Biophysics and structure-function relationship of T-type Ca2+ channels

T-type channels are distinguished among voltage-gated Ca2+ channels by their low voltage thresholds for activation and inactivation, fast inactivation and small single channel conductance in isotonic Ba2+. Detailed biophysical and pharmacological characterization of native T-type channels indicated that these channels represent a heterogeneous family. Cloning of three family members (CaV3.1-3.3) confirmed these observations and allowed the study of the structure-function relationship of these channels. T-type channels are likely heterotetrameric structures consisting of a single polypeptide of four homologous domains (I-IV), each one containing six transmembrane spans (S1-S6), and cytoplasmic N- and C-termini. Structure-function studies have revealed that fast macroscopic inactivation of CaV3.1 is modulated by specific residues in the proximal C-terminus and in the transmembrane domain IIIS6. The particular gating properties within the T-type channel subfamily are determined by several parts of the protein, whereas differences with respect to high-voltage-activated Ca2+ channels are mostly determined by domains I, II and III. Several gating properties are affected by alternative splicing, C-terminal truncations and mutations associated to idiopathic epilepsy. Intriguingly, the aspartate residues of the EEDD locus of the selectivity filter not only determine the permeation properties and the block by Cd2+ and protons, but also activation and deactivation. Mutagenesis has also revealed that the outermost arginines of the S4 segment of domain IV influence the activation of CaV3.2, though no specific voltage-sensing amino acid has yet been properly identified. The selective modulation of CaV3.2 by G-proteins, CaMKII and PKA is determined by the II-III linker and the high-affinity inhibition of CaV3.2 by Ni2+ relies on a histidine residue in the IS3-S4 linker. Certainly, more structure-function studies are needed for a better understanding of T-type channel physiology and the rational design of treatments against T-type channel-related pathologies.     

Hemicalcin, a new toxin from the Iranian scorpion Hemiscorpius lepturus which is active on ryanodine-sensitive Ca2+ channels

In the present work, we purified and characterized a novel toxin named hemicalcin from the venom of the Iranian chactoid scorpion Hemiscorpius lepturus where it represents 0.6% of the total protein content. It is a 33-mer basic peptide reticulated by three disulfide bridges, and that shares between 85 and 91% sequence identity with four other toxins, all known or supposed to be active on ryanodine-sensitive calcium channels. Hemicalcin differs from these other toxins by seven amino acids at positions 9 (leucine/arginine), 12 (alanine/glutamic acid), 13 (aspartic acid/asparagine), 14 (lysine/asparagine), 18 (serine/glycine), 26 (threonine/alanine) and 28 (proline/isoleucine/alanine). In spite of these differences, hemicalcin remains active on ryanodine-sensitive Ca2+ channels, since it increases [3H]ryanodine binding on RyR1 (ryanodine receptor type 1) and triggers Ca2+ release from sarcoplasmic vesicles. Bilayer lipid membrane experiments, in which the RyR1 channel is reconstituted and its gating properties are analysed, indicate that hemicalcin promotes an increase in the opening probability at intermediate concentration and induces a long-lasting subconductance level of 38% of the original amplitude at higher concentrations. Mice intracerebroventricular inoculation of 300 ng of hemicalcin induces neurotoxic symptoms in vivo, followed by death. Overall, these data identify a new biologically active toxin that belongs to a family of peptides active on the ryanodine-sensitive channel.     

Solution structure of BmP08, a novel short-chain scorpion toxin from Buthus martensi Karsch

A novel short-chain scorpion toxin BmP08 was purified from the venom of the Chinese scorpion Buthus martensi Karsch by a combination of gel-filtration, ion exchange, and reversed-phase chromatography. The primary sequence of BmP08 was determined using the tandem MS/MS technique and Edman degradation, as well as results of NMR sequential assignments. It is composed of 31 amino acid residues including six cysteine residues and shares less than 25% sequence identity with the known alpha-KTx toxins. BmP08 shows no inhibitory activity on all tested voltage-dependent and Ca(2+)-activated potassium channels. The 3D-structure of BmP08 has been determined by 2D-NMR spectroscopy and molecular modeling techniques. This toxin adopts a common alpha/beta-motif, but shows a distinctive local conformation and features a 3(10)-helix and a shorter beta-sheet. The unique structure is closely related to the distinct primary sequence of the toxin, especially to the novel arrangement of S-S linkages in the molecule, in which two disulfide bridges (C(i)-C(j) and C(i+3)-C(j+3)) link covalently the 3(10)-helix with one strand of the beta-sheet structure. The electrostatic potential surface analysis of the toxin reveals salt bridges and hydrogen bonds between the basic residues and negatively charged residues nearby in BmP08, which may be unfavorable for its binding with the known voltage-dependent and Ca(2+)-activated potassium channels. Thus, finding the target for this toxin should be an interesting task in the future.     

