ABSTRACT
The EAG (ether-à-go-go) family of voltage-gated K+ channels are important regulators of neuronal and cardiac action potential firing (excitability) and have major roles in human diseases such as epilepsy, schizophrenia, cancer, and sudden cardiac death. A defining feature of EAG (Kv10-12) channels is a highly conserved domain on the N terminus, known as the eag domain, consisting of a Per-ARNT-Sim (PAS) domain capped by a short sequence containing an amphipathic helix (Cap domain). The PAS and Cap domains are both vital for the normal function of EAG channels. Using heme-affinity pulldown assays and proteomics of lysates from primary cortical neurons, we identified that an EAG channel, hERG3 (Kv11.3), binds to heme. In whole-cell electrophysiology experiments, we identified that heme inhibits hERG3 channel activity. In addition, we expressed the Cap and PAS domain of hERG3 in Escherichia coli and, using spectroscopy and kinetics, identified the PAS domain as the location for heme binding. The results identify heme as a regulator of hERG3 channel activity. These observations are discussed in the context of the emerging role for heme as a regulator of ion channel activity in cells.
Subject(s)
Cerebral Cortex/chemistry , Ether-A-Go-Go Potassium Channels/chemistry , Heme/chemistry , Neurons/chemistry , Cerebral Cortex/metabolism , Ether-A-Go-Go Potassium Channels/metabolism , Heme/metabolism , Humans , Neurons/metabolism , Protein Binding , Protein DomainsABSTRACT
The originally published version of this article contained an error in the subheading 'Heme is required for CO-dependent channel activation', which was incorrectly given as 'Hame is required for CO-dependent channel activation'. This has now been corrected in both the PDF and HTML versions of the Article.
ABSTRACT
The paralogues TRPV5 and TRPV6 belong to the vanilloid subfamily of the transient receptor potential (TRP) superfamily of ion channels, and both play an important role in overall Ca2+ homeostasis. The functioning of the channels centers on a tightly controlled Ca2+-dependent feedback mechanism in which the direct binding of the universal Ca2+-binding protein calmodulin (CaM) to the channel's C-terminal tail is required for channel inactivation. We have investigated this interaction at the atomic level and propose that under basal cellular Ca2+ concentrations CaM is constitutively bound to the channel's C-tail via CaM C-lobe only contacts. When the cytosolic Ca2+ concentration increases charging the apo CaM N-lobe with Ca2+, the CaM:TRPV6 complex rearranges and the TRPV6 C-tail further engages the CaM N-lobe via a crucial interaction involving L707. In a cellular context, mutation of L707 significantly increased the rate of channel inactivation. Finally, we present a model for TRPV6 CaM-dependent inactivation, which involves a novel so-called "two-tail" mechanism whereby CaM bridges two TRPV6 monomers resulting in closure of the channel pore.
Subject(s)
Calcium/chemistry , Calmodulin/chemistry , Multiprotein Complexes/chemistry , TRPV Cation Channels/chemistry , Amino Acid Sequence/genetics , Animals , Binding Sites , Calcium/metabolism , Calcium Signaling/genetics , Calmodulin/metabolism , HEK293 Cells , Humans , Multiprotein Complexes/genetics , Mutation , Protein Binding , Protein Conformation , Rats , TRPV Cation Channels/geneticsABSTRACT
Despite being highly toxic, carbon monoxide (CO) is also an essential intracellular signalling molecule. The mechanisms of CO-dependent cell signalling are poorly defined, but are likely to involve interactions with heme proteins. One such role for CO is in ion channel regulation. Here, we examine the interaction of CO with KATP channels. We find that CO activates KATP channels and that heme binding to a CXXHX16H motif on the SUR2A receptor is required for the CO-dependent increase in channel activity. Spectroscopic and kinetic data were used to quantify the interaction of CO with the ferrous heme-SUR2A complex. The results are significant because they directly connect CO-dependent regulation to a heme-binding event on the channel. We use this information to present molecular-level insight into the dynamic processes that control the interactions of CO with a heme-regulated channel protein, and we present a structural framework for understanding the complex interplay between heme and CO in ion channel regulation.
