regulates neuronal excitability and hippocampal activity via interaction with Task-3 channels (Goutierre et al. 2019). A  biguanide compound, CHET3, is a highly selective allosteric activator, and TASK-3 is a druggable target for treating pain (Liao et al. 2019). This channel may be present in mitochondria (Parrasia et al. 2019). Differential hydroxymethylation levels in the DNA of patient-derived neural stem cells implicated altered cortical development in bipolar disorder syndrome possibly altering KCNK9 expression (Kumar et al. 2023).

KCNK3 K⁺ channel (TASK1, OAT1, TBAK1) (the K⁺ leak conductance). TASK1 and 3 may play roles in nontumorigenic primary hyperaldosteronism (Davies et al., 2008).  KCNK3/9/15 expression limits membrane depolarization and depolarization-induced secretion at least in part by maintaining intracellular K⁺ (Huang et al. 2011). TWIK-related acid-sensitive potassium (TASK) channels, members of the two pore domain potassium (K2P) channel family, are found in neurons, cardiomyocytes and vascular smooth muscle cells, where they are involved in the regulation of heart rate, pulmonary artery tone, sleep/wake cycles and responses to volatile anaesthetics (Rödström et al. 2020). K2P channels regulate the resting membrane potential, providing background K⁺ currents controlled by numerous physiological stimuli. Unlike other K2P channels, TASK channels are able to bind inhibitors with high affinity, exceptional selectivity and very slow compound washout rates. In general, potassium channels have an intramembrane vestibule with a selectivity filter situated above and a gate with four parallel helices located below, but the K2P channels studied so far all lack a lower gate. Rödström et al. 2020 presented the X-ray crystal structure of TASK-1, and showed that it contains a lower gate designated 'X-gate', created by interaction of the two crossed C-terminal M4 transmembrane helices at the vestibule entrance. This structure is formed by six residues ((243)VLRFMT(248)) that are essential for responses to volatile anaesthetics, neurotransmitters and G-protein-coupled receptors. Mutations within the X-gate and the surrounding regions affect both the channel-open probability and the activation of the channel by anaesthetics. Structures of TASK-1 bound to two high-affinity inhibitors showed that both compounds bind below the selectivity filter and are trapped in the vestibule by the X-gate, which explains their exceptionally low washout rates (Rödström et al. 2020). TWIK-related acid-sensitive K⁺ channel 2 promotes renal fibrosis by inducing cell-cycle arrest (Zhang et al. 2022). KCNK3 dysfunction plays a role in dasatinib-associated pulmonary arterial hypertension and endothelial cell dysfunction (Ribeuz et al. 2024).

Neuronal 2-P (4 TMS) domain K⁺ membrane tension-gated channel, TRAAK (stimulated by arachidonic acid and polyunsaturated fatty acids (Fink et al., 1998). The crystal structures of conductive and nonconductive human K2P TRAAK K⁺ channel has been solved (Brohawn et al., 2012; Brohawn et al. 2014). Regulated by mechanical deformation of the membrane and temperature as well as polyunsaturated fatty acids (Brohawn et al., 2012). Multiple modalities converge on a common gate to control K2P channel function (Bagriantsev et al., 2011).  In the non-conductive state, a lipid acyl chain accesses the channel cavity through a 5 Å-wide lateral opening in the membrane inner leaflet and physically blocks ion passage. In the conductive state, rotation of transmembrane helix 4 about a central hinge seals the intramembrane opening, preventing lipid block of the cavity and permitting ion entry. Additional rotation of a membrane interacting TM2-TM3 segment, unique to mechanosensitive K2Ps, against TM4 may further stabilize the conductive conformation. Comparison of the structures provodes a biophysical explanation for TRAAK mechanosensitivity--an expansion in cross-sectional area up to 2.7 nm² in the conductive state is expected to create a membrane-tension-dependent energy difference between conformations that promotes force activation (Brohawn et al. 2014).  TM helix straightening and buckling may underlie channel activation (Lolicato et al. 2014). A lipid chain blocks the conducting path in the closed state (Rasmussen 2016).

Outward rectifying mechanosensitive 2-P (4 TMS) domain K⁺ channel, TREK-2 (KCNKA; KCNK10; K2P10). Activated by membrane stretch, acidic pH, arachidonic acid and unsaturated fatty acids.  Dong et al. 2015 described crystal structures of the human TREK-2 channel (up to 3.4 angstrom resolution) in two conformations and in complex with norfluoxetine, the active metabolite of fluoxetine (Prozac) and a state-dependent blocker of TREK channels. Norfluoxetine binds within intramembrane fenestrations found in only one of these two conformations. Channel activation by arachidonic acid and mechanical stretch involves conversion between these states through movement of the pore-lining helices. This provides an explanation for TREK channel mechanosensitivity, regulation by diverse stimuli, and possible off-target effects of the serotonin reuptake inhibitor Prozac (Dong et al. 2015).  The unique gating properties of TREK-2 and the mechanisms by which extracellular and intracellular stimuli harness pore gating allosterically have been studied (Zhuo et al. 2016). TREK-2 moves from the "down" to the "up" conformation in direct response to membrane stretch. Aryal et al. 2017 showed how state-dependent interactions with lipids affect the movement of TREK-2, and how stretch influences both the inner pore and selectivity filter. They also demonstrated why direct pore block by lipid tails does not represent theprincipal mechanism of mechanogating (Aryal et al. 2017). The M2-hinges of TREK-1 and TREK-2 channels control their macroscopic current, subcellular localization and gating (Zhuo et al. 2017). TREK-2 responds to a diverse range of stimuli. Two states, "up" and "down", are known from x-ray structural crystallographic studies and have been suggested to differ in conductance. Brennecke and de Groot 2018 found that the down state is less conductive than the up state. The introduction of membrane stretch in the simulations shifts the state of the channel toward an up configuration. Membrane pressure changes the conformation of the transmembrane helices directly and consequently also influences the channel conductance (Brennecke and de Groot 2018). 3-d structures are known (PDB 4XDJ_!-D). Phosphatidyl-(3,5)-bisphosphate (PI(3,5)P2) activates (Kirsch et al. 2018).

AMPA-selective glutamate ionotropic channel receptor (GIC; AMPAR), kainate-subtype, GluR-K1; GluR1; GluR-A; GluA1; Gria1 of 906 aas (preferentially monovalent cation selective). A mature complex contains GluR1, TARPs, and PSD-95 (Fukata et al. 2005). The receptor contributes to amygdala-dependent emotional learning and fear conditioning (Humeau et al., 2007). Transmembrane AMPAR regulatory protein (TARP) gamma-7 (TC#8.A.16.2.5) selectively enhances the synaptic expression of Ca²⁺-permeable (CP-AMPARs) and suppresses calcium-impermeable (CI-AMPAR) activities (Studniarczyk et al. 2013).  Thus, TARPs modulate the pharmacology and gating of AMPA-type glutamate receptors (Soto et al. 2014).  TARPs interact with the N-terminal domain of the AMPAR and control channel gating; residues in the receptor and the TARP involved in this interaction have been identified (Cais et al. 2014).  The auxilary protein, Shisa9 or CKAMP44 (UniProt acc# B4DS77), has a C-terminal PDZ domain that allows interaction with scaffolding proteins and AMPA glutamate receptors (Karataeva et al. 2014).  The transmembrane domain alone can tetramerize (Gan et al. 2016). The most potent and well-tolerated AMPA receptor inhibitors, used to treat epilepsy, act via a noncompetitive mechanism.  The crystal structures of the rat AMPA-subtype GluA2 receptor in complex with three noncompetitive inhibitors have been solved. The inhibitors bind to a binding site, completely conserved between rat and human, at the interface between the ion channel and linkers connecting it to the ligand-binding domains (Yelshanskaya et al. 2016). The endogenous neuropeptide, cyclopropylglycine, at a physiological concentration of 1 μM, enhances the transmembrane AMPA currents in rat cerebellar Purkinje cells (Gudasheva et al. 2016). The energetics of glutamate binding have been estimated (Yu and Lau 2017). The TMEM108 protein (Q6UXF1 of 575 aas and 2 TMSs, N- and C-terminal, is required for surface expression of AMPA receptors (Jiao et al. 2017). CERC-611 is a selective antagonist of AMPA receptors containing transmembrane AMPA receptor regulatory protein (TARP; TC# 8.A.16) gamma-8 (Witkin et al. 2017). Drug effects, regulatory protein modulators and positive allosteric modulators have been reviewed (Fu et al. 2019). Herguedas et al. 2019 presented a cryo-EM structure of the heteromeric GluA1/2 receptor associated with two transmembrane AMPAR regulatory protein (TARP) gamma8 auxiliary subunits, the principal AMPAR complex at hippocampal synapses.  The native heterotetrameric AMPA-R adopts various conformations, which reflect a variable separation of the two dimeric extracellular amino-terminal domains; members of the stargazin/TARP family of transmembrane proteins co-purify with AMPA-Rs and contribute to the density representing the transmembrane region of the complex. Glutamate and cyclothiazide altered the conformational equilibrium of the channel complex, suggesting that desensitization is related to separation of the N-terminal domains (Nakagawa et al. 2005). Positive allosteric modulators (PAMs) of AMPA receptors boost cognitive performance in clinical studies, and mibampator and BIIB104 discriminate between AMPARs complexed with distinct TARPs, and particularly those with lower stargazin/gamma2 efficacy such as BIIB104 (Ishii et al. 2020). Yelshanskaya et al. 2020 identified trans-4-butylcyclohexane carboxylic acid (4-BCCA) binding sites in the transmembrane domain of AMPA receptors, at the lateral portals formed by TMSs M1-M4. At this binding site, 4-BCCA is very dynamic, assumes multiple poses and can enter the ion channel pore. Cannabidiol inhibits febrile seizure by modulating AMPA receptor kinetics through its interaction with the N-terminal domain of GluA1/GluA2 (Yu et al. 2020). Inhibition of AMPA receptors (AMPARs, e.g., TC# 1.A.10.1.1) containing transmembrane AMPAR regulatory protein gamma-8 (TC# 8.A.61.1.10) with JNJ-55511118 (TC#8.A.179.1.1) shows preclinical efficacy in reducing chronic repetitive alcohol self-administration (Hoffman et al. 2021). Mechanisms underlying TARP modulation of the GluA1/2-gamma8 AMPA receptor have been studied (Herguedas et al. 2022). L-Glutamate is the main excitatory neurotransmitter in the central nervous system (CNS). Its associated receptors, localized on neuronal and non-neuronal cells, mediate rapid excitatory synaptic transmission in the CNS and regulate a wide range of processes in the brain, spinal cord, retina, and peripheral nervous system. Glutamate receptors selective to alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) also play an important role in numerous neurological disorders.  Golubeva et al. 2022 examined the structural diversity of chemotypes of agonists, competitive AMPA receptor antagonists, positive and negative allosteric modulators, TARP-dependent allosteric modulators, ion channel blockers ans their binding sites.