BmP09, a "long chain" scorpion peptide blocker of BK channels

A novel "long chain" toxin BmP09 has been purified and characterized from the venom of the Chinese scorpion Buthus martensi Karsch. The toxin BmP09 is composed of 66 amino acid residues, including eight cysteines, with a mass of 7721.0 Da. Compared with the B. martensi Karsch AS-1 as a Na(+) channel blocker (7704.8 Da), the BmP09 has an exclusive difference in sequence by an oxidative modification at the C terminus. The sulfoxide Met-66 at the C terminus brought the peptide a dramatic switch from a Na(+) channel blocker toaK(+) channel blocker. Upon probing the targets of the toxin BmP09 on the isolated mouse adrenal medulla chromaffin cells, where a variety of ion channels coexists, we found that the toxin BmP09 specifically blocked large conductance Ca(2+)- and voltage-dependent K(+) channels (BK) but not Na(+) channels at a range of 100 nm concentration. This was further confirmed by blocking directly the BK channels encoded with mSlo1 alpha-subunits in Xenopus oocytes. The half-maximum concentration EC(50) of BmP09 was 27 nm, and the Hill coefficient was 1.8. In outside-out patches, the 100 nm BmP09 reduced approximately 70% currents of BK channels without affecting the single-channel conductance. In comparison with the "short chain" scorpion peptide toxins such as Charybdotoxin, the toxin BmP09 behaves much better in specificity and reversibility, and thus it will be a more efficient tool for studying BK channels. A three-dimensional simulation between a BmP09 toxin and an mSlo channel shows that the Lys-41 in BmP09 lies at the center of the interface and plugs into the entrance of the channel pore. The stable binding between the toxin BmP09 and the BK channel is favored by aromatic pi -pi interactions around the center.     

Interaction of SK(Ca) channels and L-type Ca(2+) channels in catecholamine secretion in the rat adrenal gland

We elucidated the interaction of small-conductance Ca(2+)-activated K(+) (SK(Ca)) channels and L-type Ca(2+) channels in muscarinic receptor-mediated control of catecholamine secretion in the isolated perfused rat adrenal gland. The muscarinic agonist methacholine (10-300 microM) produced concentration-dependent increases in adrenal output of epinephrine and norepinephrine. The SK(Ca) channel blocker apamin (1 microM) enhanced the methacholine-induced catecholamine responses. The facilitatory effect of apamin on the methacholine-induced catecholamine responses was not observed during treatment with the L-type Ca(2+) channel blocker nifedipine (3 microM) or Ca(2+)-free solution. Nifedipine did not affect the methacholine-induced catecholamine responses, but it inhibited the responses during treatment with apamin. The L-type Ca(2+) channel activator Bay k 8644 (1 microM) enhanced the methacholine-induced catecholamine responses, whereas the enhancement of the methacholine-induced epinephrine and norepinephrine responses were prevented and attenuated by apamin, respectively. These results suggest that SK(Ca) channels are activated by muscarinic receptor stimulation, which inhibits the opening of L-type Ca(2+) channels and thereby attenuates adrenal catecholamine secretion.     

Characterisation of a novel toxin active on potassium apanaine channels, purified from Buthus occitanus tunetanus scorpion venom

A new peptidyl inhibitor of the small-conductance Ca(2+)-activated K+ channels (SKca) was purified to homogeneity from the venom of the Tunisian scorpion Buthus occitanus tunetanus. The molecular mass determined by SDS-PAGE, shows that it's a short peptide (3300 Da). The primary sequence of this toxin shows that it is a 31-residue polypeptide cross-linked by three disulfide bridges and structurally related to subfamily 5 of short scorpion toxins. This molecule shows similar pharmacological properties with this group of peptides inducing high toxicity in mice after intracerebro-ventricular injection, and competing with iodinated apamin for binding to its receptor site from rat brain synaptosomes (K0.5 = 4 nM).     

