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BACKGROUND AND PURPOSE
APETx2, a toxin from the sea anemone Anthropleura elegantissima, inhibits acid-sensing ion channel 3 (ASIC3)-containing homo- and heterotrimeric channels with IC50 values < 100 nM and 0.1–2 µM respectively. ASIC3 channels mediate acute acid-induced and inflammatory pain response and APETx2 has been used as a selective pharmacological tool in animal studies. Toxins from sea anemones also modulate voltage-gated Na+ channel (Nav) function. Here we tested the effects of APETx2 on Nav function in sensory neurones.EXPERIMENTAL APPROACH
Effects of APETx2 on Nav function were studied in rat dorsal root ganglion (DRG) neurones by whole-cell patch clamp.KEY RESULTS
APETx2 inhibited the tetrodotoxin (TTX)-resistant Nav 1.8 currents of DRG neurones (IC50, 2.6 µM). TTX-sensitive currents were less inhibited. The inhibition of Nav 1.8 currents was due to a rightward shift in the voltage dependence of activation and a reduction of the maximal macroscopic conductance. The inhibition of Nav 1.8 currents by APETx2 was confirmed with cloned channels expressed in Xenopus oocytes. In current-clamp experiments in DRG neurones, the number of action potentials induced by injection of a current ramp was reduced by APETx2.CONCLUSIONS AND IMPLICATIONS
APETx2 inhibited Nav 1.8 channels, in addition to ASIC3 channels, at concentrations used in in vivo studies. The limited specificity of this toxin should be taken into account when using APETx2 as a pharmacological tool. Its dual action will be an advantage for the use of APETx2 or its derivatives as analgesic drugs. 相似文献3.
Sea anemones produce a myriad of toxic peptides and proteins of which a large group acts on voltage-gated Na+ channels. However, in comparison to other organisms, their venoms and toxins are poorly studied. Most of the known voltage-gated Na+ channel toxins isolated from sea anemone venoms act on neurotoxin receptor site 3 and inhibit the inactivation of these channels. Furthermore, it seems that most of these toxins have a distinct preference for crustaceans. Given the close evolutionary relationship between crustaceans and insects, it is not surprising that sea anemone toxins also profoundly affect insect voltage-gated Na+ channels, which constitutes the scope of this review. For this reason, these peptides can be considered as insecticidal lead compounds in the development of insecticides. 相似文献
4.
The K+ channel encoded by the Ether-á-go-go-Related Gene (ERG) is expressed in different tissues of different animal species. There are at least three subtypes of this channel, being the sub-type 1 (ERG1) crucial in the repolarization phase of the cardiac action potential. Mutations in this gene can affect the properties of the channel producing the type II long QT syndrome (LQTS2) and many drugs are also known to affect this channel with a similar side effect. Various scorpion, spider and sea anemone toxins affect the ERG currents by blocking the ion-conducting pore from the external side or by modulating channel gating through binding to the voltage-sensor domain. By doing so, these toxins become very useful tools for better understanding the structural and functional characteristics of these ion channels. This review discusses the interaction between the ERG channels and the peptides isolated from venoms of these animals. Special emphasis is placed on scorpion toxins, although the effects of several spider venom toxins and anemone toxins will be also revised. 相似文献
5.
Kazuo Shiomi 《Toxicon》2009,54(8):1112-546
Sea anemones are a rich source of peptide toxins acting on ion channels. Two classes of peptide toxins, site-3 sodium channel toxins and Kv1 potassium channel toxins, have been well characterized and some of them used as valuable pharmacological reagents. Recently, the following six peptides toxins, which structurally constitute a new family but target different ion channels, have been isolated: BDS-I and -II (Kv3 potassium channel toxins) from Anemonia sulcata, APETx1 (human ether-a-go-go-related gene potassium channel toxin) and APETx2 (acid-sensing sodium channel toxin) from Anthopleura elegantissima, BcIV (sodium channel toxin) from Bunodosoma caissarum and Am II (whose target is unknown) from Antheopsis maculata. In addition, the following structurally novel peptide toxins have also emerged in sea anemones: gigantoxin I (epidermal growth factor-like toxin) from Stichodactyla gigantea and acrorhagins I and II from acrorhagi (specialized aggressive organs) of Actinia equina. This review deals with the structural and functional features of these recently isolated sea anemone peptide toxins that are promising tools in studying the physiology of diverse ion channels. 相似文献
6.