The homo- and heteromeric glutamate receptor, GLR3.3/3.4 (Desensitized in 3 patterns: (1) by Glu alone; (2) by Ala, Cys, Glu or Gly; (3) by Ala, Cys, Glu, Gly, Ser or Asn (Stephens et al., 2008). A regulatory mechanism underlies Ca²⁺ homeostasis by sorting and activation of AtGLRs by AtCNIHs (see for example, 8.A.61.1.9) (Wudick et al. 2018).  May be responsible in part for Cd²⁺ uptake (Chen et al. 2018). GLR3.3 and GLC3.6 (TC# 1.A.10.1.24) (but not GLR3.4) play different roles in nervous system-like signaling in plant defense by a mechanism that differs substantially from that in animals (Toyota et al. 2018). Members of the banana GLR gene family have been identified, and expression analyses in response to low temperature and abscisic acid/methyl jasmonate concentrations have been reported (Luo et al. 2023).

GriK2; GluK2; GluR6 glutamate receptor, ionotropic kainate 2. The 3-d structure is known (2XXY_A). The domain organization and function have been analyzed by Das et al. (2010).  Two auxiliary subunits, Neto1 and Neto2 (Neuropilin and tolloid-like proteins) alter the trafficking, channel kinetics and pharmacology of the receptors (Howe 2014).  They reduce inward rectification without altering calcium permeability (Fisher and Mott 2012). Interactions between the pore helix (M2) and adjacent segments of the transmembrane inner (M3) and outer (M1) helices may be involved in gating (Lopez et al. 2013). Mutations in the human GRIK2 (GLUR6) cause moderate-to-severe nonsyndromic autosomal recessive mental retardation (Motazacker et al. 2007). Kainate receptors regulate KCC2 (TC# 1.A.10.1.11) expression in the hippocampus (Pressey et al. 2017). GluR6, carrying the pore loop plus adjacent transmembrane domains of the prokaryotic, glutamate-gated, K⁺-selective GluR0 (TC# 1.A.10.2.1), adopted several electrophysiological properties of the donor pore upon pore transplantation (Hoffmann et al. 2006). Clustered mutations in the GRIK2 kainate receptor subunit gene underlie diverse neurodevelopmental disorders (Stolz et al. 2021). Concanavalin A modulation of kainate receptor function is mediated by a shift in the conformation of the kainate receptor toward a tightly packed extracellular domain (Gonzalez et al. 2021). Partial agonism in heteromeric GLUK2/GLUK5 kainate receptor has been documented, and partial agonism observed with AMPA binding is mediated primarily due to differences in the GluK2 subunit, highlighting the distinct contributions of the subunits towards activation (Paudyal et al. 2023).

The NMDA receptor. The crystal structure of the N-terminal domains (GluN1 and GluN2) have been determined (PDB#3QEL; Talukder and Wollmuth, 2011). The ligand-specific deactivation time courses of GluN1/GluN2D NMDA receptors have been measured (Vance et al., 2011).  NMDA receptors are Hebbian-like coincidence detectors, requiring binding of glycine and glutamate in combination with the relief of voltage-dependent magnesium block to open an ion conductive pore. Lee et al. 2014 presented X-ray structures of the Xenopus laevis GluN1-GluN2B NMDA receptor with the allosteric inhibitor, Ro25-6981, partial agonists and the ion channel blocker, MK-801. Receptor subunits are arranged in a 1-2-1-2 fashion, demonstrating extensive interactions between the amino-terminal and ligand-binding domains. The 3-TMS transmembrane domains harbour a closed-blocked ion channel, a pyramidal central vestibule lined by residues implicated in binding ion channel blockers and magnesium, and a approximately twofold symmetric arrangement of ion channel pore loops. GRIN2D mediates developmental and epileptic encephalopathy (XiangWei et al. 2019).

Glu2 AMPA receptor (GluR-2; GluR2-flop; CX614; GluA2).  The 3-d structure is known at 3.6 Å resolution.  It shows a 4-fold axis of symmetry in the transmembrane domain, and a 2-fold axis of symmetry overall, although it is a homotetramer (Sobolevsky et al. 2009). A structure showing an agoniar-bound form of the rat GluA2 receptor revealed conformational changes that occur during gating (Yelshanskaya et al. 2014). GluR2 interacts directly with β3 integrin (Pozo et al., 2012).  In general, integrin receptors form macromolelcular complexes with ion channels (Becchetti et al. 2010).  TARPS are required for AMP receptor function and trafficking, but seven other auxiliary subunits have also been identified (Sumioka 2013). For example, AMPA receptors are regulated by S-SCAM through TARPs (Danielson et al. 2012).  The C-terminal domains of various TARPs (TC#8.A.16.2) play direct roles in the regulation of GluRs (Sager et al. 2011).  Whole-genome analyses have linked multiple TARP loci to childhood epilepsy, schizophrenia and bipolar disorders (Kato et al. 2010). Thus, TARPs emerge as vital components of excitatory synapses that participate both in signal transduction and in neuropsychiatric disorders. The architecture of a fully occupied GluR2-TARP complex has been illucidated by cryoEM, showing the homomeric GluA2 AMPA receptor saturated with TARP Υ2 subunits, showing how the TARPs are arranged with four-fold symmetry around the ion channel domain, making extensive interactions with the M1, M2 and M4 TMSs (Zhao et al. 2016). The binding mode and sites for prototypical negative allosteric modulators at the GluA2 AMPA receptor revealing new details of the molecular basis of molulator binding and mechanisms of action (Stenum-Berg et al. 2019). Drug effects, regulatory protein modulators and positive allosteric modulators have been reviewed (Fu et al. 2019). TARP γ2 converts the desensitized state to the high-conductance state which exhibits tighter coupling between subunits in the extracellular parts of the receptor (Carrillo et al. 2020).

Glutamate receptor 4, GIC, AMPA-subtype, GluR4, GRIA4 or GluR-D (preferentially monovalent cation selective). Binding of the excitatory neurotransmitter, L-glutamate, induces a conformation change, leading to the opening of the cation channel, thereby converting the chemical signal to an electrical impulse. The receptor then desensitizes rapidly and enters a transient inactive state, characterized by the presence of bound agonist. In the presence of CACNG4, CACNG7 or CACNG8, GluR4 shows resensitization characterized by a delayed accumulation of current flux upon continued application of glutamate (Gill et al. 2008; Birdsey-Benson et al. 2010). De novo variants in GRIA4 lead to intellectual disability with or without seizures, gait abnormalities, problems of social behavior, and other variable features (Martin et al. 2017).