Chemical synthesis and characterization of Pi1, a scorpion toxin from Pandinus imperator active on K+ channels.

Pi1 is a 35-residue toxin cross-linked by four disulfide bridges that has been isolated from the venom of the chactidae scorpion Pandinus imperator. Due to its very low abundance in the venom, we have chemically synthesized this toxin in order to study its biological activity. Enzyme-based proteolytic cleavage of the synthetic Pi1 (sPi1) demonstrates half-cystine pairings between Cys4-Cys25, Cys10-Cys30, Cys14-Cys32 and Cys20-Cys35, which is in agreement with the disulfide bridge organization initially reported on the natural toxin. In vivo, intracerebroventricular injection of sPi1 in mice produces lethal effects with an LD50 of 0.2 microgram per mouse. In vitro, the application of sPi1 induces drastic inhibition of Shaker B (IC50 of 23 nM) and rat Kv1.2 channels (IC50 of 0.44 nM) heterologously expressed in Xenopus laevis oocytes. No effect was observed on rat Kv1.1 and Kv1.3 currents upon synthetic peptide application. Also, sPi1 is able to compete with 125I-labeled apamin for binding onto rat brain synaptosomes with an IC50 of 55 pM. Overall, these results demonstrate that sPi1 displays a large spectrum of activities by blocking both SK- and Kv1-types of K+ channels; a selectivity reminiscent of that of maurotoxin, another structurally related four disulfide-bridged scorpion toxin that exhibits a different half-cystine pairing pattern.     

Brownian dynamics simulations of the recognition of the scorpion toxin P05 with the small-conductance calcium-activated potassium channels

The recognition of the scorpion toxin P05 and the small-conductance, calcium-activated potassium (SK) channels, rsk1, rsk2, and rsk3, has been studied by means of the Brownian dynamics (BD) method. All of the 25 available structures of P05 in the RCSB Protein Data Bank determined by NMR were considered during the simulation, which indicated that the conformation of P05 affects both the recognition and binding between the two proteins significantly. Comparing the top four high-frequency structures of P05 binding to the SK channels, we found that the rsk2 channel, with high frequencies and lowest electrostatic interaction energies (E (int)(ES)), is the most favorable for P05 binding, while rsk3 is intermediate, and rsk1 is the least favorable. Among the 25 structures of P05, the 13th structure docks into the binding site of the rsk2 channel with the highest probability and most favorable electrostatic interactions. From the P05-rsk2 channel binding model, we identified the residues critical for the recognition of these two proteins through triplet contact analyses. P05 locates around the extracellular mouth of the SK channels and contacts the SK channels using its alpha-helix rather than beta-sheets. The critical triplet contacts for recognition between P05 and the rsk2 channel are Arg6 (P05)-Asp364 (SK), Arg7 (P05)-Asn368 (SK), and Arg13 (P05)-Asp341 (SK). The structure of the P05-rsk2 complex with the most favorable electrostatic interaction energy was further refined by molecular mechanics, showing that six hydrogen bonding interactions exist between P05 and the rsk2 channel: one hydrogen bond is formed between Arg6 (P05) and Asp364(D) (rsk2); Arg7 (P05) forms three hydrogen bonds with Asp341(B) (rsk2)) and Asp364(C) (rsk2); two hydrogen bonds are formed by Arg13 (P05) with Asp341(A) (rsk2) and Asp364(B) (rsk2). The simulation results are in good agreement with the previous molecular biological experiments and can explain the binding phenomena between P05 and SK channels at the level of molecular structure. The consistency between the results of the BD simulations and the experimental data indicated that our 3D model of the P05-rsk2 channel complex is reasonable and can be employed in further biological studies, such as rational design of the novel therapeutic agents blocking the small-conductance, calcium-activated and apamin-sensitive potassium channels, and for mutagenesis studies in both toxins and SK channels. In particular, both the BD simulations and the molecular mechanics refinements indicate that residue Asp364 of the rsk2 channel is critical for its recognition and binding functionality towards P05. This phenomenon has not been appreciated in the previous mutagenesis experiments, indicating that this might be a new clue for further functional study of SK channels.     