Subject(s)
Carbon Monoxide/metabolism , Ion Channels/metabolism , Amino Acid Motifs , Amino Acid Sequence , HEK293 Cells , Heme/metabolism , Humans , Ion Channel Gating , KATP Channels/metabolism , Models, Molecular , Spectrum Analysis, Raman , Sulfonylurea Receptors/chemistry , Sulfonylurea Receptors/metabolismSubject(s)
Ether-A-Go-Go Potassium Channels , Inosine Triphosphate , Amines , ERG1 Potassium ChannelABSTRACT
The ether à go-go family of voltage-gated potassium channels is structurally distinct. The N terminus contains an eag domain (eagD) that contains a Per-Arnt-Sim (PAS) domain that is preceded by a conserved sequence of 25-27 amino acids known as the PAS-cap. The C terminus contains a region with homology to cyclic nucleotide binding domains (cNBHD), which is directly linked to the channel pore. The human EAG1 (hEAG1) channel is remarkably sensitive to inhibition by intracellular calcium (Ca(2+) i) through binding of Ca(2+)-calmodulin to three sites adjacent to the eagD and cNBHD. Here, we show that the eagD and cNBHD interact to modulate Ca(2+)-calmodulin as well as voltage-dependent gating. Sustained elevation of Ca(2+) i resulted in an initial profound inhibition of hEAG1 currents, which was followed by a phase when current amplitudes partially recovered, but activation gating was slowed and shifted to depolarized potentials. Deletion of either the eagD or cNBHD abolished the inhibition by Ca(2+) i However, deletion of just the PAS-cap resulted in a >15-fold potentiation in response to elevated Ca(2+) i Mutations of residues at the interface between the eagD and cNBHD have been linked to human cancer. Glu-600 on the cNBHD, when substituted with residues with a larger volume, resulted in hEAG1 currents that were profoundly potentiated by Ca(2+) i in a manner similar to the ΔPAS-cap mutant. These findings provide the first evidence that eagD and cNBHD interactions are regulating Ca(2+)-dependent gating and indicate that the binding of the PAS-cap with the cNBHD is required for the closure of the channels upon CaM binding.
Subject(s)
Calcium/metabolism , Calmodulin/metabolism , Ether-A-Go-Go Potassium Channels/metabolism , Amino Acid Sequence , Animals , Calmodulin/genetics , Ether-A-Go-Go Potassium Channels/genetics , Humans , Protein Domains , Sequence Deletion , Xenopus laevisABSTRACT
Heme iron has many and varied roles in biology. Most commonly it binds as a prosthetic group to proteins, and it has been widely supposed and amply demonstrated that subtle variations in the protein structure around the heme, including the heme ligands, are used to control the reactivity of the metal ion. However, the role of heme in biology now appears to also include a regulatory responsibility in the cell; this includes regulation of ion channel function. In this work, we show that cardiac KATP channels are regulated by heme. We identify a cytoplasmic heme-binding CXXHX16H motif on the sulphonylurea receptor subunit of the channel, and mutagenesis together with quantitative and spectroscopic analyses of heme-binding and single channel experiments identified Cys628 and His648 as important for heme binding. We discuss the wider implications of these findings and we use the information to present hypotheses for mechanisms of heme-dependent regulation across other ion channels.
Subject(s)
Heme/metabolism , KATP Channels/metabolism , Sulfonylurea Receptors/chemistry , Amino Acid Motifs/genetics , Animals , Cell Line , HEK293 Cells , Humans , KATP Channels/genetics , Myocardium/metabolism , Protein Binding/genetics , Protein Structure, Tertiary , Rats , Rats, Wistar , Sulfonylurea Receptors/geneticsABSTRACT
BACKGROUND: Ivabradine is a specific bradycardic agent used in coronary artery disease and heart failure, lowering heart rate through inhibition of sinoatrial nodal HCN-channels. This study investigated the propensity of ivabradine to interact with KCNH2-encoded human Ether-à-go-go-Related Gene (hERG) potassium channels, which strongly influence ventricular repolarization and susceptibility to torsades de pointes arrhythmia. METHODS AND RESULTS: Patch clamp recordings of hERG current (IhERG) were made from hERG expressing cells at 37°C. Ih ERG was inhibited with an IC50 of 2.07 µmol/L for the hERG 1a isoform and 3.31 µmol/L for coexpressed hERG 1a/1b. The voltage and time-dependent characteristics of Ih ERG block were consistent with preferential gated-state-dependent channel block. Inhibition was partially attenuated by the N588K inactivation-mutant and the S624A pore-helix mutant and was strongly reduced by the Y652A and F656A S6 helix mutants. In docking simulations to a MthK-based homology model of hERG, the 2 aromatic rings of the drug could form multiple π-π interactions with the aromatic side chains of both Y652 and F656. In monophasic action potential (MAP) recordings from guinea-pig Langendorff-perfused hearts, ivabradine delayed ventricular repolarization and produced a steepening of the MAPD90 restitution curve. CONCLUSIONS: Ivabradine prolongs ventricular repolarization and alters electrical restitution properties at concentrations relevant to the upper therapeutic range. In absolute terms ivabradine does not discriminate between hERG and HCN channels: it inhibits Ih ERG with similar potency to that reported for native If and HCN channels, with S6 binding determinants resembling those observed for HCN4. These findings may have important implications both clinically and for future bradycardic drug design.
