Ligand activation mechanisms of human KCNQ2 channel.

The human voltage-gated potassium channel KCNQ2/KCNQ3 carries the neuronal M-current, which helps to stabilize the membrane potential. KCNQ2 can be activated by analgesics and antiepileptic drugs but their activation mechanisms remain unclear. Here we report cryo-electron microscopy (cryo-EM) structures of human KCNQ2-CaM in complex with three activators, namely the antiepileptic drug cannabidiol (CBD), the lipid phosphatidylinositol 4,5-bisphosphate (PIP2), and HN37 (pynegabine), an antiepileptic drug in the clinical trial, in an either closed or open conformation. The activator-bound structures, along with electrophysiology analyses, reveal the binding modes of two CBD, one PIP2, and two HN37 molecules in each KCNQ2 subunit, and elucidate their activation mechanisms on the KCNQ2 channel. These structures may guide the development of antiepileptic drugs and analgesics that target KCNQ2.

Ancient medicinal plant rosemary contains a highly efficacious and isoform-selective KCNQ potassium channel opener.

Voltage-gated potassium (Kv) channels in the KCNQ subfamily serve essential roles in the nervous system, heart, muscle and epithelia. Different heteromeric KCNQ complexes likely serve distinct functions in the brain but heteromer subtype-specific small molecules for research or therapy are lacking. Rosemary (Salvia rosmarinus) is an evergreen plant used medicinally for millennia for neurological and other disorders. Here, we report that rosemary extract is a highly efficacious opener of heteromeric KCNQ3/5 channels, with weak effects on KCNQ2/3. Using functional screening we find that carnosic acid, a phenolic diterpene from rosemary, is a potent, highly efficacious, PIP2 depletion-resistant KCNQ3 opener with lesser effects on KCNQ5 and none on KCNQ1 or KCNQ2. Carnosic acid is also highly selective for KCNQ3/5 over KCNQ2/3 heteromers. Medicinal chemistry, in silico docking, and mutagenesis reveal that carboxylate-guanidinium ionic bonding with an S4-5 linker arginine underlies the KCNQ3 opening proficiency of carnosic acid, the effects of which on KCNQ3/5 suggest unique therapeutic potential and a molecular basis for ancient neurotherapeutic use of rosemary.

Intracellular zinc protects Kv7 K+ channels from Ca2+/calmodulin-mediated inhibition.

Zinc (Zn) is an essential trace element; it serves as a cofactor for a great number of enzymes, transcription factors, receptors, and other proteins. Zinc is also an important signaling molecule, which can be released from intracellular stores into the cytosol or extracellular space, for example, during synaptic transmission. Amongst cellular effects of zinc is activation of Kv7 (KCNQ, M-type) voltage-gated potassium channels. Here, we investigated relationships between Kv7 channel inhibition by Ca2+/calmodulin (CaM) and zinc-mediated potentiation. We show that Zn2+ ionophore, zinc pyrithione (ZnPy), can prevent or reverse Ca2+/CaM-mediated inhibition of Kv7.2. In the presence of both Ca2+ and Zn2+, the Kv7.2 channels lose most of their voltage dependence and lock in an open state. In addition, we demonstrate that mutations that interfere with CaM binding to Kv7.2 and Kv7.3 reduced channel membrane abundance and activity, but these mutants retained zinc sensitivity. Moreover, the relative efficacy of ZnPy to activate these mutants was generally greater, compared with the WT channels. Finally, we show that zinc sensitivity was retained in Kv7.2 channels assembled with mutant CaM with all four EF hands disabled, suggesting that it is unlikely to be mediated by CaM. Taken together, our findings indicate that zinc is a potent Kv7 stabilizer, which may protect these channels from physiological inhibitory effects of neurotransmitters and neuromodulators, protecting neurons from overactivity.

Triclosan is a KCNQ3 potassium channel activator.

