Purification, characterization, and biosynthesis of margatoxin, a component of Centruroides margaritatus venom that selectively inhibits voltage-dependent potassium channels.

García‐Calvo, Margarita; Leonard, Reid J.; Novick, J; Stevens, Scott P.; Schmalhofer, William A.; Kaczorowski, Gregory J.; García, María L.

doi:10.1016/s0021-9258(17)46707-x

Cited by 311 publications

(64 citation statements)

References 74 publications

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“…The homotetrameric arrangement of voltage-gated potassium channels was first revealed by the blocking analyses of charybdotoxin (ChTX, from Leiurus quinquestriatus) to wild-type and mutant channels (MacKinnon, 1991). Many potent polypeptide inhibitors of the lymphocyte potassium channels have been discovered in scorpion venom (Alessandri-Haber et al, 1999;Blanc et al, 1998;Garcia-Calvo et al, 1993;Gilquin et al, 2002;Goldstein et al, 1994). ChTX was the first polypeptide shown to block all Kv channels, inhibiting Kv1.3 with nanomolar affinity.…”

Section: Introductionmentioning

confidence: 99%

“…ChTX was the first polypeptide shown to block all Kv channels, inhibiting Kv1.3 with nanomolar affinity. Other scorpion toxins include noxiustoxin, kaliotoxin, margatoxin, agitoxin-2,hongotoxin, HsTx1, maurotoxin, and Pandinus toxins 1-3 (Pi1, Pi2, and Pi3) (Bontems et al, 1992;Fernandez et al, 1994;Garcia-Calvo et al, 1993;Goldstein et al, 1994;Krezel et al, 1995). In general, channel inhibitors can be pore blockers or gating modifiers (Goldstein et al, 1994;Harvey et al, 1995;Miller, 1995).…”

Section: Introductionmentioning

confidence: 99%

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Computational Simulations of Interactions of Scorpion Toxins with the Voltage-Gated Potassium Ion Channel

Liu

et al. 2004

Biophysical Journal

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Based on a homology model of the Kv1.3 potassium channel, the recognitions of the six scorpion toxins, viz. agitoxin2, charybdotoxin, kaliotoxin, margatoxin, noxiustoxin, and Pandinus toxin, to the human Kv1.3 potassium channel have been investigated by using an approach of the Brownian dynamics (BD) simulation integrating molecular dynamics (MD) simulation. Reasonable three-dimensional structures of the toxin-channel complexes have been obtained employing BD simulations and triplet contact analyses. All of the available structures of the six scorpion toxins in the Research Collaboratory for Structural Bioinformatics Protein Data Bank determined by NMR were considered during the simulation, which indicated that the conformations of the toxin significantly affect both the molecular recognition and binding energy between the two proteins. BD simulations predicted that all the six scorpion toxins in this study use their beta-sheets to bind to the extracellular entryway of the Kv1.3 channel, which is in line with the primary clues from the electrostatic interaction calculations and mutagenesis results. Additionally, the electrostatic interaction energies between the toxins and Kv1.3 channel correlate well with the binding affinities (-logK(d)s), R(2) = 0.603, suggesting that the electrostatic interaction is a dominant component for toxin-channel binding specificity. Most importantly, recognition residues and interaction contacts for the binding were identified. Lys-27 or Lys-28, residues Arg-24 or Arg-25 in the separate six toxins, and residues Tyr-400, Asp-402, His-404, Asp-386, and Gly-380 in each subunit of the Kv1.3 potassium channel, are the key residues for the toxin-channel recognitions. This is in agreement with the mutation results. MD simulations lasting 5 ns for the individual proteins and the toxin-channel complexes in a solvated lipid bilayer environment confirmed that the toxins are flexible and the channel is not flexible in the binding. The consistency between the results of the simulations and the experimental data indicated that our three-dimensional models of the toxin-channel complex are reasonable and can be used as a guide for future biological studies, such as the rational design of the blocking agents of the Kv1.3 channel and mutagenesis in both toxins and the Kv1.3 channel. Moreover, the simulation result demonstrates that the electrostatic interaction energies combined with the distribution frequencies from BD simulations might be used as criteria in ranking the binding configuration of a scorpion toxin to the Kv1.3 channel.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Computational Simulations of Interactions of Scorpion Toxins with the Voltage-Gated Potassium Ion Channel