Subject(s)
Benzazepines/pharmacology , Ether-A-Go-Go Potassium Channels/antagonists & inhibitors , Potassium Channel Blockers/pharmacology , Animals , Bradycardia/drug therapy , ERG1 Potassium Channel , Ether-A-Go-Go Potassium Channels/drug effects , Guinea Pigs , HEK293 Cells , Heart/drug effects , Humans , Ivabradine , Male , Patch-Clamp TechniquesABSTRACT
KV11.1 (hERG1) channels are often overexpressed in human cancers. In leukemias, KV11.1 regulates pro-survival signals that promote resistance to chemotherapy, raising the possibility that inhibitors of KV11.1 could be therapeutically beneficial. However, because of the role of KV11.1 in cardiac repolarization, blocking these channels may cause cardiac arrhythmias. We show that CD-160130, a novel pyrimido-indole compound, blocks KV11.1 channels with a higher efficacy for the KV11.1 isoform B, in which the IC50 (1.8 µM) was approximately 10-fold lower than observed in KV11.1 isoform A. At this concentration, CD-160130 also had minor effects on Kir2.1, KV 1.3, Kv1.5, and KCa3.1. In vitro, CD-160130 induced leukemia cell apoptosis, and could overcome bone marrow mesenchymal stromal cell (MSC)-induced chemoresistance. This effect was caused by interference with the survival signaling pathways triggered by MSCs. In vivo, CD-160130 produced an antileukemic activity, stronger than that caused by cytarabine. Consistent with its atypical target specificity, CD-160130 did not bind to the main binding site of the arrhythmogenic KV11.1 blockers (the Phe656 pore residue). Importantly, in guinea pigs CD-160130 produced neither alteration of the cardiac action potential shape in dissociated cardiomyocytes nor any lengthening of the QT interval in vivo. Moreover, CD-160130 had no myelotoxicity on human bone marrow-derived cells. Therefore, CD-160130 is a promising first-in-class compound to attempt oncologic therapy without cardiotoxicity, based on targeting KV11.1. Because leukemia and cardiac cells tend to express different ratios of the A and B KV11.1 isoforms, the pharmacological properties of CD-160130 may depend, at least in part, on isoform specificity.
Subject(s)
Antineoplastic Agents/pharmacology , Cardiotoxins , Ether-A-Go-Go Potassium Channels/antagonists & inhibitors , Indoles/pharmacology , Pyrimidines/chemistry , Pyrimidines/pharmacology , Pyrimidinones/pharmacology , Animals , Antineoplastic Agents/chemistry , Antineoplastic Agents/therapeutic use , CHO Cells , Cell Line, Tumor , Cricetinae , Cricetulus , ERG1 Potassium Channel , Ether-A-Go-Go Potassium Channels/metabolism , Female , Guinea Pigs , HEK293 Cells , HL-60 Cells , Humans , Indoles/chemistry , Indoles/therapeutic use , Leukemia, B-Cell/drug therapy , Leukemia, B-Cell/metabolism , Leukemia, B-Cell/pathology , Male , Mice , Protein Isoforms/antagonists & inhibitors , Protein Isoforms/metabolism , Pyrimidines/therapeutic use , Pyrimidinones/chemistry , Pyrimidinones/therapeutic use , Treatment Outcome , Xenograft Model Antitumor Assays/methodsABSTRACT
Both human ether-à-go-go-related gene (hERG1) and the closely related human ether-à-go-go (hEAG1) channel are aberrantly expressed in a large proportion of human cancers. In the present study, we demonstrate that transfection of hERG1 into mouse fibroblasts is sufficient to induce many features characteristic of malignant transformation. An important finding of this work is that this transformation could be reversed by chronic incubation (for 2-3 weeks) with the hERG channel blocker dofetilide (100 nM), whereas more acute applications (for 1-2 days) were ineffective. The hERG1 expression resulted in a profound loss of cell contact inhibition, multiple layers of overgrowing cells, and high saturation densities. Cells also changed from fibroblast-like to a more spindle-shaped morphology, which was associated with a smaller cell size, a dramatic increase in cell polarization, a reduction in the number of actin stress fibers, and less punctate