KCNQ channels participate in the physiology of several cell types. In neurons of the central nervous system, the primary subunits are KCNQ2, 3, and 5. Activation of these channels silence the neurons, limiting action potential duration and preventing high-frequency action potential burst. Loss-of-function mutations of the KCNQ channels are associated with a wide spectrum of phenotypes characterized by hyperexcitability. Hence, pharmacological activation of these channels is an attractive strategy to treat epilepsy and other hyperexcitability conditions as are the evolution of stroke and traumatic brain injury. In this work we show that triclosan, a bactericide widely used in personal care products, activates the KCNQ3 channels but not the KCNQ2. Triclosan induces a voltage shift in the activation, increases the conductance, and slows the closing of the channel. The response is independent of PIP2. Molecular docking simulations together with site-directed mutagenesis suggest that the putative binding site is in the voltage sensor domain. Our results indicate that triclosan is a new activator for KCNQ channels.

A novel KCNQ2 missense variant in non-syndromic intellectual disability causes mild gain-of-function of Kv7.2 channel.

BACKGROUND: Heterozygous variants of KCNQ2 can cause KCNQ2 associated neurodevelopmental disorder, mainly are benign (familial) neonatal or infantile epilepsy (B(F)NE or B(F)IE) and developmental epileptic encephalopathy(DEE). Moreover, some intermediate phenotypes, including intellectual disability (ID), and myokymia are related to the gene. METHODS: We collected a non-syndromic ID male patient with a novel KCNQ2 missense variant. Whole cell electrophysiology, western blotting, and immunofluorescence were adopted to analyze the variant's functional alterations. RESULTS: The patient presented with global developmental delay since his infancy. He still had profound ID but did not have epilepsy at the adolescence. The de novo KCNQ2 variant p.R75C (NM_172107) in the NH2 domain identified here showed a slightly hyperpolarized shift of activation curves and larger current density in homomeric configurations, which could be abolished in co-expression with Kv7.2 or Kv7.3 wild-type. Western blotting and immunocytochemistry supported that the expression of variant p.R75C is lower than the Kv7.2 wild-type. The findings indicated variant p.R75C causes mild gain-of-function (GOF) of Kv7.2 channel. CONCLUSIONS: We report a non-syndromic ID patient with a KCNQ2 mild GOF variant, adding evidence for this rare clinical phenotype in the disorder. We propose that individuals with KCNQ2 GOF variants are prone to have cognitive impairments.

KCNQ2 Selectivity Filter Mutations Cause Kv7.2 M-Current Dysfunction and Configuration Changes Manifesting as Epileptic Encephalopathies and Autistic Spectrum Disorders.

KCNQ2 mutations can cause benign familial neonatal convulsions (BFNCs), epileptic encephalopathy (EE), and mild-to-profound neurodevelopmental disabilities. Mutations in the KCNQ2 selectivity filter (SF) are critical to neurodevelopmental outcomes. Three patients with neonatal EE carry de novo heterozygous KCNQ2 p.Thr287Ile, p.Gly281Glu and p.Pro285Thr, and all are followed-up in our clinics. Whole-cell patch-clamp analysis with transfected mutations was performed. The Kv7.2 in three mutations demonstrated significant current changes in the homomeric-transfected cells. The conduction curves for V1/2, the K slope, and currents in 3 mutations were lower than those for the wild type (WT). The p.Gly281Glu had a worse conductance than the p.Thr287Ile and p.Pro285Thr, the patient compatible with p.Gly281Glu had a worse clinical outcome than patients with p.Thr287Ile and p.Pro285Thr. The p.Gly281Glu had more amino acid weight changes than the p.Gly281Glu and p.Pro285Thr. Among 5 BFNCs and 23 EE from mutations in the SF, the greater weight of the mutated protein compared with that of the WT was presumed to cause an obstacle to pore size, which is one of the most important factors in the phenotype and outcome. For the 35 mutations in the SF domain, using changes in amino acid weight between the WT and the KCNQ2 mutations to predict EE resulted in 80.0% sensitivity and 80% specificity, a positive prediction rate of 96.0%, and a negative prediction rate of 40.0% (p = 0.006, χ2 (1, n = 35) = 7.56; odds ratio 16.0, 95% confidence interval, 1.50 to 170.63). The findings suggest that p.Thr287Ile, p.Gly281Glu and p.Pro285Thr are pathogenic to KCNQ2 EE. In mutations in SF, a mutated protein heavier than the WT is a factor in the Kv7.2 current and outcome.