Heteromeric ionotropic NMDA receptor (NMDAR) consisting of two subunits, GluN1 (938 aas) and GluN2A (1464 aas).  Positions of the Mg²⁺ and Ca²⁺ ions in the ion channel divalent cation binding site have been proposed, and differences in the structural and dynamic behavior of the channel proteins in the presence of Mg²⁺ or Ca²⁺ have been analyzed (Mesbahi-Vasey et al. 2017). GRIN variants in receptor M2 channel pore-forming loop are associated with neurological diseases (Li et al. 2019). Disease-associated variants have revealed mechanistic aspect of the NMDA receptor (Amin et al. 2021). Cross-subunit interactions that stabilize open states mediate gating in NMDA receptors (Iacobucci et al. 2021). The gating mechanism and a modulatory niche of human GluN1-GluN2A NMDA receptors have been reported (Wang et al. 2021). GluN2A and GluN2B NMDA receptors apparently use distinct allosteric routes (Tian et al. 2021). A negative allosteric modulatory site in the GluN1 M4 determines the efficiency of neurosteroid modulation (Langer et al. 2021). Excitatory signaling mediated by NMDAR is critical for brain development and function, as well as for neurological diseases and disorders. Channel blockers of NMDARs can be used for treating depression, Alzheimer's disease, and epilepsy. Chou et al. 2022 monitored the binding of three clinically important channel blockers: phencyclidine, ketamine, and memantine in GluN1-2B NMDARs at local resolutions of 2.5-3.5 Å around the binding site. The channel blockers form interactions with pore-lining residues, which control mostly off-speeds but not on-speeds (Chou et al. 2022).  NMDAR channel blockers include MK-801, phencyclidine, ketamine, and the Alzheimer's disease drug memantine, can bind and unbind only when the NMDAR channel is open. NMDAR channel blockers can enter the channel through two routes: the well-known hydrophilic path from extracellular solution to channel through the open channel gate, and also a hydrophobic path from plasma membrane to channel through a gated fenestration (Wilcox et al. 2022). Pregnane-based steroids are  positive NMDA receptor modulators that may compensate for the effect of loss-of-function disease-associated GRIN mutations (Kysilov et al. 2022). The NMDA receptor C-terminal domain signals in development, maturity, and disease (Haddow et al. 2022). Blood tissue Plasminogen Activator (tPA) of liver origin contributes to neurovascular coupling involving brain endothelial N-Methyl-D-Aspartate (NMDA) receptors (Furon et al. 2023). Two gates mediate NMDA receptor activity and are under subunit-specific regulation (Amin et al. 2023). One of the main molecular mechanisms of ketamine action is the blockage of NMDA-activated glutamate receptors (Pochwat 2022). The S1-M1 linker of the NMDA receptor controls channel opening (Xie et al. 2023). Binding and dynamics demonstrated the destabilization of ligand binding for the S688Y mutation in the NMDA receptor GluN1 subunit (Chen et al. 2023). The functional effects of disease-associated NMDA receptor variants have been reviewed (Moody et al. 2023). Co-activation of NMDAR and mGluRs controls protein nanoparticle-induced osmotic pressure in neurotoxic edema (Zheng et al. 2023).  Disease-associated nonsense and frame-shift variants resulting in the truncation of the GluN2A or GluN2B C-terminal domain decreases NMDAR surface expression and reduces potentiating effects of neurosteroids (Kysilov et al. 2024).  De novo GRIN variants in the M3 helix associated with neurological disorders control channel gating of the NMDA receptor (Xu et al. 2024).

AMPA glutamate receptor 3 (GluR3, GluA3. GRIA3. LLUR3. GLURC) (non-selective monovalent cation channel and Ca²⁺  channel) (Ayalon et al., 2005; Midgett et al., 2012). Regulated by AMPA receptor regulatory proteins (TARPs) including stargazin and CNIH auxiliary subunits (Kim et al., 2010; Straub and Tomita, 2011; Jackson and Nicoll, 2011; Bats et al., 2012; Rigby et al. 2015). The domain architecture of a calcium-permeable AMPA receptor in a ligand-free conformation has been solved (Midgett et al., 2012). The TARP, stargazin, is elevated in the somatosensory cortex of Genetic Absence Epilepsy Rats (Kennard et al. 2011). TARPs alter the conformation of pore-forming subunits and thereby affect antagonist interactions (Cokić and Stein 2008).  The structural basis of AMPAR regulation by TARP gamma2, or stargazin (STZ) involves variable interaction stoichiometries of the AMPAR-TARP complex, with one or two TARP molecules binding one tetrameric AMPAR (Twomey et al. 2016).  The ion channel extracellular collar plays a role in gating and represents a hub for powerful allosteric modulation of AMPA receptor function (Yelshanskaya et al. 2017). The A653T mutation stabilizes the closed configuration of the channel and affects duration of sleep and awake periods in both humans and mice (Davies et al. 2017). The tetramer exhibits 4 distinct conductase leves due to independent subunit activation.  Perampanel is an anticonvulsant drug that regulates gating (Yuan et al. 2019). Tetramerization of the AMPA receptor glutamate-gated ion channel is regulated by auxiliary subunits (Certain et al. 2023). A synopsis of multitarget therapeutic effects of anesthetics on depression has been published (Wu and Xu 2023).

The homomeric cation channel/glutamate receptor/kainate 1, GluR5, GluK1, GRIK1 of 918 aas and  (weakly responsive to glutamate) (expressed in the developing nervous system) (Bettler et al., 1990).  The 3-d structures of the protein have been determined with agonists and antagonists.  The agonist, domoic acid, stabilizes the ligand-binding core of the iGluR5 complex in a conformation that is 11 degrees more open than the conformation observed when the full agonist, (S)-glutamate, is bound (Hald et al. 2007). Kainate receptors regulate KCC2 expression in the hippocampus (Pressey et al. 2017). GluR5/ERK signaling is regulated by the phosphorylation and function of the glycine receptor alpha1ins subunit (TC# 9.A.14.3.4) in the spinal dorsal horn of mice (Zhang et al. 2019). The human ortholog is 918 aas long and 97% identical to the rat homolog. (-)-Arctigenin and a series of new analogues are AMPA and kainate receptor antagonists of human homomeric GluA1 and GluK2 receptors (Rečnik et al. 2021).

The heteromeric monovalent cation/Ca²⁺ channel/glutamate (NMDA) receptor NMDAR1/NMDAR2A/NMDAR2B/NMDAR2C) (Monyer et al., 1992). Note: NR2B is the same as NR3, GluN2A, GRIN2A or subunit epsilon (Schüler et al., 2008). Mediates voltage- and Mg²⁺-dependent control of Na⁺ and Ca²⁺ permeability (Yang et al., 2010).  Mutations in the subunit, GRIN1, a 1464 aa protein, identified in patients with early-onset epileptic encephalopathy and profound developmental delay, are located in the transmembrane domain and the linker region between the ligand-binding and transmembrane domains (Yuan et al. 2014; Ohba et al. 2015).  Karakas and Furukawa 2014 determined the crystal structure of the heterotetrameric GluN1-GluN2B NMDA receptor ion channel at 4 Å resolution. The receptor is arranged as a dimer of GluN1-GluN2B heterodimers with the twofold symmetry axis running through the entire molecule composed of an amino terminal domain, a ligand-binding domain, and a transmembrane domain.  The GluN2 subunit regulates synaptic trafficking of AMPA in the neonatal mouse brain (Hamada et al. 2014).  GRIN1 and GRIN2A mutations are associated with severe intellectual disability with cortical visual impairment, epilepsy and oculomotor and movement disorders being discriminating phenotypic features (Lemke et al. 2016; Chen et al. 2017).The cryoEM structure of a triheteromeric receptor including GluN1 (glycine binding), GluN2A and GluN2B (both glutamate binding) has been solved with and without a GluN2B allosteric antagonist, Ro 25-6981 (Lü et al. 2017). Ogden et al. 2017 implicated the pre-M1 region in gating, providing insight into how different subunits contribute to gating, and suggesting that mutations in the pre-M1 helix, such as those that cause epilepsy and developmental delays, can compromise neuronal health. The severity of GRIN2A (Glu2A)-related disorders can be predicted based on the positions of the mutations in the encoding gene (Strehlow et al. 2019). Knock-in mice expressing an ethanol-resistant GluN2A NMDA receptor subunit show altered responses to ethanol (Zamudio et al. 2019). Results of McDaniel et al. 2020 revealed the role of the pre-M1 helix in channel gating, implicated the surrounding amino acid environment in this mechanism, and suggested unique subunit-specific contributions of pre-M1 helices to GluN1 and GluN2 gating. The human ortholog is 998.5% identical. An autism-associated mutation in GluN2B prevents NMDA receptor trafficking and interferes with dendrite growth (Sceniak et al. 2019). The binding of calcium-calmodulin to the C-terminus of GluN1 has long range allosteric effects on the extracellular segments of the receptor that may contribute to the calcium-dependent inactivation (Bhatia et al. 2020). GluN1 interacts with PCDH7 (O60245) to regulate dendritic spine morphology and synaptic function (Wang et al. 2020).Pluripotential GluN1 (NMDA NR1) functions in cellular nuclei in pain/nociception (McNearney and Westlund 2023).