Fernando Lazcano-Prez Karina Bermeo Hctor Castro Zayil Salazar Campos Isabel Arenas Ariana Zavala-Moreno Sheila Narayn Chvez-Villela Irma Jimnez Roberto Arreguín-Espinosa Reyna Fierro Humberto Gonzlez-Mrquez David E. Garcia Judith Snchez-Rodríguez 《Toxins》2022,14(4)
Sea anemones produce venoms characterized by a complex mixture of low molecular weight compounds, proteins and peptides acting on voltage-gated ion channels. Mammal sperm cells, like neurons, are characterized by their ion channels. Calcium channels seem to be implicated in pivotal roles such as motility and capacitation. In this study, we evaluated the effect of a low molecular weight fraction from the venom of the sea anemone Lebrunia neglecta on boar sperm cells and in HVA calcium channels from rat chromaffin cells. Spermatozoa viability seemed unaffected by the fraction whereas motility and sperm capacitation were notoriously impaired. The sea anemone fraction inhibited the HVA calcium current with partial recovery and no changes in chromaffin cells’ current kinetics and current–voltage relationship. These findings might be relevant to the pharmacological characterization of cnidarian venoms and toxins on voltage-gated calcium channels. 相似文献
7.
Small peptides isolated from the venom of animals are potential scaffolds for ion channel drug discovery. This review article mainly focuses on the computational studies that have advanced our understanding of how various toxins interfere with the function of K+ channels. We introduce the computational tools available for the study of toxin-channel interactions. We then discuss how these computational tools have been fruitfully applied to elucidate the mechanisms of action of a wide range of venom peptides from scorpions, spiders, and sea anemone. 相似文献
8.
《Toxin reviews》2013,32(4):451-466
During the past thirty years sea anemones turned out to be important producer of biologically highly active polypeptides acting on Na+ and K+ ion channels. They are compoused from 27 to 59 aminoacids having molecular weights from 3000 to 7000 Da. All contain six cystein molecules which are interconnected to three sisulfide bridges. Because of their specifique mode of action on nerve cell exitation and on the mammalian heart muscle they became very important tools in neurophysiology and pharmacology. In the present review the research on sea anemone polypeptid toxins is summarized from a historical point of view, focussed on the polypeptides of Anemonia sulcata, Condylactis gigantea and Anthopleura elegantissima. 相似文献
9.
The voltage-gated sodium (Nav) channel is a target for a number of drugs, insecticides, and neurotoxins. These bind to at least seven identified neurotoxin binding sites and either block conductance or modulate sodium channel gating and/or kinetics. A number of polypeptide toxins from the venoms of araneomorph and mygalomorph spiders have been isolated and characterized that interact with several of these sites. Certain huwentoxins and hainantoxins appear to target site 1 to block Nav channel conductance. The δ -atracotoxins and Magi 4 slow Nav-channel inactivation via an interaction with neurotoxin site 3. The δ -palutoxins, and most likely μ -agatoxins and curtatoxins, target site 4. However, their action is complex with the μ -agatoxins causing a hyperpolarizing shift in the voltage-dependence of activation, an action analogous to scorpion β -toxins, but with both δ -palutoxins and μ -agatoxins slowing Nav channel inactivation, a site 3-like action. Many spider toxins target undefined sites, while others are likely to cross-react with other ion channels due to conserved structures within domains of voltage-gated ion channels. It is already clear, however, that many spider toxins represent highly potent and specific molecular tools to define novel links between sites modulating channel activation and inactivation. Other spider toxins show phyla specificity and are being considered as lead compounds for the development of biopesticides. Others display tissue specificity via interactions with specific Nav channel subtypes and should provide useful tools to delineate the molecular determinants to target ligands to these channel subtypes. These studies are being greatly assisted by the determination of the pharmacophore of these toxins, but without precise identification of their binding site and mode of action their potential in the mentioned areas remains underdeveloped. 相似文献
10.