labeling of focal adhesions. Analysis of single-cell migration and scratch-wound closure clearly demonstrated that hERG1-expressing cells migrated more rapidly than vector-transfected control cells. In contrast to previous studies on hEAG1, there were no increases in rates of proliferation, or loss of growth factor dependency; however, hERG1-expressing cells were capable of substrate-independent growth. Allogeneic transplantation of hERG1-expressing cells into nude mice resulted in an increased incidence of tumors. In contrast to hEAG1, the mechanism of cellular transformation is dependent on ion conduction. Trafficking-deficient and conduction-deficient hERG1 mutants also prevented cellular transformation. These results provide evidence that hERG1 expression is sufficient to induce cellular transformation by a mechanism distinct from hEAG1. The most important conclusion of this study is that selective hERG1 channel blockers have therapeutic potential in the treatment of hERG1-expressing cancers.
Subject(s)
Cell Transformation, Neoplastic/drug effects , Ether-A-Go-Go Potassium Channels/antagonists & inhibitors , Ether-A-Go-Go Potassium Channels/metabolism , Potassium Channel Blockers/pharmacology , Actins/metabolism , Animals , Cell Line , Cell Movement/drug effects , Cell Proliferation/drug effects , Cell Transformation, Neoplastic/metabolism , Fibroblasts/drug effects , Focal Adhesions/metabolism , Humans , Mice , Mice, Nude , NIH 3T3 Cells , Stress Fibers/metabolism , TransfectionABSTRACT
Molecular knowledge of hERG blocking liability can offer the possibility of optimizing lead compounds in a way that eliminates potentially lethal side effects. In this study, we computationally designed, synthesized, and tested a small series of "minimally structured" molecules. Some of these compounds were remarkably potent against hERG (6, IC(50) = 2.4 nM), allowing us to identify the minimal structural requirements for hERG blocking liability.
Subject(s)
Ether-A-Go-Go Potassium Channels/drug effects , Potassium Channel Blockers/chemical synthesis , Drug Design , Humans , Long QT Syndrome/chemically induced , Potassium Channel Blockers/adverse effects , Quantitative Structure-Activity RelationshipABSTRACT
Human ether-à-go-go-related gene (hERG) K(+) channels have a critical role in cardiac repolarization. hERG channels close (deactivate) very slowly, and this is vital for regulating the time course and amplitude of repolarizing current during the cardiac action potential. Accelerated deactivation is one mechanism by which inherited mutations cause long QT syndrome and potentially lethal arrhythmias. hERG deactivation is highly dependent upon an intact EAG domain (the first 135 amino acids of the N terminus). Importantly, deletion of residues 2-26 accelerates deactivation to a similar extent as removing the entire EAG domain. These and other experiments suggest the first 26 residues (NT1-26) contain structural elements required to slow deactivation by stabilizing the open conformation of the pore. Residues 26-135 form a Per-Arnt-Sim domain, but a structure for NT1-26 has not been forthcoming, and little is known about its site of interaction on the channel. In this study, we present an NMR structure for the entire EAG domain, which reveals that NT1-26 is structurally independent from the Per-Arnt-Sim domain and contains a stable amphipathic helix with one face being positively charged. Mutagenesis and electrophysiological studies indicate that neutralizing basic residues and breaking the amphipathic helix dramatically accelerate deactivation. Furthermore, scanning mutagenesis and molecular modeling studies of the cyclic nucleotide binding domain suggest that negatively charged patches on its cytoplasmic surface form an interface with the NT1-26 domain. We propose a model in which NT1-26 obstructs gating motions of the cyclic nucleotide binding domain to allosterically stabilize the open conformation of the pore.