Molecular basis for ligand activation of the human KCNQ2 channel.

The voltage-gated potassium channel KCNQ2 is responsible for M-current in neurons and is an important drug target to treat epilepsy, pain and several other diseases related to neuronal hyper-excitability. A list of synthetic compounds have been developed to directly activate KCNQ2, yet our knowledge of their activation mechanism is limited, due to lack of high-resolution structures. Here, we report cryo-electron microscopy (cryo-EM) structures of the human KCNQ2 determined in apo state and in complex with two activators, ztz240 or retigabine, which activate KCNQ2 through different mechanisms. The activator-bound structures, along with electrophysiology analysis, reveal that ztz240 binds at the voltage-sensing domain and directly stabilizes it at the activated state, whereas retigabine binds at the pore domain and activates the channel by an allosteric modulation. By accurately defining ligand-binding sites, these KCNQ2 structures not only reveal different ligand recognition and activation mechanisms, but also provide a structural basis for drug optimization and design.

Heteromeric Assembly of Truncated Neuronal Kv7 Channels: Implications for Neurologic Disease and Pharmacotherapy.

Neuronal voltage-gated potassium channels (Kv) are critical regulators of electrical activity in the central nervous system. Mutations in the KCNQ (Kv7) ion channel family are linked to epilepsy and neurodevelopmental disorders. These channels underlie the neuronal "M-current" and cluster in the axon initial segment to regulate the firing of action potentials. There is general consensus that KCNQ channel assembly and heteromerization are controlled by C-terminal helices. We identified a pediatric patient with neurodevelopmental disability, including autism traits, inattention and hyperactivity, and ataxia, who carries a de novo frameshift mutation in KCNQ3 (KCNQ3-FS534), leading to truncation of ∼300 amino acids in the C terminus. We investigated possible molecular mechanisms of channel dysfunction, including haplo-insufficiency or a dominant-negative effect caused by the assembly of truncated KCNQ3 and functional KCNQ2 subunits. We also used a recently recognized property of the KCNQ2-specific activator ICA-069673 to identify assembly of heteromeric channels. ICA-069673 exhibits a functional signature that depends on the subunit composition of KCNQ2/3 channels, allowing us to determine whether truncated KCNQ3 subunits can assemble with KCNQ2. Our findings demonstrate that although the KCNQ3-FS534 mutant does not generate functional channels on its own, large C-terminal truncations of KCNQ3 (including the KCNQ3-FS534 mutation) assemble efficiently with KCNQ2 but fail to promote or stabilize KCNQ2/KCNQ3 heteromeric channel expression. Therefore, the frequent assumption that pathologies linked to KCNQ3 truncations arise from haplo-insufficiency should be reconsidered in some cases. Subtype-specific channel activators like ICA-069673 are a reliable tool to identify heteromeric assembly of KCNQ2 and KCNQ3. SIGNIFICANCE STATEMENT: Mutations that truncate the C terminus of neuronal Kv7/KCNQ channels are linked to a spectrum of seizure disorders. One role of the multifunctional KCNQ C terminus is to mediate subtype-specific assembly of heteromeric KCNQ channels. This study describes the use of a subtype-specific Kv7 activator to assess assembly of heteromeric KCNQ2/KCNQ3 (Kv7.2/Kv7.3) channels and demonstrates that large disease-linked and experimentally generated C-terminal truncated KCNQ3 mutants retain the ability to assemble with KCNQ2.

Molecular Mechanisms and Structural Basis of Retigabine Analogues in Regulating KCNQ2 Channel.