Kbot1, a three disulfide bridges toxin from Buthus occitanus tunetanus venom highly active on both SK and Kv channels

On attempts to identify toxins showing original profile of activity among K+ channels, we purified Kbot1, a scorpion toxin that blocks Kv1 and SK potassium channels. With 28 amino-acid residues, Kbot1 is the shortest toxin sequenced in Buthus occitanus scorpion. It is linked by three disulfide bridges and its primary structure is 93% identical to that of BmP02 isolated from the venom of the Chinese scorpion Buthus martensi Karsch [Eur. J. Biochem. 245 (1996) 457]. Kbot1 exhibited a low neurotoxicity in mice after intracerebroventricular injection (LD50 approximately or = 0.8 microg per mouse). It competes with iodinated apamin for its rat brain synaptosomal membrane-binding site (IC50 of 20 nM). Despite 30% sequence identity between Kbot1 and ChTX, competitive experiments on the [125I] charybdotoxin, show that Kbot1 inhibits its binding to its rat brain synaptosomes with IC50 of 10 nM. This result was supported by electrophysiological experiments on cloned voltage-dependent K+ channels from rat brain, expressed in Xenopus oocytes. Kbot1 blocks Kv1.1, Kv1.2 and Kv1.3 currents with IC50 of 145, 2.5 and 15 nM, respectively. Based on these data, Kbot1 may be considered as the first member of subfamily 9 of scorpion toxins [Trends Pharmacol. Sci. 20 (1999) 444], highly active on both Kv and SK channels.     

Decreased expression of small-conductance Ca2+-activated K+ channels SK1 and SK2 in human chronic atrial fibrillation

AimsSmall-conductance Ca^2 +-activated K⁺ (SK) channels are recognized as new ion channel candidates in atrial fibrillation (AF), with pivotal implications as novel drug targets due to their atrial-selective distribution in humans. The purpose of this study was to investigate whether SK channels and the Ca^2 +-activated K⁺ current (I_K,Ca) are involved in electrical remodeling of human chronic AF (cAF) and whether they display the differential distribution between the right (RA) and left atria (LA).Main methodsThe right (RAA) and left atrial appendage (LAA) myocytes were obtained from 29 sinus rhythm (SR) and 22 cAF patients. The I_K,Ca and action potential (AP) were recorded using the patch-clamp technique. Three SK channel subtypes (SK1–3) expressions were assayed by western blot and real-time quantitative PCR analysis.Key findingsThe I_K,Ca was decreased and its role in AP repolarization was attenuated in cAF, concomitant with a significant decrease in protein and mRNA levels of SK1 and SK2. In either SR or cAF, there was no difference in the I_K,Ca density and protein and mRNA expression levels of SK1–3 between RAA and LAA myocytes.SignificanceOur results demonstrated that SK1 and SK2 are involved in electrical remodeling of cAF. SK1–3 and I_K,Ca do not display the inter-atrial differential distribution in SR or cAF. These findings provide a new insight into mechanisms of electrical remodeling of human cAF.     

OD1, the first toxin isolated from the venom of the scorpion Odonthobuthus doriae active on voltage-gated Na+ channels

In this study, we isolated and pharmacologically characterized the first alpha-like toxin from the venom of the scarcely studied Iranian scorpion Odonthobuthus doriae. The toxin was termed OD1 and its primary sequence was determined: GVRDAYIADDKNCVYTCASNGYCNTECTKNGAESGYCQWIGRYGNACWCIKLPDEVPIRIPGKCR. Using the two-electrode voltage clamp technique, the pharmacological effects of OD1 were studied on three cloned voltage-gated Na+ channels expressed in Xenopus laevis oocytes (Na(v)1.2/beta1, Na(v)1.5/beta1, para/tipE). The inactivation process of the insect channel, para/tipE, was severely hampered by 200 nM of OD1 (EC50 = 80+/-14 nM) while Na(v)1.2/beta1 still was not affected at concentrations up to 5 microM. Na(v)1.5/beta1 was influenced at micromolar concentrations.     

Chemical synthesis and structure-activity relationships of Ts kappa, a novel scorpion toxin acting on apamin-sensitive SK channel.