Schiavon E Pedraza-Escalona M Gurrola GB Olamendi-Portugal T Corzo G Wanke E Possani LD 《Toxicon》2012,59(2):283-293
The β-toxins purified from the New World scorpion venoms of the Centruroides species affect several voltage-gated sodium channels (VGSCs) and thus are essential tools not only for the discrimination of different channel sub-types but also for studying the structure-function relationship between channels and toxins. This communication reports the results obtained with four different peptides purified from three species of Centruroides scorpions and assayed on seven distinct isoforms of VGSC (Nav1.1-Nav1.7) by specific functional analysis conducted through single cell electrophysiology. The toxins studied were CssII from Centruroides suffusus suffusus, Cll1 and Cll2 from Centruroides limpidus limpidus and a novel toxin from Centruroides noxius, which was characterized for the first time here. It has 67 amino acid residues and four disulfide bridges with a molecular mass of 7626 Da. Three different functional features were identified: current reduction of macroscopic conductance, left shift of the voltage-dependent activation and induction of resurgent currents at negative voltages following brief, strong depolarizations. The isoforms which revealed to be more affected resulted to be Nav1.6 > 1.1 > 1.2 and, for the first time, a β-toxin is here shown to induce resurgent current also in isoforms different from Nav1.6. Additionally, these results were analyzed with molecular modelling. In conclusion, although the four toxins have a high degree of identity, they display tri-modal function, each of which shows selectivity among the different sub-types of Na+-channels. Thus, they are invaluable as tools for structure-function studies of β-toxins and offer a basis for the design of novel ion channel-specific drugs. 相似文献
11.
Mandë Holford Min-Min Zhang Layla Azam Maren Watkins Doju Yoshikami Baldomero M. Olivera 《Toxicon》2009,53(1):90-10307
Described herein is a general approach to identify novel compounds using the biodiversity of a megadiverse group of animals; specifically, the phylogenetic lineage of the venomous gastropods that belong to the genus Conus (“cone snails”). Cone snail biodiversity was exploited to identify three new μ-conotoxins, BuIIIA, BuIIIB and BuIIIC, encoded by the fish-hunting species Conus bullatus. BuIIIA, BuIIIB and BuIIIC are strikingly divergent in their amino acid composition compared to previous μ-conotoxins known to target the voltage-gated Na channel skeletal muscle subtype Nav1.4. Our preliminary results indicate that BuIIIB and BuIIIC are potent inhibitors of Nav1.4 (average block ∼96%, at a 1 μM concentration of peptide), displaying a very slow off-rate not seen in previously characterized μ-conotoxins that block Nav1.4. In addition, the three new C. bullatus μ-conopeptides help to define a new branch of the M-superfamily of conotoxins, namely M-5. The exogene strategy used to discover these Na channel-inhibiting peptides was based on both understanding the phylogeny of Conus, as well as the molecular genetics of venom μ-conotoxin peptides previously shown to generally target voltage-gated Na channels. The discovery of BuIIIA, BuIIIB and BuIIIC Na channel blockers expands the diversity of ligands useful in determining the structure-activity relationship of voltage-gated sodium channels. 相似文献
12.
C. Ian Spencer 《Toxicon》2009,53(1):78-89
Voltage-gated Na+ channels underlie the action potential upstroke in excitable cells, and both natural and synthetic inactivation inhibitors prolong the Na+ current (INa). The effects of Na+ channel mutations on these pharmacological actions are incompletely investigated. Therefore, I compared the effects of inactivation inhibitors on INa in WT or mutant (ΔKPQ) human cardiac Na+ channels expressed in HEK-293 cells, by measuring difference currents sensitive to 50 μM tetrodotoxin. Veratridine and the pyrethroid tefluthrin prolonged INa in WT and ΔKPQ without obvious differential effects, while a sea anemone toxin (ATX-II) and a synthetic inotrope (SDZ 201-106) prolonged WT INa, but apparently blocked INa in the ΔKPQ mutant. This block was due, at least in-part, to enhanced steady-state inactivation, with half-inactivation potentials shifted by up to −17 mV. Inactivation enhancement by ATX-II also persisted when conditioning depolarizations were abbreviated, and was unaffected by the additional presence of SDZ 201-106 consistent with these agents having unique interactions with ΔKPQ Na+ channels. It is concluded that the toxin-binding sites for ATX-II and SDZ 201-106 have allosteric effects converging on a common path affecting steady-state inactivation of ΔKPQ INa. Pharmacological modulation of this path to increase inactivation in mutant Na+ channels could potentially produce therapeutic benefits. 相似文献
13.