KCNQ2 channel is one of the important members of potassium voltage-gated channel. KCNQ2 is closely related to neuronal excitatory diseases including epilepsy and neuropathic pain, and also acts as a drug target of the anti-epileptic drug, retigabine (RTG). In the past few decades, RTG has shown strong efficacy in the treatment of refractory epilepsy but has been withdrawn from clinical use due to its multiple adverse effects in clinical phase III trials. To overcome the drawbacks of RTG, several RTG analogues have been developed with different activation potency to KCNQ2. However, the detailed molecular mechanism by which these RTG analogues regulate KCNQ2 channel remains obscure. In this study, we used molecular simulations to analyse the interaction mode between the RTG analogues and KCNQ2, and to determine their molecular mechanism of action. Our data show that the van der Waals interactions, hydrophobic interactions, hydrogen bond, halogen bond, and π-π stacking work together to maintain the binding stability of the drugs in the binding pocket. On an atomic scale, the amide group in the carbamate and the amino group in the 2-aminophenyl moiety of RTG and RL648_81 are identified as key interaction sites. Our finding provides insight into the molecular mechanism by which KCNQ2 channels are regulated by RTG analogues. It also provides direct theoretical support for optimizing design of the KCNQ2 channel openers in the future, which will help treat refractory epilepsy caused by nerve excitability.

In silico re-engineering of a neurotransmitter to activate KCNQ potassium channels in an isoform-specific manner.

Voltage-gated potassium (Kv) channel dysfunction causes a variety of inherited disorders, but developing small molecules that activate Kv channels has proven challenging. We recently discovered that the inhibitory neurotransmitter γ-aminobutyric acid (GABA) directly activates Kv channels KCNQ3 and KCNQ5. Here, finding that inhibitory neurotransmitter glycine does not activate KCNQs, we re-engineered it in silico to introduce predicted KCNQ-opening properties, screened by in silico docking, then validated the hits in vitro. Attaching a fluorophenyl ring to glycine optimized its electrostatic potential, converting it to a low-nM affinity KCNQ channel activator. Repositioning the phenyl ring fluorine and/or adding a methylsulfonyl group increased the efficacy of the re-engineered glycines and switched their target KCNQs. Combining KCNQ2- and KCNQ3-specific glycine derivatives synergistically potentiated KCNQ2/3 activation by exploiting heteromeric channel composition. Thus, in silico optimization and docking, combined with functional screening of only three compounds, facilitated re-engineering of glycine to develop several potent KCNQ activators.

Epileptic Encephalopathy In A Patient With A Novel Variant In The Kv7.2 S2 Transmembrane Segment: Clinical, Genetic, and Functional Features.

Kv7.2 subunits encoded by the KCNQ2 gene provide a major contribution to the M-current (IKM), a voltage-gated K+ current crucially involved in the regulation of neuronal excitability. Heterozygous missense variants in Kv7.2 are responsible for epileptic diseases characterized by highly heterogeneous genetic transmission and clinical severity, ranging from autosomal-dominant Benign Familial Neonatal Seizures (BFNS) to sporadic cases of severe epileptic and developmental encephalopathy (DEE). Here, we describe a patient with neonatal onset DEE, carrying a previously undescribed heterozygous KCNQ2 c.418G > C, p.Glu140Gln (E140Q) variant. Patch-clamp recordings in CHO cells expressing the E140Q mutation reveal dramatic loss of function (LoF) effects. Multistate structural modelling suggested that the E140Q substitution impeded an intrasubunit electrostatic interaction occurring between the E140 side chain in S2 and the arginine at position 210 in S4 (R210); this interaction is critically involved in stabilizing the activated configuration of the voltage-sensing domain (VSD) of Kv7.2. Functional results from coupled charge reversal or disulfide trapping experiments supported such a hypothesis. Finally, retigabine restored mutation-induced functional changes, reinforcing the rationale for the clinical use of Kv7 activators as personalized therapy for DEE-affected patients carrying Kv7.2 LoF mutations.

KCNQ2 related early-onset epileptic encephalopathies in Chinese children.