Subject(s)
Ether-A-Go-Go Potassium Channels/chemistry , Ion Channel Gating , Models, Molecular , Amino Acid Substitution , ERG1 Potassium Channel , Ether-A-Go-Go Potassium Channels/genetics , Ether-A-Go-Go Potassium Channels/metabolism , Humans , Mutagenesis , Mutation, Missense , Nuclear Magnetic Resonance, Biomolecular , Protein Structure, TertiaryABSTRACT
The hERG (human ether-à-go-go related gene) channel is a member of the eag voltage-gated K(+) channel family. In common with other members of this family, it has a subunit topology of six trans-membrane helices that tetramerise to form a functional ion-channel. In addition, hERG has an N-terminal PAS (Per, Arnt and Sim) domain and a C-terminal cyclic nucleotide binding domain (cNBD). Both these cytosolic domains are involved in regulation of the gating of the ion channel as demonstrated by inheritable mutations in these domains that result in either a loss, or a gain, in function. Here we report near complete backbone and side chain (15)N, (13)C and (1)H assignments for the N-terminal domain (residues 1-135) including the functionally critical first 26 residues. Comparison with the secondary structure of the crystal structure (residues 26-135) suggests that the solution and crystal structures are very similar except that the solution structure contains an additional helix between residues 12-23; a region of the protein important for channel gating.
Subject(s)
Ether-A-Go-Go Potassium Channels/chemistry , Nuclear Magnetic Resonance, Biomolecular , Amino Acid Sequence , Humans , Molecular Sequence Data , Protein Structure, Secondary , Protein Structure, TertiaryABSTRACT
Potassium (K (+)) channels can regulate ionic conduction through their pore by a mechanism, involving the selectivity filter, known as C-type inactivation. This process is rapid in the hERG K (+) channel and is fundamental to its physiological role. Although mutations within hERG are known to remove this process, a structural basis for the inactivation mechanism has yet to be characterized. Using MD simulations based on homology modeling, we observe that the carbonyl of the filter aromatic, Phe627, forming the S 0 K (+) binding site, swiftly rotates away from the conduction axis in the wild-type channel. In contrast, in well-characterized non-inactivating mutant channels, this conformational change occurs less frequently. In the non-inactivating channels, interactions with a water molecule located behind the selectivity filter are critical to the enhanced stability of the conducting state. We observe comparable conformational changes in the acid sensitive TASK-1 channel and propose a common mechanism in these channels for regulating efflux of K (+) ions through the selectivity filter.
Subject(s)
Hydrogen-Ion Concentration , Potassium Channels/chemistry , Potassium Channels/physiology , Amino Acid Sequence , Computer Simulation , Humans , Lipid Bilayers , Models, Molecular , Molecular Sequence Data , Phosphatidylcholines , Protein ConformationABSTRACT
hERG potassium channels have a critical role in the normal electrical activity of the heart. The block of hERG channels can cause the drug-induced form of long QT syndrome, a cardiac disorder that carries an increased risk of cardiac arrhythmias and sudden death. hERG channels are extraordinarily sensitive to block by large numbers of structurally diverse drugs. In previous years, the risk of compounds causing this cardiotoxic side effect has been a common reason for the failure of compounds in preclinical safety trials. Pharmaceutical companies have successfully utilized and developed higher throughput techniques for the early detection of compounds that block hERG, and this has helped reduce the number of compounds that fail in the late stages of development. Nevertheless, this screening-based approach is expensive, consumes chemistry resources, and bypasses the problem rather than shedding light on it. Crystal structures of potassium channels have facilitated studies into the structural basis for the gating and block of hERG channels. Most drugs bind within the inner cavity, and the individual amino acids that form the drug binding site have been identified by site-directed mutagenesis approaches. Gating processes have an important influence on the drug-binding site. Recent advances in our understanding of channel activation and inactivation are providing insight into why hERG channels are more susceptible to block than other K (+) channels. Knowledge of the structure of the drug-binding site and precise nature of interactions with drug molecules should assist efforts to develop drugs without the propensity to cause cardiac arrhythmias.