Peigneur S Billen B Derua R Waelkens E Debaveye S Béress L Tytgat J 《Biochemical pharmacology》2011,(1):81-90
Sea anemone venom is a known source of interesting bioactive compounds, including peptide toxins which are invaluable tools for studying structure and function of voltage-gated potassium channels. APEKTx1 is a novel peptide isolated from the sea anemone Anthopleura elegantissima, containing 63 amino acids cross-linked by 3 disulfide bridges. Sequence alignment reveals that APEKTx1 is a new member of the type 2 sea anemone peptides targeting voltage-gated potassium channels (KVs), which also include the kalicludines from Anemonia sulcata. Similar to the kalicludines, APEKTx1 shares structural homology with both the basic pancreatic trypsin inhibitor (BPTI), a very potent Kunitz-type protease inhibitor, and dendrotoxins which are powerful blockers of voltage-gated potassium channels. In this study, APEKTx1 has been subjected to a screening on a wide range of 23 ion channels expressed in Xenopus laevis oocytes: 13 cloned voltage-gated potassium channels (KV1.1–KV1.6, KV1.1 triple mutant, KV2.1, KV3.1, KV4.2, KV4.3, hERG, the insect channel Shaker IR), 2 cloned hyperpolarization-activated cyclic nucleotide-sensitive cation non-selective channels (HCN1 and HCN2) and 8 cloned voltage-gated sodium channels (NaV1.2–NaV1.8 and the insect channel DmNaV1). Our data show that APEKTx1 selectively blocks KV1.1 channels in a very potent manner with an IC50 value of 0.9 nM. Furthermore, we compared the trypsin inhibitory activity of this toxin with BPTI. APEKTx1 inhibits trypsin with a dissociation constant of 124 nM. In conclusion, this study demonstrates that APEKTx1 has the unique feature to combine the dual functionality of a potent and selective blocker of KV1.1 channels with that of a competitive inhibitor of trypsin. 相似文献
14.
Rebecca A Roof David L Roman Samuel T Clements Katarzyna Sobczyk-Kojiro Levi L Blazer Shodai Ota Henry I Mosberg Richard R Neubig 《BMC pharmacology》2009,9(1):1-11
Background
K+ and Na+ channel toxins constitute a large set of polypeptides, which interact with their ion channel targets. These polypeptides are classified in two different structural groups. Recently a new structural group called birtoxin-like appeared to contain both types of toxins has been described. We hypothesized that peptides of this group may contain two conserved structural motifs in K+ and/or Na+ channels scorpion toxins, allowing these birtoxin-like peptides to be active on K+ and/or Na+ channels.Results
Four multilevel motifs, overrepresented and specific to each group of K+ and/or Na+ ion channel toxins have been identified, using GIBBS and MEME and based on a training dataset of 79 sequences judged as representative of K+ and Na+ toxins. Unexpectedly birtoxin-like peptides appeared to present a new structural motif distinct from those present in K+ and Na+ channels Toxins. This result, supported by previous experimental data, suggests that birtoxin-like peptides may exert their activity on different sites than those targeted by classic K+ or Na+ toxins. Searching, the nr database with these newly identified motifs using MAST, retrieved several sequences (116 with e-value < 1) from various scorpion species (test dataset). The filtering process left 30 new and highly likely ion channel effectors. Phylogenetic analysis was used to classify the newly found sequences. Alternatively, classification tree analysis, using CART algorithm adjusted with the training dataset, using the motifs and their 2D structure as explanatory variables, provided a model for prediction of the activity of the new sequences.Conclusion
The phylogenetic results were in perfect agreement with those obtained by the CART algorithm. Our results may be used as criteria for a new classification of scorpion toxins based on functional motifs. 相似文献15.