Glutamate-gated ionotropic K⁺ channel receptor, GluR0 (5TMSs). X-ray structures are available (PDB: 1IIT) (Lee et al. 2005; Lee et al. 2008)  GluR6 (TC# 1.A.10.1.11), carrying the pore loop plus adjacent transmembrane domains of this prokaryotic, glutamate-gated, K⁺-selective GluR0, adopted several electrophysiological properties of the donor pore upon pore transplantation (Hoffmann et al. 2006).

The mixed lineage kinase domain-like (MLKL) protein of 471 aas and 5 TMSs; forms Mg²⁺-selective channels in the presence of Na⁺ and K⁺ (Xia et al. 2016).

The Transmembrane and coiled-coil domains protein 1 (TMCO1, TMCC4, PNAS-10, PNAS-136, UNQ155) of 188 aas and 3 TMSs. It is an ER transmembrane protein that actively prevents Ca²⁺ stores from overfilling, acting as a "Cacium Load-activated Calcium channel" or ""CLAC"" channel. TMCO1 undergoes reversible homotetramerization in response to ER Ca²⁺ overloading and disassembly upon Ca²⁺ depletion to form a Ca²⁺-selective ion channel as demonstrated in liposomes (Wang et al. 2016). TMCO1 knockout mice reproduce the main clinical features of human cerebrofaciothoracic (CFT) dysplasia spectrum, a developmental disorder linked to TMCO1 dysfunction, and exhibit severe mishandling of ER Ca²⁺ in cells (Alanay et al. 2014). Thus, TMCO1 provides a protective mechanism to prevent overfilling of ER stores with calcium ions (Wang et al. 2016). It regulates Ca²⁺ stores in granulosa cells (Sun et al. 2018).  TMCO1-mediated Ca²⁺ leak underlies osteoblast functions via CaMKII signaling (Li et al. 2019). The TMCO1 gene is a tumor suppressor in urinary bladder urothelial carcinomas (UBUC). TMCO1 dysregulates cell-cycle progression via suppression of the AKT pathway, and S60 of the TMCO1 protein is crucial for its tumor-suppressor roles (Li et al. 2017). Batchelor-Regan et al. 2021 published a short review about the clinical and scientific advances made with TMCO1. Ca²⁺ homeostasis maintained by TMCO1 underlies corpus callosum development via ERK signaling (Yang et al. 2022). A mechanism of metformin action, restoring cellular ER homeostasis, enabled the development of a nanocarrier-mediated ER targeting strategy for remodeling diabetic periodontal tissue (Zhong et al. 2022). TMCO1 regulates cell proliferation, metastasis and EMT signaling through CALR, promoting ovarian cancer progression and cisplatin resistance (Sun et al. 2024).  TMCO1 is a crucial regulator of ovarian cancer progression, influencing VDAC1 through CALR and impacting diverse cellular processes (Sun et al. 2024).

Pore-forming Hemoglobin-α of 142 aas (Morrill and Kostellow 2016).

Pore-forming hemoglobin-β of 147 aas (Morrill and Kostellow 2016).

Pore-forming myoglobin of 154 aas (Morrill and Kostellow 2016).

Pore-forming neuroglobin of 151 aas (Morrill and Kostellow 2016).

Pore-forming cytoglobin of 154 aas (Morrill and Kostellow 2016).

Fibroblast growth factor 2, FGF2, of 288 aas.  Forms oligomeric pores in the plasma membrane, and these are involved in its secretion to the external medium where it exerts its action (La Venuta et al. 2016; see family description). The ATP1A1 (TC# 3.A.3.1.1) serves as the receptor for membrane insertion (Zacherl et al. 2015).

Fibroblast growth factor 23, FGF23 of 251 aas.  Functions through FGF receptor 4 to regulate the level of the calcium/inorganic cation channel, TrpC6 or TRP6 (TC# 1.A.4.1.5) (Smith et al. 2017).

Interleukin-1 alpha, IL1α or IL1A, of 271 aas and 0 TMSs. Regulates gap junctional communication in Sertoli cells, which is critical for sertoli cell barrier/blood-testis barrier (BTB) restructuring (Chojnacka et al. 2016). 

Interleukin 1β, IL1β, IL1beta, IL1B, IL1F2, of 269 aas.  Its release depends on its insertion into the membrane with the formation of a transmembrane pore (Martín-Sánchez et al. 2016). IL-1beta is generated by macrophages upon activation of intracellular NLRP3 (NOD-like, leucine rich repeat domains, and pyrin domain-containing protein 3), part of the functional NLRP3 inflammasome complex that detects pathogenic microorganisms and stressors, while neutrophils are enhanced by increasing levels of IL-1beta (Yaqinuddin and Kashir 2020).

Ammonia transporter and regulatory sensor, AmtB (Blauwkamp and Ninfa, 2003; Khademi et al., 2004).  It has a cleavable N-terminal signal peptide, and while Amt proteins in Gram-negative bacteria appear to utilize a signal peptide, the homologous proteins in Gram-positive organisms do not (Thornton et al. 2006).

Rhesus (Rh) type C glycoprotein NH₃/NH₄⁺ transporter, RhCG (also called tumor-related protein DRC2) (Bakouh et al., 2004; Worrell et al., 2007). Zidi-Yahiaoui et al. (2009) have described characteristics of the pore/vestibule. The structure is known to 2.1 Å resolution (Gruswitz et al., 2010). Each monomer contains 12 transmembrane helices, one more than in the bacterial homologs. Reconstituted into proteoliposomes, RhCG conducts NH₃ to raise the internal pH. Models of the erythrocyte Rh complex based on the RhCG structure suggest that the erythrocytic Rh complex is composed of stochastically assembled heterotrimers of RhAG, RhD, and RhCE (Gruswitz et al., 2010). Rh proteins also transport CO₂ (Michenkova et al. 2021).

The RH50 NH₃ channel (most like human Rh proteins TC# 1.A.11.4.1 and 2; 36-38% identity) (Cherif-Zahar et al., 2007). The Rh CO₂ channel protein (3-D structure ± CO₂ available) (3B9Z_A; 3B9Y_A) (Li et al., 2007; Lupo et al., 2007) (also transports methyl ammonia) (Weidinger et al., 2007).

The OTOP3 protopn channel protein of 681 aas and 12 TMSs. The cryo-EM structure along with functional characteristics have been described (Chen et al. 2019). XtOTOP3 forms a homodimer with each subunit containing 12 transmembrane helices that can be divided into two structurally homologous halves; each half assembles as an alpha-helical barrel that could serve as a proton conduction pore.

Kidney metal (Mg²⁺) transporter, Cyclin (CNN) M2 isoform CRA_b (CNNM2). Defects cause hypomagnesemia. It has an extracellular N-terminus, an N-terminal TMS, a hydrophilic domain followed by 4 TMSs, another hydrophilic domain, and an intracellular C-terminus (de Baaij et al., 2012). CNNM2a forms heterodimers with the smaller isoform CNNM2b.  The human splice variant 1 of CNNM2 (ACDP2; Q9H8M5) is a Mg²⁺ transporter (Brandao et al. 2012).  The Bateman module is involved in AMP binding and Mg²⁺ sensing, and their binding causes a conformational change in the CBS module, transmitted to the transmembrane domain (Corral-Rodríguez et al. 2014). It may be able to transport divalent metal cations, Mg²⁺, Co²⁺, Mn²⁺, Sr²⁺, Ba²⁺, Cu²⁺, Fe²⁺ and monvalent cation, Na⁺. In prokaryotes, homologs are CorB/C.

Metal transporter CNNM3 (Ancient conserved domain-containing protein 3) (mACDP3) (Cyclin-M3) of 713 aas and probably 5 TMSs with one N-terminal, and four together in the first half of the protein (Chen et al. 2018). See the family description for the domain order of the CNNM proteins.