Tityus kappa (Ts kappa), a novel toxin from the venom of the scorpion Tityus serrulatus, is a 35-residue polypeptide cross-linked by three disulphide bridges and acts on small-conductance calcium-activated potassium channels (SK channels). Ts K was chemically synthesized using the solid-phase method and characterized. The synthetic product, sTs kappa, was indistinguishable from the natural toxin when tested in vitro in competition assay with radiolabelled apamin for binding to rat brain synaptosomes (IC50 = 3 nM). The sTs kappa was further tested in vivo for lethal activity to mice following intracerebroventricular inoculation (LD50 = 70 ng per mouse). The half-cystine pairings were formerly established by enzyme-based cleavage of sTs kappa; they were between Cys7-Cys28, Cys13-CyS33 and Cys17-Cys35, which is a disulphide bridge pattern similar to that of other short scorpion toxins. According to previous studies on SK channel-acting toxins, the putative influence of certain basic residues of Ts kappa (i.e. Arg6, Arg9, Lys18, Lys19) in its pharmacological activity was investigated using synthetic point-mutated analogues of the toxin with an Ala substitution at these positions. Data from binding assay, together with conformational analysis of the synthetic analogues by 1H-NMR, suggest that Arg6, and to a lesser extent Arg9, are important residues for an high-affinity interaction of this toxin with SK channels; interestingly these residues are located outside the alpha-helical structure, whereas the pharmacologically important basic residues from other SK channel-specific toxins had been located inside the alpha-helix.     

Evidence for domain-specific recognition of SK and Kv channels by MTX and HsTx1 scorpion toxins

Maurotoxin (MTX) and HsTx1 are two scorpion toxins belonging to the alpha-KTx6 structural family. These 34-residue toxins, cross-linked by four disulfide bridges, share 59% sequence identity and fold along the classical alpha/beta scaffold. Despite these structural similarities, they fully differ in their pharmacological profiles. MTX is highly active on small (SK) and intermediate (IK) conductance Ca(2+)-activated (K(+)) channels and on voltage-gated Kv1.2 channel, whereas HsTx1 potently blocks voltage-gated Kv1.1 and Kv1.3 channels only. Here, we designed and chemically produced MTX-HsTx1, a chimera of both toxins that contains the N-terminal helical region of MTX (sequence 1-16) and the C-terminal beta-sheet region of HsTx1 (sequence 17-34). The three-dimensional structure of the peptide in solution was solved by (1)H NMR. MTX-HsTx1 displays the activity of MTX on SK channel, whereas it exhibits the pharmacological profile of HsTx1 on Kv1.1, Kv1.2, Kv1.3, and IK channels. These data demonstrate that the helical region of MTX exerts a key role in SK channel recognition, whereas the beta-sheet region of HsTx1 is crucial for activity on all other channel types tested.     

The SK3 subunit of small conductance Ca2+-activated K+ channels interacts with both SK1 and SK2 subunits in a heterologous expression system

The aim of this study was to determine whether functional heteromeric channels can be formed by co-assembly of rat SK3 (rSK3) potassium channel subunits with either SK1 or SK2 subunits. First, to determine whether rSK3 could co-assemble with rSK2 we created rSK3VK (an SK3 mutant insensitive to block by UCL 1848). When rSK3VK was co-expressed with rSK2 the resulting currents had an intermediate sensitivity to UCL 1848 (IC50 of approximately 5 nM compared with 120 pM for rSK2 and >300 nM for rSK3VK), suggesting that rSK3 and rSK2 can form functional heteromeric channels. To detect co-assembly of SK3 with SK1, we initially used a dominant negative construct of the human SK1 subunit (hSK1YP). hSK1YP dramatically reduced the SK3 current, supporting the idea that SK3 and SK1 subunits also interact. To determine whether these assemblies were functional we created rSK3VF, an rSK3 mutant with an enhanced affinity for tetraethylammonium chloride (TEA) (IC50 of 0.3 mM). Co-transfection of rSK3VF and hSK1 produced currents with a sensitivity to TEA not different from that of hSK1 alone (IC50 approximately 15 mM). These results suggest that hSK1 does not produce functional cell-surface assemblies with SK3. Antibody-staining experiments suggested that hSK1 may reduce the number of functional SK3 subunits reaching the cell surface. Additional experiments showed that co-expression of the rat SK1 gene with SK3 also dramatically suppressed SK current. The pharmacology of the residual current was consistent with that of homomeric SK3 assemblies. These results demonstrate interactions that cause changes in protein trafficking, cell surface expression, and channel pharmacology and strongly suggest heteromeric assembly of SK3 with the other SK channel subunits.     