Eva Alonso Amparo Alfonso Mercedes R. Vieytes Luis M. Botana 《Archives of toxicology》2016,90(2):479-488
Although voltage-gated sodium channels (Nav) are the cellular target of paralytic shellfish poisoning (PSP) toxins and that patch clamp electrophysiology is the most effective way of studying direct interaction of molecules with these channels, nowadays, this technique is still reduced to more specific analysis due to the difficulties of transforming it in a reliable throughput system. Actual functional methods for PSP detection are based in binding assays using receptors but not functional Nav channels. Currently, the availability of automated patch clamp platforms and also of stably transfected cell lines with human Nav channels allow us to introduce this specific and selective method for fast screenings in marine toxin detection. Taking advantage of the accessibility to pure PSP standards, we calculated the toxicity equivalent factors (TEFs) for nine PSP analogs obtaining reliable TEFs in human targets to fulfill the deficiencies of the official analytic methods and to verify automated patch clamp technology as a fast and reliable screening method for marine toxins that interact with the sodium channel. The main observation of this work was the large variation of TEFs depending on the channel subtype selected, being remarkable the variation of potency in the 1.7 channel subtype and the suitability of Nav 1.6 and 1.2 channels for PSP screening. 相似文献
16.
Scorpion β-toxins represent a particular pharmacological group of voltage-gated sodium channel (VGSC) neurotoxins. They typically shift the voltage dependence of activation to more hyperpolarizing potentials and reduce the peak current amplitude by binding to receptor-site 4. Here, we report the purification and functional characterization of the first voltage-gated sodium channel toxins, CeII8 and CeII9, isolated from the scorpion Centruroides elegans (Thorell, 1876), which is responsible for deadly cases of intoxication in Mexico. The soluble venom was fractionated by gel filtration and ion-exchange chromatography, followed by reversed-phase HPLC. The toxins CeII8 and CeII9 were further purified and both their amino acid sequence and molecular weight were determined. Both toxins were electrophysiologically characterized on four mammalian VGSCs (rNav1.2, rNav1.4, hNav1.5 and rNav1.7) expressed heterologously in Xenopus laevis oocytes, using the two-electrode voltage-clamp technique. Although CeII8 has the highest sequence similarity with scorpion α-toxins, inhibiting the inactivation of VGSCs, 300 nM toxin had a clear β-toxin effect and was selective towards Nav1.7, involved in short-term and inflammatory pain. To the best of our knowledge, CeII8 is the first β-toxin active on Nav1.7. CeII9, a typical anti-mammalian β-toxin, selectively modulated Nav1.4 at a concentration of 700 nM and was, in contrast to CeII8, found to be lethal to mice. Interestingly, both toxins, despite their differences in amino acid sequence, only altered the biophysical properties of a fraction of the expressed sodium channels. Since these effects have also been reported for the β-toxin CssIV, the bioactive surfaces of the toxins have been compared to each other. 相似文献
17.
Comparison of sea anemone and scorpion toxins binding to Kv1 channels: an example of convergent evolution 总被引:1,自引:0,他引:1
Comparison of data from functional mapping carried out on scorpion and sea anemones toxins blocking currents through voltage-gated potassium channels revealed that, despite their different 3D structures, the binding cores of these toxins displayed some similarities. Further molecular modeling studies suggested that these similarities reflect the use by these toxins of a common binding mode to exert their blocking function. Therefore, scorpion and sea anemone toxins offer an example of mechanistic convergent evolution. 相似文献
18.