Metal transporter CNNM3 (Ancient conserved domain-containing protein 3) (Cyclin-M3).  As of 2018, the function of this protein as a Mg²⁺ transporter is under debate (Schäffers et al. 2018).

Metal transporter CNNM4 (Ancient conserved domain-containing protein 4) (Cyclin-M4).  As of 2018, the function of this protein as a Mg²⁺ transporter was under debate (Schäffers et al. 2018).

Putative Mg²⁺ exporter of 875 aas and 5 TMSs, CNNM2 or ACDP2 (Chen et al. 2018). The bacterial CorC is involved in resistance to antibiotic exposure and to the survival of pathogenic microorganisms in their host environments. CorC possesses a cytoplasmic region containing the (regulatory ?) ATP-binding site (Huang et al. 2021). An inhibitor, IGN95a, targets the ATP-binding site and blocks both ATP binding and Mg²⁺ export. The cytoplasmic domain structure in complex with IGN95a was determined (Huang et al. 2021). With ATP bound to the cytoplasmic domain, the conformational equilibrium of CorC shifts toward the inward-facing state of the transmembrane domain (Huang et al. 2021). These considerations suggest that CorC may be an ATP-driven Mg²⁺ efflux porter, and if so, the family belongs in TC sub-class, 3.A. CorC homologs may be able to export Mg²⁺, Co²⁺, Mn²⁺, Sr²⁺, Ba²⁺, Cu²⁺ and Fe²⁺.

Cyclotide, kalata-B1, kB1. Targets membranes in a process that depends on lipid interactions; hemolytic; can disrupt HIV membranes (Henriques et al., 2011).

The Varv peptide A/Kalata-B1

Nuclear chloride channel-27, NCC27 or CLIC1 (Br^- > Cl^- > I^-) (241 aas).  CLIC1 has two charged residues, K37 and R29, in its single TMS which are important for the biophysical properties of the channel (Averaimo et al. 2013).  A putative Lys37-Trp35 cation-pi interaction stabilizes the active dimeric form of the CLIC1 TMS in membranes (Peter et al. 2013).  This channel may play a role in cancer (Peretti et al. 2014).  A positively charged motif at the C-terminus of the single TMS enhances membrane partitioning and insertion via electrostatic contacts.  It also functions as an electrostatic plug to anchor the TMS in membranes and is involved in orientating the TMS with respect to the cis and trans faces of the membrane (Peter et al. 2014).  The CLIC1 protein accumulates in the circulating monocyte membrane during neurodegeneration (Carlini et al. 2020). The involvement of CLIC1 protein functions in physiological and in pathological conditions has been reviewed (Cianci and Verduci 2021).

The Janus protein, CLIC2. The 3-D structure of its water soluble form has been determined at 1.8 Å resolution (Cromer et al., 2007). CLIC2 interacts with the skeletal ryanodine receptor (RyR1) and modulates its channel activity (Meng et al., 2009).

Chloride intracellular channel protein 4, CLIC4.  Regulates the histamine H3 receptor (Maeda et al., 2008)) Discriminates poorly between anions and cations (Singh and Ashley, 2007). 76% identical to CLIC5; it may play a role in cancer (Peretti et al. 2014). CLIC5 was the first mitochondrial chloride channel to be identified in the inner mitochondrial membrane, while CLIC4 is located predominantly in the outer mitochondrial membrane. Gururaja Rao et al. 2020 discussed the intracellular chloride channels, their roles in pathologies, such as cardiovascular, cancer, and neurodegenerative diseases, and developments concerning their usage as theraputic targets in humans.

Intracellular Cl^- channel-3 (CLIC3). The 3-d structure is known (3FY7). This protein is associated with pregnancy disorders (Murthi et al., 2012). 

Putative Glutathione S-transferase.  Pore formation has not been demonstrated in prokaryotes.

Ca⁺²-activated Cl^- channel, or Chloride Channel Accessory 1 (CLCA1 or CACC1), of 914 aas.  CLCA1 may play a role in inflammatory airway diseases (Sala-Rabanal et al. 2015). It is thought to play a role in Cl^- secretion in the intestine (A. Quach, personal communication).

The 7 TMS proton-sensitive Ca²⁺ leak channel, YetJ.  The activity and high resolution 3-d structure have been determined (Chang et al. 2014). BsYetJ in lipid nanodiscs is structurally different from those crystallized in detergents. Li et al. 2020 showed that the BsYetJ conformation is pH-sensitive in the apo state (lacking calcium), whereas in a calcium-containing solution, it is stuck in an intermediate state, inert to pH changes. Only when the transmembrane calcium gradient is established can the calcium-release activity of holo-BsYetJ occur and be mediated by pH-dependent conformational changes, suggesting a dual gating mechanism. Conformational substates involved in the process and a key residue, D171, relevant to the gating of calcium were identified. Thus, BsYetJ/TMBIM6 is a pH-dependent, voltage-gated calcium channel (Li et al. 2020). The transmembrane BAX inhibitor-1-containing motif 6 (TMBIM6) protein may modulate apoptosis by regulating calcium homeostasis in the endoplasmic reticulum (ER). Lan et al. 2023 investigated all negatively charged residues in BsYetJ, a bacterial homolog of TMBIM6. They reconstituted BsYetJ in membrane vesicles with a lipid composition similar to that of the ER. The charged residues E49 and R205 work together as a major gate, regulating calcium conductance in these ER-like lipid vesicles. However, these residues become largely inactive when reconstituted in other lipid environments. D195 acts as a minor filter compared to the E49-R205 dyad (Lan et al. 2023).

NS channel protein-1 or Sec62 protein of 399aas and 2 TMSs.  Efficient secretion of small proteins in mammalian cells relies on Sec62-dependent posttranslational translocation (Lakkaraju et al. 2012).

Formate uptake/efflux permease, FocA.  It catalyzes bidirectional transport, has a pentameric aquaporin-like (TC# 1.A.8) structure, and may function by a channel-type mechanism (Falke et al., 2009; Wang et al. 2009). The structure at 2.25 Å resolution has been determined (Wang et al., 2009).  The protein is encoded in an operon with pyruvate-formate lyase, PflB.  A pyruvate:formate antiport mechanism has been suggested (Moraes and Reithmeier 2012). The C-terminal 6 aas are required for formate transport, but not for homopentamer formation (Hunger et al. 2017).  The N-terminus of FocA interacts with PflB, and this interaction is essential for optimal formate translocation (Doberenz et al. 2014). In fact, the GREs, TdcE and PflB, interact with the FNT channel protein, probably to control formate translocation by FocA (Falke et al. 2016). The lipophilic constrictions of FocA mainly act as barriers to isolate the central histidine from the aqueous bulk, preventing protonation via proton wires. Thus, an FNT transport model is supported in which the central histidine is uncharged, and weak acid substrate anion protonation occurs in the vestibule regions of the transporter before passing the constrictions (Schmidt and Beitz 2021). An interplay between the conserved pore residues Thr-91 and His-209 controls formate translocation through the FocA channel (Kammel et al. 2022). T91 is essential for formate permeation in both directions; however, it is particularly important to allow anion efflux. H209 is essential for formate uptake by FocA, strongly suggesting that protonation-deprotonation of this residue plays a role in formate uptake. These observations substantiate the premise that efflux and influx of formate by FocA are mechanistically distinct processes that are controlled by the interplay between T91 and H209 (Kammel et al. 2022).

Nitrite uptake/efflux channel (Jia et al. 2009).

Hydrosulfide (hydrogen sulfide; HS^-), Fnt3 (Hsc) channel.  Also probably transports chloride, formate and nitrite. The 3-d crystal structure (2.2Å resolution in the closed state) is known (PDB# 3TE2) (Czyzewski and Wang, 2012). The Fnt3 gene is linked to the asrABC operon encoding the sulfite (SO₃^2-) reductase that gives HS^- as the product (Czyzewski and Wang 2012).

TMEM16 of 735 aas and 10 TMSs.  Operates as a Ca²⁺-activated lipid scramblase (Wang et al. 2018). Each subunit of the homodimer contains a hydrophilic membrane-traversing cavity that is exposed to the lipid bilayer as a potential site of catalysis. This cavity harbours a conserved Ca²⁺-binding site, located within the hydrophobic core of the membrane. Mutations of residues involved in Ca²⁺ coordination affect both lipid scrambling in N. haematococca TMEM16 and ion conduction in the Cl^- channel of TMEM16A. The structure reveals the general architecture of the family and its mode of Ca²⁺ activation (Brunner et al. 2014). While the cytoplasmic portion of the protein is important for function, it does not appear to regulate scramblase activity via a detectable conformational change (Andra et al. 2018). Dynamic modulation of the lipid translocation groove generates a conductive ion channel in Ca²⁺-bound nhTMEM16 (Khelashvili et al. 2019) (see family description). Permeation of potassium ions through the lipid scrambling path of nhTMEM16 has been documented (Cheng et al. 2022). Citral amide derivatives possess antifungal activity against Rhizoctonia. solani (Zhang et al. 2024).