Subject(s)
Arrhythmias, Cardiac/chemically induced , Arrhythmias, Cardiac/metabolism , Cardiotoxins/toxicity , Ether-A-Go-Go Potassium Channels/chemistry , Ether-A-Go-Go Potassium Channels/metabolism , Binding Sites , Cardiotoxins/chemistry , Ether-A-Go-Go Potassium Channels/antagonists & inhibitors , Humans , Pharmaceutical Preparations/chemistry , Protein ConformationABSTRACT
The opening of ion channels is proposed to arise from bending of the pore inner helices that enables them to pivot away from the central axis creating a cytosolic opening for ion diffusion. The flexibility of the inner helices is suggested to occur either at a conserved glycine located adjacent to the selectivity filter (glycine gating hinge) and/or at a second site occupied by glycine or proline containing motifs. Sequence alignment with other K+ channels shows that hERG possesses glycine residues (Gly648 and Gly657) at each of these putative hinge sites. In apparent contrast to the hinge hypotheses, substitution of both glycine residues for alanine causes little effect on either the voltage-dependence or kinetics of channel activation, and open state block by intracellular blockers. Substitution of the glycines with larger hydrophobic residues causes a greater propensity for the channel to open. We propose that in contrast to Shaker the pore of hERG is intrinsically more stable in the open than the closed conformation and that substitution at Gly648 or Gly657 further shifts the gating equilibrium to favor the open state. Molecular dynamics simulations indicate the S6 helices of hERG are inherently flexible, even in the absence of the glycine residues. Thus hERG activation gating exhibits important differences to other Kv channels. Our findings indicate that the hERG inner helix glycine residues are required for the tight packing of the channel helices and that the flexibility afforded by glycine or proline residues is not universally required for activation gating.
Subject(s)
Ether-A-Go-Go Potassium Channels/chemistry , Ether-A-Go-Go Potassium Channels/metabolism , Amino Acid Sequence , Amino Acid Substitution , Animals , Ether-A-Go-Go Potassium Channels/genetics , Humans , Molecular Sequence Data , Mutagenesis, Site-Directed , Protein Structure, Secondary , Sequence Alignment , Thermodynamics , Xenopus laevisABSTRACT
Many commonly used, structurally diverse, drugs block the human ether-a-go-go-related gene (hERG) K(+) channel to cause acquired long QT syndrome, which can lead to sudden death via lethal cardiac arrhythmias. This undesirable side effect is a major hurdle in the development of safe drugs. To gain insight about the structure of hERG and the nature of drug block we have produced structural models of the channel pore domain, into each of which we have docked a set of 20 hERG blockers. In the absence of an experimentally determined three-dimensional structure of hERG, each of the models was validated against site-directed mutagenesis data. First, hERG models were produced of the open and closed channel states, based on homology with the prokaryotic K(+) channel crystal structures. The modeled complexes were in partial agreement with the mutagenesis data. To improve agreement with mutagenesis data, a KcsA-based model was refined by rotating the four copies of the S6 transmembrane helix half a residue position toward the C-terminus, so as to place all residues known to be involved in drug binding in positions lining the central cavity. This model produces complexes that are consistent with mutagenesis data for smaller, but not larger, ligands. Larger ligands could be accommodated following refinement of this model by enlarging the cavity using the inherent flexibility about the glycine hinge (Gly648) in S6, to produce results consistent with the experimental data for the majority of ligands tested.
Subject(s)
Ether-A-Go-Go Potassium Channels/antagonists & inhibitors , Ether-A-Go-Go Potassium Channels/chemistry , Potassium Channel Blockers/pharmacology , Amino Acid Sequence , Amino Acid Substitution , Crystallography, X-Ray , ERG1 Potassium Channel , Ether-A-Go-Go Potassium Channels/genetics , Heart/physiology , Humans , Models, Molecular , Molecular Sequence Data , Neurons/physiology , Potassium Channel Blockers/chemistry , Protein Conformation , Sequence Alignment , Sequence Homology, Amino AcidABSTRACT
The human ether-a-go-go-related gene (hERG) encodes the pore-forming alpha-subunit of a voltage-gated potassium (K(+)) channel. A variety of unrelated compounds reduce K(+ )current in the heart by blocking the pore or disrupting trafficking of the hERG channel to the membrane surface. This induces a syndrome known as long QT, which arises from abnormalities in action potential repolarisation and can degenerate into lethal cardiac arrhythmias. As a result, this undesirable side effect has severely hindered safe drug development. This review describes progress in understanding the molecular basis for drug binding to hERG, outlines the characteristics of hERG ligands and discusses experimental and in silico approaches for identifying compounds with QT liabilities. Recent developments should enable recognition of hERG-positive compounds at the early stages of their development.