OBJECTIVE: To study the phenotype, genotype, treatment strategies, and short-term prognosis of Chinese children with KCNQ2 (potassium voltage-gated channel subfamily Q member 2) related early-onset epileptic encephalopathies (KCNQ2-EOEEs) in Southwest China. METHODS: We used targeted next-generation sequencing (NGS) to identify KCNQ2 variants in Chinese patients with EOEEs. And patients with KCNQ2-EOEEs were confirmed after clinical and genetic analyses. We followed them in our cohort and analyzed their clinical data. RESULTS: 122 patients with EOEEs were registered from August 2015 to October 2017, and 78 underwent targeted NGS. Seven among them were confirmed to be caused by pathogenic KCNQ2 variants, 6 of that were de novo and 1 was inherited. The median seizure onset age of the 7 patients was 5 days. Tonic-clonic and tonic seizures were the major seizure types; the electroencephalograms of all patients showed multifocal sharp waves initially. When new seizure types appeared in infancy, the most common type was epileptic spasm. At the last follow-up, seizures persisted in only one patient, and another patient had seizure recurrence. The identified pathogenic KCNQ2 variants introduced amino acid missense changes, or in one instance, frameshift variant, four of which have not been reported. Valproic acid (VPA) was effective as concomitant treatment in three patients, and all patients had intellectual/developmental disabilities (IDDs). CONCLUSIONS: The KCNQ2 missense variant plays an important role in EOEE pathogenesis, and patients with KCNQ2-EOEEs mainly present with intractable seizures and IDDs. Moreover, VPA has potential as an effective therapeutic strategy.

Homomeric Kv7.2 current suppression is a common feature in KCNQ2 epileptic encephalopathy.

OBJECTIVE: To gain insight into the mechanisms underlying KCNQ2 encephalopathy by examining the electrophysiologic properties of mutant Kv7.2 channels in different multimeric configurations. METHODS: We analyzed the genotype-phenotype relationship in 4 patients with KCNQ2 encephalopathy and performed electrophysiologic analysis of M-currents mediated by homomeric Kv7.2 or heteromeric Kv7.2/Kv7.3 channels. RESULTS: Negligible or no current was recorded in cells expressing homomeric E130K, W270R, or G281R de novo mutants, and it was reduced by more than 90% for the L243F maternally inherited mutant. The E130K and G281R mutants presented a marked dominant-negative behavior, whereas the current density was partially reduced (L243F) or not affected (W270R) when coexpressed with wild-type Kv7.2 subunits. In contrast, the extent of Kv7.3 "rescue," which yields negligible currents on its own, followed the sequence E130K > L243F > W270R, whereas no rescue was observed with the G281R mutant. No significant effects on current density were observed when subunits were expressed in a 0.5:0.5:1.0 (Kv7.2:mutant:Kv7.3) DNA ratio to mimic the genetic balance. There was an increase in sensitivity to phosphatidylinositol 4,5-bisphosphate (PIP2 ) depletion for W270R/Kv7.3, but no substantial differences were observed when the mutated subunits were coexpressed with Kv7.2 or both Kv7.2 and Kv7.3. SIGNIFICANCE: There was a marked disparity of the impact of these mutations on Kv7.2 function, which varied on association with Kv7.2 or Kv7.3 subunits. Current density of homomeric channels was the most reliable property relating Kv7.2 function to encephalopathy, but other factors are required to explain the milder phenotype for some individuals carrying the maternally inherited L243F mutation. We hypothesize that the role of homomeric Kv7.2 channels for fine-tuning neuronal connections during development is critical for the severity of the KCNQ2 encephalopathy.

Lack of correlation between surface expression and currents in epileptogenic AB-calmodulin binding domain Kv7.2 potassium channel mutants.