Solution structure of Pi4, a short four-disulfide-bridged scorpion toxin specific of potassium channels

Pi4 is a short toxin found at very low abundance in the venom of Pandinus imperator scorpions. It is a potent blocker of K(+) channels. Like the other members of the alpha-KTX6 subfamily to which it belongs, it is cross-linked by four disulfide bonds. The synthetic analog (sPi4) and the natural toxin (nPi4) have been obtained by solid-phase synthesis or from scorpion venom, respectively. Analysis of two-dimensional (1)H NMR spectra of nPi4 and sPi4 indicates that both peptides have the same structure. Moreover, electrophysiological recordings of the blocking of Shaker B K(+) channels by sPi4 (K(D) = 8.5 nM) indicate that sPi4 has the same blocking activity of nPi4 (K(D) = 8.0 nM), previously described. The disulfide bonds have been independently determined by NMR and structure calculations, and by Edman-degradation/mass-spectrometry identification of peptides obtained by proteolysis of nPi4. Both approaches indicate that the pairing of the half-cystines is (6)C-(27)C, (12)C-(32)C, (16)C-(34)C, and (22)C-(37)C. The structure of the toxin has been determined by using 705 constraints derived from NMR data on sPi4. The structure, which is well defined, shows the characteristic alpha/beta scaffold of scorpion toxins. It is compared to the structure of the other alpha-KTX6 subfamily members and, in particular, to the structure of maurotoxin, which shows a different pattern of disulfide bridges despite its high degree of sequence identity (76%) with Pi4. The structure of Pi4 and the high amounts of synthetic peptide available, will enable the detailed analysis of the interaction of Pi4 with K(+) channels.     

Physiology, pharmacology and modelling of potassium channels: focus on SK channels

Various types of ion channels are involved in the control of neuronal activity. Among them, SK channels represent an interesting therapeutic target. Indeed, they underlie medium duration after hyperpolarizations in many types of neurons, thus inhibiting cell excitability. A thorough knowledge of the physiology of these channels and the discovery of non-peptidic selective modulators able to cross the blood-brain barrier are essential in view of developing future drugs for brain diseases such as those related to a dysfunction of dopaminergic and serotonergic systems.     

Ca(2+)-activated K+ channels: molecular determinants and function of the SK family

Ca(2+)-activated K(+) (K(Ca)) channels of small (SK) and intermediate (IK) conductance are present in a wide range of excitable and non-excitable cells. On activation by low concentrations of Ca(2+), they open, which results in hyperpolarization of the membrane potential and changes in cellular excitability. K(Ca)-channel activation also counteracts further increases in intracellular Ca(2+), thereby regulating the concentration of this ubiquitous intracellular messenger in space and time. K(Ca) channels have various functions, including the regulation of neuronal firing properties, blood flow and cell proliferation. The cloning of SK and IK channels has prompted investigations into their gating, pharmacology and organization into calcium-signalling domains, and has provided a framework that can be used to correlate molecularly identified K(Ca) channels with their native currents.     

Interaction of the scorpion toxin discrepin with Kv4.3 channels and A-type K channels in cerebellum granular cells

Background

The peptide discrepin from the α-KTx15 subfamily of scorpion toxins preferentially affects transient A-type potassium currents, which regulate many aspects of neuronal function in the central nervous system. However, the specific Kv channel targeted by discrepin and the molecular mechanism of interaction are still unknown.

Methods

Different variant peptides of discrepin were chemically synthesized and their effects were studied using patch clamp technique on rat cerebellum granular cells (CGC) and HEK cells transiently expressing Kv4.3 channels.

Results

Functional analysis indicated that nanomolar concentrations of native discrepin blocked Kv4.3 expressed channels, as previously observed in CGC. Similarly, the apparent affinities of all mutated peptides for Kv4.3 expressed channels were analogous to those found in CGC. In particular, in the double variant [V6K, D20K] the apparent affinity increased about 10-fold, whereas in variants carrying a deletion (ΔK13) or substitution (K13A) at position K13, the blockage was removed and the apparent affinity decreased more than 20-fold.

Conclusion

These results indicate that Kv4.3 is likely the target of discrepin and highlight the importance of the basic residue K13, located in the α-helix of the toxin, for current blockage.

General significance

We report the first example of a Kv4 subfamily potassium channel blocked by discrepin and identify the amino acid residues responsible for the blockage. The availability of discrepin variant peptides stimulates further research on the functions and pharmacology of neuronal Kv4 channels and on their possible roles in neurodegenerative disorders.