L Kang MQ Zheng M Morishima Y Wang T Kaku K Ono 《British journal of pharmacology》2009,157(3):404-414
Background and purpose:
Bepridil is an anti-arrhythmic agent with anti-electrical remodelling effects that target many cardiac ion channels, including the voltage-gated Na+ channel. However, long-term effects of bepridil on the Na+ channel remain unclear. We explored the long-term effect of bepridil on the Na+ channel in isolated neonatal rat cardiomyocytes and in a heterologous expression system of human Nav1.5 channel.Experimental approach:
Na+ currents were recorded by whole-cell voltage-clamp technique. Na+ channel message and protein were evaluated by real-time RT-PCR and Western blot analysis.Key results:
Treatment of cardiomyocytes with 10 µmol·L−1 bepridil for 24 h augmented Na+ channel current (INa) in a dose- and time-dependent manner. This long-term effect of bepridil was mimicked or masked by application of W-7, a calmodulin inhibitor, but not KN93 [2-[N-(2-hydroxyethyl)-N-(4-methoxy benzenesulphonyl)]-amino-N-(4-chlorocinnamyl)-N-methylbenzylamine], a Ca2+/calmodulin-dependent kinase inhibitor. During inhibition of protein synthesis by cycloheximide, the INa increase due to bepridil was larger than the increase without cycloheximide. Bepridil and W-7 significantly slowed the time course of Nav1.5 protein degradation in neonatal cardiomyocytes, although the mRNA levels of Nav1.5 were not modified. Bepridil and W-7 did not increase INa any further in the presence of the proteasome inhibitor MG132 [N-[(phenylmethoxy)carbonyl]-L-leucyl-N-[(1S)-1-formyl-3-methylbutyl]-L-leucinamide]. Bepridil, W-7 and MG132 but not KN93 significantly decreased 20S proteasome activity in a concentration-dependent manner.Conclusions and implications:
We conclude that long-term exposure of cardiomyocytes to bepridil at therapeutic concentrations inhibits calmodulin action, which decreased degradation of the Nav1.5 α-subunit, which in turn increased Na+ current. 相似文献19.
Site-3 toxins are small polypeptide venoms from scorpions, sea anemones, and spiders that bind with a high specificity to the extracellular surface of voltage-gated Na channels. After binding to a site near the S4 segment in domain IV the toxin causes disruption of the normal fast inactivation transition resulting in a marked prolongation of the action potentials of excitable tissues including those of cardiac and skeletal muscle and nerve. In this review we discuss the specific binding interactions between residues of the toxin and those of the Na channel, and the specific modification of Na channel kinetic behavior leading to a change in fast inactivation focusing on interactions deduced primarily from the study of sea anemone toxins and the cardiac Na channel (Na(V)1.5). We also illustrate the usefulness of site-3 toxins in the study of altered Na channel behavior by drug-modification. 相似文献
20.
V Schulze C Stoetzer AO O'Reilly E Eberhardt N Foadi J Ahrens F Wegner A Lampert J Roche A Leffler 《British journal of pharmacology》2014,171(2):427-437
Background and Purpose: Treatment with methadone is associated with severe cardiac arrhythmias, a side effect that seems to result from an inhibition of cardiac hERG K+ channels. However, several other opioids are inhibitors of voltage-gated Na+ channels. Considering the common assumption that an inhibition of the cardiac Na+ channel Nav1.5, is the primary mechanism for local anaesthetic (LA)-induced cardiotoxicity, we hypothesized that methadone has LA-like properties leading to a modulation of Nav1.5 channels.Experimental Approach: The whole-cell patch clamp technique was applied to investigate the effects of methadone on wild-type and mutant human Nav1.5 channels expressed in HEK293 cells. A homology model of human Nav1.5 channels was used to perform automated ligand-docking studies.Key Results: Methadone inhibited Nav1.5 channels in a state-dependent manner, that is, tonic block was stronger with inactivated channels than with resting channels and a use-dependent block at 10 Hz. Methadone induced a concentration-dependent shift of the voltage dependency of both fast and slow inactivation towards more hyperpolarized potentials, and impaired recovery from fast and slow inactivation. The LA-insensitive mutants N406K and F1760A exhibited reduced tonic and use-dependent block by methadone, and docking predictions positioned methadone in a cavity that was delimited by the residue F1760. Dextromethadone and levomethadone induced discrete stereo-selective effects on Nav1.5 channels.Conclusions and Implications: Methadone interacted with the LA-binding site to inhibit Nav1.5 channels. Our data suggest that these channels are a hitherto unrecognized molecular component contributing to cardiac arrhythmias induced by methadone. 相似文献