Heteromers of Kv7.2/Kv7.3 subunits constitute the main substrate of the neuronal M-current that limits neuronal hyper-excitability and firing frequency. Calmodulin (CaM) binding is essential for surface expression of Kv7 channels, and disruption of this interaction leads to diseases ranging from mild epilepsy to early onset encephalopathy. In this study, we addressed the impact of a charge neutralizing mutation located at the periphery of helix B (K526N). We found that, CaM binding and surface expression was impaired, although current amplitude was not altered. Currents were reduced at a faster rate after activation of a voltage-dependent phosphatase, suggesting that phosphatidylinositol-4,5-bisphosphate (PIP2) binding was weaker. In contrast, a charge neutralizing mutation located at the periphery of helix A (R333Q) did not affect CaM binding, but impaired trafficking and led to a reduction in current amplitude. Taken together, these results suggest that disruption of CaM-dependent or CaM-independent trafficking of Kv7.2/Kv7.3 channels can lead to pathology regardless of the consequences on the macroscopic ionic flow through the channel.

Reduced axonal surface expression and phosphoinositide sensitivity in Kv7 channels disrupts their function to inhibit neuronal excitability in Kcnq2 epileptic encephalopathy.

Neuronal Kv7/KCNQ channels are voltage-gated potassium channels composed of Kv7.2/KCNQ2 and Kv7.3/KCNQ3 subunits. Enriched at the axonal membrane, they potently suppress neuronal excitability. De novo and inherited dominant mutations in Kv7.2 cause early onset epileptic encephalopathy characterized by drug resistant seizures and profound psychomotor delay. However, their precise pathogenic mechanisms remain elusive. Here, we investigated selected epileptic encephalopathy causing mutations in calmodulin (CaM)-binding helices A and B of Kv7.2. We discovered that R333W, K526N, and R532W mutations located peripheral to CaM contact sites decreased axonal surface expression of heteromeric channels although only R333W mutation reduced CaM binding to Kv7.2. These mutations also altered gating modulation by phosphatidylinositol 4,5-bisphosphate (PIP2), revealing novel PIP2 binding residues. While these mutations disrupted Kv7 function to suppress excitability, hyperexcitability was observed in neurons expressing Kv7.2-R532W that displayed severe impairment in voltage-dependent activation. The M518 V mutation at the CaM contact site in helix B caused most defects in Kv7 channels by severely reducing their CaM binding, K+ currents, and axonal surface expression. Interestingly, the M518 V mutation induced ubiquitination and accelerated proteasome-dependent degradation of Kv7.2, whereas the presence of Kv7.3 blocked this degradation. Furthermore, expression of Kv7.2-M518V increased neuronal death. Together, our results demonstrate that epileptic encephalopathy mutations in helices A and B of Kv7.2 cause abnormal Kv7 expression and function by disrupting Kv7.2 binding to CaM and/or modulation by PIP2. We propose that such multiple Kv7 channel defects could exert more severe impacts on neuronal excitability and health, and thus serve as pathogenic mechanisms underlying Kcnq2 epileptic encephalopathy.

Calmodulin: A Multitasking Protein in Kv7.2 Potassium Channel Functions.

The ubiquitous calcium transducer calmodulin (CaM) plays a pivotal role in many cellular processes, regulating a myriad of structurally different target proteins. Indeed, it is unquestionable that CaM is the most relevant transductor of calcium signals in eukaryotic cells. During the last two decades, different studies have demonstrated that CaM mediates the modulation of several ion channels. Among others, it has been indicated that Kv7.2 channels, one of the members of the voltage gated potassium channel family that plays a critical role in brain excitability, requires CaM binding to regulate the different mechanisms that govern its functions. The purpose of this review is to provide an overview of the most recent advances in structure⁻function studies on the role of CaM regulation of Kv7.2 and the other members of the Kv7 family.

A Calmodulin C-Lobe Ca2+-Dependent Switch Governs Kv7 Channel Function.