TMem16A or Anoctamin-1 (Ano1) Ca²⁺-activated anion (Cl^-) channel of 960 aas and 10 TMSs.  Its structure has been solved by cryoEM (Paulino et al. 2017). The protein shows a similar organization to the fungal nhTMEM16, except for changes at the site of catalysis. There, the conformation of transmembrane helices, constituting a membrane-spanning furrow that provides a path for lipids in scramblases, is replaced to form an enclosed aqueous pore that is largely shielded from the membrane (Paulino et al. 2017). It thus provides a pathway for anions such as Cl^-.  During activation, the binding of Ca²⁺ to a site located within the transmembrane domain, in the vicinity of the pore, alters the electrostatic properties of the ion conduction path and triggers a conformational rearrangement of an α-helix that comes into physical contact with the bound ligand, and thereby directly couples ligand binding and pore opening (Paulino et al. 2017).  The E143A mutant showed reduced sensitivity to Ca²⁺ but not to high temperatures, whereas the E705V mutant exhibited reduced sensitivity to both Ca²⁺ and noxious heat (Choi et al. 2018). Loss of TMEM16A resulted in reduced nephron number and, subsequently, albuminuria and tubular damage (Schenk et al. 2018). mAno1 expression is regulated via alternative promoters, and its transcriptional variation results in variation of the N-terminal sequence of the Ano1 protein due to alternative translation initiation sites (Kamikawa et al. 2018). The Ca²⁺ gating mechanism of TMEM16A, involving a Ca²⁺-sensing element close to the anion pore, alters conduction and substrate selection. De Jesús-Pérez et al. 2022 studied the gating-permeant anion relationship using mouse TMEM16A, showing that the apparent Ca²⁺ sensitivity increases with highly permeant anions and SCN^- mole fractions, likely by stabilizing bound Ca²⁺. Conversely, mutations in crucial gating elements, including the Ca²⁺-binding site 1,TMS 6, and the hydrophobic gate, impaired anion permeability and selectivity. Thus, there is a reciprocal rationship between gating and selectivity (De Jesús-Pérez et al. 2022).  Propagation of pacemaker activity and peristaltic contractions in the mouse renal pelvis rely on Ca²⁺-activated Cl^- Channels such as Ano1 and T-type Ca²⁺ channels (Grainger et al. 2022).

Anoctamin-10 or TMEM16K of 660 aas and 9 or 10 TMSs.  In the presence of Ca²⁺, TMEM16K directly binds Ca²⁺ to form a stable complex (Ishihara et al. 2016). In the absence of Ca²⁺, TMEM16K and TMEM16F (TC# 1.A.17.1.4) aggregate, suggesting that their structures are stabilized by Ca²⁺.  Mutagenesis of acidic residues in TMEM16K's cytoplasmic and transmembrane regions identified five residues that are critical for binding Ca²⁺. These residues are well conserved between TMEM16F and 16K, and point mutations of these residues in TMEM16F reduced its ability to support Ca²⁺-dependent phospholipid scrambling (Ishihara et al. 2016). Phosphatidyl serine in the ER of mammalian cells is predominantly localized to the cytoplasmic leaflet, but TMEM16K directly or indirectly mediates Ca²⁺-dependent phospholipid scrambling (Tsuji et al. 2019). Ano10 plays roles in cell division, migration, apoptosis, cell signalling, and developmental processes (Chrysanthou et al. 2022).  There is structural heterogeneity within the ion and lipid channel of TMEM16F (Ye et al. 2024).  It coordinates organ morphogenesis in the urochordate notochord (Liang et al. 2024)

Structural heterogeneity of the ion and lipid channel TMEM16F.

Anoctamin-8, Ano8 or TMEM16H, of 1232 aas and 9 TMSs. It tethers the endoplasmic reticulum and plasma membrane for assembly of Ca²⁺ signaling complexes at the ER/PM compartment (Jha et al. 2019). ANO8 is a key tether in the formation of the ER/PM junctions that are essential for STIM1-STIM1 interaction and STIM1-Orai1 interaction and channel activation at a ER/PM PI(4,5)P₂-rich compartment. Moreover, ANO8 assembles all core Ca²⁺ signaling proteins: Orai1, PMCA, STIM1, IP₃ receptors, and SERCA2 at the ER/PM junctions to mediate a novel form of Orai1 channel inactivation by markedly facilitating SERCA2-mediated Ca²⁺ influx into the ER. This controls the efficiency of receptor-stimulated Ca²⁺ signaling, Ca²⁺ oscillations, and the duration of Orai1 activity to prevent Ca²⁺ toxicity (Jha et al. 2019).

The non-rectifying, plasma membrane, calcium-permeable, stress-gated, cation channel 1 (CSC1) of 771 aas (Hou et al. 2014).  Activated by hyperosmotic shock.  Permeable to Ca²⁺, K⁺ and Na⁺.  Inactivation or closure is Ca²⁺-dependent.  The N-terminal region contains 3 TMSs, the first of which may be a cleavable signal peptide., and the C-terminal region contains 6 TMSs corresponding to the DUF221 domain.  Arabidopsis contains at least 15 CSCs ((Hou et al. 2014).  Some plant homologues are transcriptionally upregulated in response to vaious abiotic and biotic stresses involving mechanical perturbation (Kiyosue et al. 1994).

OSCA1.2 of 772 aas and 11 TMSs.  It is a dimer containing eleven TMSs per subunit, similar to other TMEM16 proteins. Jojoa Cruz et al. 2018 located the ion permeation pathway within each subunit by demonstrating that a conserved acidic residue is a determinant of channel conductance. Molecular dynamics simulations revealed membrane interactions, suggesting a role of lipids in gating.  The high resolution structure of this hyperosmolality-gated calcium-permeable channel has been determined (Liu et al. 2018). It contains 11 TMSs and forms a homodimer. The pore-lining residues were clearly identified. Its cytosolic domain contains an RNA recognition motif and two unique long helices. The linker between these two helices forms an anchor in the lipid bilayer and may be essential to osmosensing. Genome-wide analyses of OSCA gene family members in Vigna radiata have revealed their involvement in the osmotic response (Yin et al. 2021). There are 42 OSCA channel proteins in Triticum aestivum, and  their diverse roles during development and stress responses have been evaluated (Kaur et al. 2022).

Dimeric OSCA1.2 of 766 aas and 11 TMSs.  The 3-D structure has been determined (K. Maity et al., PNAS, in press).  This protein is 69% identical to the A. thaliana ortholog (TC# 1.A.17.5.10). It is a putative early stress-responsive osmolality-sensing ion channel protein. A model has been proposed by which it may mediate hyperosmolality-sensing and consequent gating of ion transport. It has a cytosolic domain structurally related to RNA recognition proteins that includes helical arms paralell to the plane of the membrane.  They may sense lateral tension in the inner leaflet, caused by changes in turgor pressure, allowing gating of the channel via coupling of the two domains.

Tic110 channel protein. The x-ray structure (4.2Å resolution) of Tic110B and C from Cyanidioschyzon merolae is known (Tsai et al., 2013). The C-terminal half of Tic110 posesses a rod-shaped helix-repeat structure that is too flattened and elongated to be a channel. The structure is most similar to the HEAT-repeat motif that functions as scaffolds for protein-protein interactions (Tsai et al., 2013).

Matrix protein, M2, an acid activated drug-sensitive proton channel.  Transport involves binding to the four His-37s and transfer to water molecules on the inside of the channel (Acharya et al., 2010).  Functional properties and structure are known (Hong and Degrado 2012). The cytoplasmic tail facilitates proton conduction (Liao et al. 2015).  It is a dimer of dimers (Andreas et al. 2015).  The four TMSs flanking the channel lumen alone seem to determine the proton conduction mechanism (Liang et al. 2016). His-37 forms a planar tetrad in the configuration of the bundle accepting and translocating the incoming protons from the N terminal side, exterior of the virus, to the C terminal side, inside the virus (Kalita and Fischer 2017). The cholesterol binding site in M2 that mediates membrane scission in a cholesterol-dependent manner to cause virus budding and release has been identified (Elkins et al. 2017).Transport-related conformational changes coupled to water and H⁺ movements have been studied (Mandala et al. 2018). The L46P mutant confers a novel allosteric mechanism of resistance towards the influenza A virus M2 S31N proton channel blockers (Musharrafieh et al. 2019). The C-terminal domain of M2 may serve as a sensor that regulates how M2 participates in critical events in the viral infection cycle (Kim et al. 2019). The M2 proton channel protein  self-assembles into tetramers that retain the ability to bind to the drug amantadine, and the effects of phospholipid acyl chain length and cholesterol on the peptide association were investigated. Association of the helices depends on the thickness of the bilayer and cholesterol levels present in the phospholipid bilayer. The most favorable folding occurred when there was a good match between the width of the apolar region of the bilayer and the hydrophobic length of the transmembrane helix with tighter association upon inclusion of cholesterol in the lipid bilayer (Cristian et al. 2003).

