ELUCIDATING THE DIFFERENTIAL IMPACTS OF EQUIVALENT GATING-CHARGE MUTATIONS IN VOLTAGE-GATED SODIUM CHANNELS.

Voltage-gated sodium (Na) channels are pivotal for cellular signaling and mutations in Na channels can lead to excitability disorders in cardiac, muscular, and neural tissues. A major cluster of pathological mutations localizes in the voltage-sensing domains (VSDs), resulting in either gain-of-function (GoF), loss-of-function (LoF) effects, or both. However, the mechanism behind this functional divergence of mutations at equivalent positions remains elusive.

The chemistry of electrical signaling in sodium channels from bacteria and beyond.

Electrical signaling is essential for all fast processes in biology, but its molecular mechanisms have been uncertain. This review article focuses on studies of bacterial sodium channels in order to home in on the essential molecular and chemical mechanisms underlying transmembrane ion conductance and voltage-dependent gating without the overlay of complex protein interactions and regulatory mechanisms in mammalian sodium channels. This minimalist approach has yielded a nearly complete picture of sodium channel function at the atomic level that are mostly conserved in mammalian sodium channels, including sodium selectivity and conductance, voltage sensing and activation, electromechanical coupling to pore opening and closing, slow inactivation, and pathogenic dysfunction in a debilitating channelopathy.

Pathogenic gating pore current conducted by autism-related mutations in the Na1.2 brain sodium channel.

Autism spectrum disorder (ASD) is a complex neurodevelopmental condition characterized by social and communication deficits and repetitive behaviors. The genetic heterogeneity of ASD presents a challenge to the development of an effective treatment targeting the underlying molecular defects. ASD gating charge mutations in the /K7 potassium channel cause gating pore currents (I) and impair action potential (AP) firing of dopaminergic neurons in brain slices.

Dual receptor-sites reveal the structural basis for hyperactivation of sodium channels by poison-dart toxin batrachotoxin.

The poison dart toxin batrachotoxin is exceptional for its high potency and toxicity, and for its multifaceted modification of the function of voltage-gated sodium channels. By using cryogenic electron microscopy, we identify two homologous, but nonidentical receptor sites that simultaneously bind two molecules of toxin, one at the interface between Domains I and IV, and the other at the interface between Domains III and IV of the cardiac sodium channel. Together, these two bound toxin molecules stabilize α/π helical conformation in the S6 segments that gate the pore, and one of the bound BTX-B molecules interacts with the crucial Lys1421 residue that is essential for sodium conductance and selectivity via an apparent water-bridged hydrogen bond.

Structural basis for severe pain caused by mutations in the voltage sensors of sodium channel NaV1.7.

Voltage-gated sodium channels in peripheral nerves conduct nociceptive signals from nerve endings to the spinal cord. Mutations in voltage-gated sodium channel NaV1.7 are responsible for a number of severe inherited pain syndromes, including inherited erythromelalgia (IEM).

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

Heterologous expression of recombinant ion channel subunits in cell lines is often limited by the presence of a low number of channels at the cell surface level. Here, we introduce a combination of two techniques: viral expression using the baculovirus system plus cell-cycle arrest at the G1/S boundary using either thymidine or hydroxyurea. This method achieved a manifold increase in the peak current density of expressed ion channels compared with the classical liposome-mediated transfection methods.

Structural basis for severe pain caused by mutations in the S4-S5 linkers of voltage-gated sodium channel Na1.7.

Gain-of-function mutations in voltage-gated sodium channel Na1.7 cause severe inherited pain syndromes, including inherited erythromelalgia (IEM). The structural basis of these disease mutations, however, remains elusive.

When the Gates Swing Open Only: Arrhythmia Mutations That Target the Fast Inactivation Gate of Na1.5.

Na1.5 is the main voltage-gated sodium channel found in cardiac muscle, where it facilitates the fast influx of Na ions across the cell membrane, resulting in the fast depolarization phase-phase 0 of the cardiac action potential. As a result, it plays a major role in determining the amplitude and the upstroke velocity of the cardiac impulse.

Convergent regulation of Ca1.2 channels by direct phosphorylation and by the small GTPase RAD in the cardiac fight-or-flight response.

The L-type calcium currents conducted by the cardiac Ca1.2 calcium channel initiate excitation-contraction coupling and serve as a key regulator of heart rate, rhythm, and force of contraction. Ca1.

Druggability of Voltage-Gated Sodium Channels-Exploring Old and New Drug Receptor Sites.

Voltage-gated ion channels are important drug targets because they play crucial physiological roles in both excitable and non-excitable cells. About 15% of clinical drugs used for treating human diseases target ion channels. However, most of these drugs do not provide sufficient specificity to a single subtype of the channels and their off-target side effects can be serious and sometimes fatal.

Fenestropathy of Voltage-Gated Sodium Channels.

Voltage-gated sodium channels (Na) are responsible for the initiation and propagation of action potentials in excitable cells. From pain to heartbeat, these integral membrane proteins are the ignition stations for every sensation and action in human bodies. They are large (>200 kDa, 24 transmembrane helices) multi-domain proteins that couple changes in membrane voltage to the gating cycle of the sodium-selective pore.

Autism-associated mutations in K7 channels induce gating pore current.