Kv7 (KCNQ) voltage-gated potassium channels control excitability in the brain, heart, and ear. Calmodulin (CaM) is crucial for Kv7 function, but how this calcium sensor affects activity has remained unclear. Here, we present X-ray crystallographic analysis of CaM:Kv7.4 and CaM:Kv7.5 AB domain complexes that reveal an Apo/CaM clamp conformation and calcium binding preferences. These structures, combined with small-angle X-ray scattering, biochemical, and functional studies, establish a regulatory mechanism for Kv7 CaM modulation based on a common architecture in which a CaM C-lobe calcium-dependent switch releases a shared Apo/CaM clamp conformation. This C-lobe switch inhibits voltage-dependent activation of Kv7.4 and Kv7.5 but facilitates Kv7.1, demonstrating that mechanism is shared by Kv7 isoforms despite the different directions of CaM modulation. Our findings provide a unified framework for understanding how CaM controls different Kv7 isoforms and highlight the role of membrane proximal domains for controlling voltage-gated channel function. VIDEO ABSTRACT.

Structural basis and energy landscape for the Ca2+ gating and calmodulation of the Kv7.2 K+ channel.

The Kv7.2 (KCNQ2) channel is the principal molecular component of the slow voltage-gated, noninactivating K+ M-current, a key controller of neuronal excitability. To investigate the calmodulin (CaM)-mediated Ca2+ gating of the channel, we used NMR spectroscopy to structurally and dynamically describe the association of helices hA and hB of Kv7.2 with CaM, as a function of Ca2+ concentration. The structures of the CaM/Kv7.2-hAB complex at two different calcification states are reported here. In the presence of a basal cytosolic Ca2+ concentration (10-100 nM), only the N-lobe of CaM is Ca2+-loaded and the complex (representative of the open channel) exhibits collective dynamics on the millisecond time scale toward a low-populated excited state (1.5%) that corresponds to the inactive state of the channel. In response to a chemical or electrical signal, intracellular Ca2+ levels rise up to 1-10 µM, triggering Ca2+ association with the C-lobe. The associated conformational rearrangement is the key biological signal that shifts populations to the closed/inactive channel. This reorientation affects the C-lobe of CaM and both helices in Kv7.2, allosterically transducing the information from the Ca2+-binding site to the transmembrane region of the channel.

Migration of PIP2 on KCNQ2 Surface Revealed by Molecular Dynamics Simulations.

Lipids and membrane proteins are the main components of cell membranes. Lipid-protein interactions are dynamic because these interactions typically occur on shallow protein surface clefts. Molecular dynamics (MD) simulations provide a tool for studying the dynamics of these interactions. Here, we describe the interactions of phosphatidylinositol-4,5-bisphosphate (PIP2) with both the open and closed states of a KCNQ2 channel. Through these methods, we show that a lipid can migrate between different binding sites in a protein and this migration modulates protein functions.

Pharmacological Targeting of Neuronal Kv7.2/3 Channels: A Focus on Chemotypes and Receptor Sites.

BACKGROUND: The Kv7 (KCNQ) subfamily of voltage-gated potassium channels consists of 5 members (Kv7.1-5) each showing characteristic tissue distribution and physiological roles. Given their functional heterogeneity, Kv7 channels represent important pharmacological targets for the development of new drugs for neuronal, cardiovascular and metabolic diseases. OBJECTIVE: In the present manuscript, we focus on describing the pharmacological relevance and potential therapeutic applications of drugs acting on neuronally-expressed Kv7.2/3 channels, placing particular emphasis on the different chemotypes, and highlighting their pharmacodynamic and, whenever possible, pharmacokinetic peculiarities. METHODS: The present work is based on an in-depth search of the currently available scientific literature, and on our own experience and knowledge in the field of neuronal Kv7 channel pharmacology. Space limitations impeded to describe the full pharmacological potential of Kv7 channels; thus, we have chosen to focus on neuronal channels composed of Kv7.2 and Kv7.3 subunits, and to mainly concentrate on their involvement in epilepsy. RESULTS: An astonishing heterogeneity in the molecular scaffolds exploitable to develop Kv7.2/3 modulators is evident, with important structural/functional peculiarities of distinct compound classes. CONCLUSION: In the present work we have attempted to show the current status and growing potential of the Kv7 pharmacology field. We anticipate a bright future for the field, and express our hopes that the efforts herein reviewed will result in an improved treatment of hyperexcitability (or any other) diseases.