G-protein-activated inward rectifying K⁺ channel, Kir3.2, KATP2, KCNJ6, KCNJ7 or GIRK2 of 423 aas and 2 TMSs (Inanobe et al., 2011; Yokogawa et al. 2011). Mutations cause the Keppen-Lubinsky syndrome (Gao et al. 2022). It functions in electrical signaling in neurons and muscle cells (Weng et al. 2021), being important in regulating heart rate and neuronal excitability.  It is activated by binding of the βγ-subunit complex to the cytoplasmic pore gate (Yokogawa et al. 2011). Chen et al. 2017 found that GIRK channels are activated by Ivermectin (IVM). Cholesterol binds to and upregulates GIRK channels (GIRK2 and 4), and the binding sites have been determined (Rosenhouse-Dantsker 2018). An inherited gain-of-function mutation in the human GIRK3.4 causes familial human sinus node dysfunction (SND). The increased activity of GIRK channels likely leads to a sustained hyperpolarization of pacemaker cells and thereby reduces heart rate (Kuß et al. 2019). GIRK2 channels are abundantly expressed in the heart and require that phosphatidylinositol bisphosphate (PIP2) is present so that intracellular channel-gating regulators such as Gbetagamma (Gβγ) and Na⁺ ions maintain the channel-open state. Li et al. 2019 determined how each regulator uses  channel domain movements to control gate transitions. Na⁺ controls the cytosolic gate of the channel through an anti-clockwise rotation, whereas Gβγ stabilizes the transmembrane gate in the open state through a rocking movement of the cytosolic domain. Both effects altered the way by which the channel interacts with PIP2 and thereby stabilizes the open states of the respective gates (Li et al. 2019). The protein plays a role in heart atrial fibrillation-valvular heart disease (VHD) (Zhao et al. 2021). Measurements of ligand binding and channel current have been made (Usher et al. 2021). CryoEM structures of GIRK2 in the presence and absence of the cholesterol analog cholesteryl hemisuccinate (CHS) and phosphatidylinositol 4,5-bisphosphate (PIP2) reveal that CHS binds near PIP2 in lipid-facing hydrophobic pockets of the transmembrane domain, suggesting that CHS stabilizes the PIP2 interaction with the channel to promote engagement of the cytoplasmic domain with the transmembrane region (Mathiharan et al. 2021). It may play a role in Parkinson's Disease (Zhou et al. 2023).

KCNJ11 or Kir6.2 or KATP of 390 aas; 96% identical to the rat homologue, TC# 1.A.2.1.7. Congenital hyperinsulinism (CHI) is characterized by persistent insulin secretion despite severe hypoglycemia. Mutations in the pancreatic ATP-sensitive K⁺ (K(ATP)) channel proteins sulfonylurea receptor 1 (SUR1) and Kir6.2, encoded by ABCC8 and KCNJ11, respectively, is the most common cause of the disease. Many mutations in SUR1 render the channel unable to traffic to the cell surface, thereby reducing channel function. Many studies have shown that for some SUR1 trafficking mutants, the defects could be corrected by treating cells with sulfonylureas or diazoxide (Yan et al. 2007). Inward rectifier potassium channels are characterized by a greater tendency to allow potassium to flow into the cell rather than out of it. Their voltage dependence is regulated by the concentration of extracellular potassium; as external potassium is raised, the voltage range of the channel opening shifts to more positive voltages. The inward rectification is mainly due to the blockage of outward current by internal magnesium (Tammaro and Ashcroft 2007). Kir6.2 is an ATP-sensitive potassium (K_ATP) channel coupling cell metabolism to electrical activity by regulating K⁺ fluxes across the plasma membrane. Channel closure is facilitated by ATP, which binds to the pore-forming subunit (Kir6.2). Conversely, channel opening is potentiated by phosphoinositol bisphosphate (PIP₂), which binds to Kir6.2 and reduces channel inhibition by ATP.  The PIP₂ binding site has been identified (Haider et al. 2007).  KATP channels are metabolic sensors that couple cell energetics to membrane excitability. In pancreatic beta-cells, channels formed by SUR1 and Kir6.2 regulate insulin secretion and are the targets of antidiabetic sulfonylureas. Martin et al. 2017 used cryo-EM to elucidate the structural basis of channel assembly and gating. The structure, determined in the presence of ATP and the sulfonylurea, glibenclamide, at ~6 Å resolution, revealed a closed Kir6.2 tetrameric core with four peripheral SUR1s, each anchored to a Kir6.2 by its N-terminal transmembrane domain (TMD0). Intricate interactions between TMD0, the loop following TMD0, and Kir6.2 near the proposed PIP2 binding site, and where ATP density is observed, suggest that SUR1 may contribute to ATP and PIP2 binding to enhance Kir6.2 sensitivity to both. The SUR1-ABC core is found in an unusual inward-facing conformation whereby the two nucleotide binding domains are misaligned along a two-fold symmetry axis, revealing a possible mechanism by which glibenclamide inhibits channel activity (Martin et al. 2017). a cryo-EM structure of a hamster SUR1/rat Kir6.2 channel bound to a high-affinity sulfonylurea drug glibenclamide and ATP has been solved at 3.63 Å resolution. The structure shows that glibenclamide is lodged in the transmembrane bundle of the SUR1-ABC core connected to the first nucleotide binding domain near the inner leaflet of the lipid bilayer (Martin et al. 2017). The activation of K(ATP) channels contributes to the shortening of action potential duration but is not the primary cause of extracellular K⁺ accumulation during early myocardial ischemia (Saito et al. 2005). KATP binds nucleotides (Usher et al. 2021). Mitochondrial KATP channels stabilize intracellular Ca²⁺ during hypoxia in retinal horizontal cells of goldfish (Carassius auratus) (Country and Jonz 2021). Medicinal plant products can interact with KATP (Rajabian et al. 2022). Remodeling of excitation-contraction coupling in transgenic mice expressing ATP-insensitive sarcolemmal KATP channels has been observed (Flagg et al. 2004). Thus, a compensatory increase in I(Ca) counteracts a mild activation of ATP-insensitive K(ATP) channels. Pharmacological inhibitors and ATP enrich a channel conformation in which the Kir6.2 cytoplasmic domain is closely associated with the transmembrane domain, while depleting one where the Kir6.2 cytoplasmic domain is extended away into the cytoplasm. This conformational change remodels a network of intra- and inter-subunit interactions as well as the ATP and PIP2 binding pockets. The structures resolved key contacts between the distal N-terminus of Kir6.2 and SUR1's ABC module involving residues implicated in channel function and showed a SUR1 residue, K134, participates in PIP2 binding. Molecular dynamics simulations revealed two Kir6.2 residues, K39 and R54, that mediate both ATP and PIP2 binding, suggesting a mechanism for competitive gating by ATP and PIP2 (Sung et al. 2022). The natural product, 7-hydroxycoumarin (7-HC), exhibits pharmacological properties linked to antihypertensive mechanisms of action. This relaxant effect induced by 7-HC relies on K⁺-channels (K_ATP, BK_Ca, and, to a lesser extent, K_v) activation and also on Ca²⁺ influx from sarcolemma and sarcoplasmic reticulum mobilization (inositol 1,4,5-triphosphate (IP3) and ryanodine receptors) (Jesus et al. 2022). Lymphatic contractile dysfunction in mouse models of Cantú Syndrome is oberved with K_ATP channel gain-of-function mutations (Davis et al. 2023). The structure of an open K (ATP) channel has revealed tandem PIP binding sites mediating the Kir6.2 and SUR1 regulatory interface (Driggers et al. 2023).