Autism spectrum disorder (ASD) adversely impacts >1% of children in the United States, causing social interaction deficits, repetitive behaviors, and communication disorders. Genetic analysis of ASD has advanced dramatically through genome sequencing, which has identified >500 genes with mutations in ASD. Mutations that alter arginine gating charges in the voltage sensor of the voltage-gated potassium (K) channel K7 (KCNQ) are among those frequently associated with ASD.

Open-state structure and pore gating mechanism of the cardiac sodium channel.

The heartbeat is initiated by voltage-gated sodium channel Na1.5, which opens rapidly and triggers the cardiac action potential; however, the structural basis for pore opening remains unknown. Here, we blocked fast inactivation with a mutation and captured the elusive open-state structure.

Novel G1481V and Q1491H SCN5A Mutations Linked to Long QT Syndrome Destabilize the Nav1.5 Inactivation State.

Background: Na1.5, which is encoded by the gene, is the predominant voltage-gated Na channel in the heart. Several mutations of this gene have been identified and reported to be involved in several cardiac rhythm disorders, including type 3 long QT interval syndrome, that can cause sudden cardiac death.

Structural basis for voltage-sensor trapping of the cardiac sodium channel by a deathstalker scorpion toxin.

Voltage-gated sodium (Na) channels initiate action potentials in excitable cells, and their function is altered by potent gating-modifier toxins. The α-toxin LqhIII from the deathstalker scorpion inhibits fast inactivation of cardiac Na1.5 channels with IC = 11.

Structural Basis for High-Affinity Trapping of the Na1.7 Channel in Its Resting State by Tarantula Toxin.

Voltage-gated sodium channels initiate electrical signals and are frequently targeted by deadly gating-modifier neurotoxins, including tarantula toxins, which trap the voltage sensor in its resting state. The structural basis for tarantula-toxin action remains elusive because of the difficulty of capturing the functionally relevant form of the toxin-channel complex. Here, we engineered the model sodium channel NaAb with voltage-shifting mutations and the toxin-binding site of human Na1.

Computational design of transmembrane pores.

Transmembrane channels and pores have key roles in fundamental biological processes and in biotechnological applications such as DNA nanopore sequencing, resulting in considerable interest in the design of pore-containing proteins. Synthetic amphiphilic peptides have been found to form ion channels, and there have been recent advances in de novo membrane protein design and in redesigning naturally occurring channel-containing proteins. However, the de novo design of stable, well-defined transmembrane protein pores that are capable of conducting ions selectively or are large enough to enable the passage of small-molecule fluorophores remains an outstanding challenge.

Structure of the Cardiac Sodium Channel.

Voltage-gated sodium channel Na1.5 generates cardiac action potentials and initiates the heartbeat. Here, we report structures of Na1.

Structure and Pharmacology of Voltage-Gated Sodium and Calcium Channels.

Voltage-gated sodium and calcium channels are evolutionarily related transmembrane signaling proteins that initiate action potentials, neurotransmission, excitation-contraction coupling, and other physiological processes. Genetic or acquired dysfunction of these proteins causes numerous diseases, termed channelopathies, and sodium and calcium channels are the molecular targets for several major classes of drugs. Recent advances in the structural biology of these proteins using X-ray crystallography and cryo-electron microscopy have given new insights into the molecular basis for their function and pharmacology.

Structural Basis for Diltiazem Block of a Voltage-Gated Ca Channel.

Diltiazem is a widely prescribed Ca antagonist drug for cardiac arrhythmia, hypertension, and angina pectoris. Using the ancestral Ca channel construct CaAb as a molecular model for X-ray crystallographic analysis, we show here that diltiazem targets the central cavity of the voltage-gated Ca channel underneath its selectivity filter and physically blocks ion conduction. The diltiazem-binding site overlaps with the receptor site for phenylalkylamine Ca antagonist drugs such as verapamil.

Resting-State Structure and Gating Mechanism of a Voltage-Gated Sodium Channel.

Voltage-gated sodium (Na) channels initiate action potentials in nerve, muscle, and other electrically excitable cells. The structural basis of voltage gating is uncertain because the resting state exists only at deeply negative membrane potentials. To stabilize the resting conformation, we inserted voltage-shifting mutations and introduced a disulfide crosslink in the VS of the ancestral bacterial sodium channel NaAb.

Fenestrations control resting-state block of a voltage-gated sodium channel.

Potency of drug action is usually determined by binding to a specific receptor site on target proteins. In contrast to this conventional paradigm, we show here that potency of local anesthetics (LAs) and antiarrhythmic drugs (AADs) that block sodium channels is controlled by fenestrations that allow drug access to the receptor site directly from the membrane phase. Voltage-gated sodium channels initiate action potentials in nerve and cardiac muscle, where their hyperactivity causes pain and cardiac arrhythmia, respectively.

Molecular dissection of multiphase inactivation of the bacterial sodium channel NaAb.

Homotetrameric bacterial voltage-gated sodium channels share major biophysical features with their more complex eukaryotic counterparts, including a slow-inactivation mechanism that reduces ion-conductance activity during prolonged depolarization through conformational changes in the pore. The bacterial sodium channel NaAb activates at very negative membrane potentials and inactivates through a multiphase slow-inactivation mechanism. Early voltage-dependent inactivation during one depolarization is followed by late use-dependent inactivation during repetitive depolarization.