Insulin secretion is regulated by ATP-sensitive potassium (K_ATP) channels in pancreatic β-cells. Peroxisome proliferator-activated receptors (PPAR)α ligands are used to treat dyslipidemia. A PPARα ligand, fenofibrate, and PPARγ ligands troglitazone and 15-deoxy-∆^12,14-prostaglandin J2 close K_ATP channels and induce insulin secretion. The PPARα ligand, pemafibrate, is used to treat dyslipidemia and improves glucose intolerance in mice treated with a high fat diet and a novel selective PPARα modulator, it may affect K_ATP channels or insulin secretion. The effect of fenofibrate and pemafibrate (both at 100 µM) on insulin secretion was measured. Addition of fenofibrate for 10 min increased insulin secretion in low glucose conditions. The K_ATP channel activity was measured. Although fenofibrate (100 µM) reduced the K_ATP channel current, it had no effect on insulin mRNA expression (Kitamura et al. 2023).

Inward rectifier potassium channel 4, KCNJ4, IRK3 or 4, of 445 aas. Its voltage dependence is regulated by the concentration of extracellular potassium; as external potassium is raised, the voltage range of the channel opening shifts to more positive voltages. The inward rectification is mainly due to the blockage of outward current by internal magnesium, and it can be blocked by extracellular barium and cesium. It may play a role in the control of polyamine-mediated channel gating and in the blocking by intracellular magnesium. Overexpression of KCNJ4 correlates with cancer progression and nfavorable prognosis in lung adenocarcinoma (Wu and Yu 2019).

G-protein enhanced inward rectifier K channel 2, IRK1, IRK2, KCNJ2, KCNJ5, Kir2.1 (Andersen-Tawil Syndrome (ATS-1) protein; the V302M mutation causing the syndrome, alters the G-loop cytoplasmic K conduction pathway) (Bendahhou et al., 2003; Ma et al., 2007). (Blocked by chloroquine which binds in the cytoplasmic pore domain (Rodriguez-Menchaca et al., 2008)). Forms heteromultimers with Kir3.1 and Kir3.4 (Ishihara et al., 2009). A C-terminal domain is critical for the sensitivity of Kir2.1 to cholesterol (Epshtein et al., 2009). Flecainide increases Kir2.1 currents by interacting with cysteine 311, decreasing the polyamine-induced rectification (Caballero et al., 2010).  The inhibitory cholesterol binding site has been identified (Fürst et al. 2014).  Polyamines and Mg²⁺ block ion flux synergistically (Huang and Kuo 2016). Long polyamines serve a dual role as both blockers and coactivators (with PIP2) of Kir2.1 channels (Xie et al. 2005).  Cytoplasmic domain structures of Kir2.1 and Kir3.1 show sites for modulating gating and rectification (Pegan et al. 2005). Loss-of-function mutations are a rare cause of long QT syndrome (Fodstad et al. 2004). Fibroblast growth factor 21 ameliorates NaV1.5 and Kir2.1 channel dysregulation in human AC16 cardiomyocytes (Li et al. 2021). The trafficking of Kir2.1 and its role in development have been reviewed (Hager et al. 2021). Cholesterol-induced suppression of Kir2 channels is mediated by decoupling at the inter-subunit interfaces (Barbera et al. 2022). CryoEM studies have revealed a well-connected network of interactions between the PIP2-binding site and the G-loop through residues R312 and H221.Moreover, the intrinsic tendency of the CTD to tether to the TMD and a movement of the secondary anionic binding site to the membrane even without PIP2 was revealed (Fernandes et al. 2022). The results revealed structural features unique to human Kir2.1. Individual protonation events change the electrostatic microenvironment of the pore, resulting in distinct, uncoordinated, and relatively long-lasting conductance states, which depend on levels of ion pooling in the pore and the maintenance of pore wetting (Maksaev et al. 2023). Subunit gating results from individual protonation events in Kir2 channels (Maksaev et al. 2023).

Kidney/pancreas/muscle ATP-senstive, ER/Golgi K⁺ channel, KATP or ROMK (Kir6.2) (Boim et al., 1995) (three alternatively spliced isoforms are called ROMK1-3). Involved in congenital hyperinsulinism (Lin et al., 2008). Regulated by Ankyrin-B (Li et al., 2010). ATP activates ATP-sensitive potassium channels composed of mutant sulfonylurea receptor 1 (SUR1) and Kir6.2 with diminished PIP2 sensitivity (Pratt and Shyng, 2011). This channel protects the myocardium from hypertrophy induced by pressure-overloading (Alvin et al., 2011). Domain organization studies show which domains in Sur and Kir6.2 interact (Wang et al. 2012).  KATP channels consisting of Kir6.2 and SUR1 couple cell metabolism to membrane excitability and regulate insulin secretion in pancreatic beta cells, and mutations in the former protein can compensate for mutations in the latter (Zhou et al. 2013).  Mutations cause inactivation of channel function by disrupting PIP2-dependent gating (Bushman et al. 2013). Thus, these proteins comprise part of the glucose sensing mechanism (Rufino et al. 2013).  A single point mutation can confer voltage sensitivity (Kurata et al. 2010).  Its involvement in type II diabetes has been reviewed by Bonfanti et al. 2015. KATP channels (Kir6.2/SUR1) in the brain and endocrine pancreas  couple metabolic status to the membrane potential. In beta-cells, increases in cytosolic [ATP/ADP] inhibit KATP channel activity, leading to membrane depolarization and exocytosis of insulin granules. Mutations in ABCC8 (SUR1) or KCNJ11 (Kir6.2) can result in gain or loss of channel activity and cause neonatal diabetes (ND) or congenital hyperinsulinism (CHI), respectively.  Nucleotide binding without hydrolysis switches SUR1 to stimulatory conformations.  Increased affinity for ATP gives rise to ND while decreased affinty gives rise to CHI (Ortiz and Bryan 2015).  Kir6.2 can associate with either SUR1 (TC# 3.A.1.208.4) or SUR2A (TC# 3.A.1.208.23) to form heteroctamers, leading to different locations and consequences (Principalli et al. 2015). IATP channels and Ca²⁺ influx play roles in purinergic vasotoxicity and cell death (Shibata et al. 2018).
       A cryo-EM structure of a hamster SUR1/rat Kir6.2 channel bound to a high-affinity sulfonylurea drug, glibenclamide, and ATP at 3.63 A resolution revealed details of the ATP and glibenclamide binding sites (Martin et al. 2017). The structure showed that glibenclamide lodges in the transmembrane bundle of the SUR1-ABC core connected to the first nucleotide binding domain near the inner leaflet of the lipid bilayer. Mutation of residues predicted to interact with glibenclamide led to reduced sensitivity to this drug (Martin et al. 2017).

Prokaryotic K⁺-selective Kir channel KirBac1.1 (selectivity: K⁺ = Rb⁺ = Cs⁺ >> Li⁺, Na⁺ or NMGM) (Enkvetchakul et al., 2004), inward rectifying (Cheng et al., 2009). Closure of the Kir1.1 pH gate results from steric occlusion of the permeation path by the convergence of four leucines (or phenylalanines) at the cytoplasmic apex of the inner transmembrane helices. In the open state, K⁺ crosses the pH gate together with its hydration shell (Sackin et al. 2005). An inhibitory cholesterol binding site has been identified (Fürst et al. 2014). Conformational changes associated with an open activation gate have been identified, and these suggest an allosteric pathway that ties the selectivity filter to the activation gate through interactions between both transmembrane helices, the turret, the selectivity filter loop, and the pore helix. Specific residues involved in this conformational exchange that are highly conserved among human Kir channels have also been identified (Amani et al. 2020). Anionic lipids, especially cardiolipin, initiate a concerted rotation of the cytoplasmic domain subunits. This action buries ionic side chains away from the bulk water, while allowing water greater access to the K⁺ conduction pathway (Borcik et al. 2020). Kv1.5 channels are regulated by PKC-mediated endocytic degradation (Du et al. 2021). Pore-forming TMSs control ion selectivity and the selectivity filter conformation in the KirBac1.1 channel (Matamoros and Nichols 2021). Key functional residues involved in gating and lipid allostery of K⁺ Kir channels have been identified (Yekefallah et al. 2022).

The KirBac3.1 K⁺ channel (a dimer of dimers with gating visualized by atomic force microscopy (Jaroslawski et al., 2007) (regulated by binding lipids, G-proteins, nucleotides, and ions (H⁺, Ca²⁺, and Mg²⁺)). The 3-D structure is available (1XL6_A).  The inhibitory cholesterol binding site has been identified (Fürst et al. 2014). The constricted opening in this, and presumably other, Kir channels does not impede potassium conduction (Black et al. 2020). The structural and dynamic properties of a KirBac3.1 mutant revealed the function of a highly conserved tryptophan in the transmembrane domain (Fagnen et al. 2021).