Liu

et al. 2004

Biophysical Journal

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“…MgTx is reported to block Kv1.3 with an IC 50 of 11.7–119 pM (Garcia‐Calvo et al ., 1993; Knaus et al ., 1995), and therefore we measured the ability of HisMgTx and QDMgTx to block Kv1.3 in HEK293 cells (Figure 4). Addition of the epitope tag lowered the affinity of the peptide inhibitor, but HisMgTx still blocked Kv1.3 with picomolar affinity (IC 50 = 487 ± 106 pM, Hill coefficient = 1).…”

Section: Resultsmentioning

confidence: 99%

“…Recombinant expression of the Kv1.3 inhibitor (HisMgTx) permitted conjugation to the Zn 2+ surface of QDs using polyhistidine‐mediated self‐assembly, while retaining a majority of predicted biophysical properties of the channel in a blocked state. Addition of a polyhistidine tag only slightly decreased the inhibitory potency of the peptide (IC 50 of 487 pM compared to reported IC 50 values of 112–119 pM for unconjugated peptide (Garcia‐Calvo et al ., 1993; Knaus et al ., 1995)). Despite this small reduction in potency, block of the channel occurred quickly.…”

Section: Discussionmentioning

confidence: 99%

“…We selected the pore‐blocker margatoxin (MgTx) to inhibit Kv1.3 current flow and label cells expressing the channel because of its reported selectivity (Garcia et al ., 1997; Garcia‐Calvo et al ., 1993; Knaus et al ., 1995) and applications in the OB (Colley et al ., 2004; Fadool & Levitan, 1998; Mast & Fadool, 2012; Schwartz et al ., 2017). Luminescent quantum dots (QDs) were utilized because of their intense photo‐properties and ability to tune their interactions with biomolecules via modification of the QDs polymer ligand (Sundberg & Bruce Martin, 1974; Wang et al., 2016).…”

Section: Introductionmentioning

confidence: 99%

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Olfactory bulb‐targeted quantum dot (QD) bioconjugate and Kv1.3 blocking peptide improve metabolic health in obese male mice

Schwartz

Kapur

Huang

et al. 2020

Journal of Neurochemistry

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The olfactory system is a driver of feeding behavior, whereby olfactory acuity is modulated by the metabolic state of the individual. The excitability of the major output neurons of the olfactory bulb (OB) can be modulated through targeting a voltage‐dependent potassium channel, Kv1.3, which responds to changes in metabolic factors such as insulin, glucose, and glucagon‐like peptide‐1. Because gene‐targeted deletion or inhibition of Kv1.3 in the periphery has been found to increase energy metabolism and decrease body weight, we hypothesized that inhibition of Kv1.3 selectively in the OB could enhance excitability of the output neurons to evoke changes in energy homeostasis. We thereby employed metal‐histidine coordination to self‐assemble the Kv1.3 inhibitor margatoxin (MgTx) to fluorescent quantum dots (QDMgTx) as a means to label cells in vivo and test changes in neuronal excitability and metabolism when delivered to the OB. Using patch‐clamp electrophysiology to measure Kv1.3 properties in heterologously expressed cells and native mitral cells in OB slices, we found that QDMgTx had a fast rate of inhibition, but with a reduced IC50, and increased action potential firing frequency. QDMgTx was capable of labeling cloned Kv1.3 channels but was not visible when delivered to native Kv1.3 in the OB. Diet‐induced obese mice were observed to reduce body weight and clear glucose more quickly following osmotic mini‐pump delivery of QDMgTx/MgTx to the OB, and following MgTx delivery, they increased the use of fats as fuels (reduced respiratory exchange ratio). These results suggest that enhanced excitability of bulbar output neurons can drive metabolic responses.

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Solution structure of maurotoxin, a scorpion toxin fromScorpio maurus, with high affinity for voltage-gated potassium channels

et al. 1997

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Maurotoxin (MTX), purified from the scorpionid Scorpio maurus is a potent ligand for potassium channels. It shows a broad specificity as being active on Kv1.1 (Kd = 37 nM), Kv1.2 (Kd = 0.8 nM), Kv1.3 (Kd = 150 nM) voltage-gated potassium channels, as well as on small-conductance calcium-activated potassium channels. It has a unique disulfide pairing among the scorpion toxins family. The solution structure of MTX has been determined by 2D-NMR techniques, which led to the full description of its 3D conformation: a bended helix from residues 6 to 16 connected by a loop to a two-stranded antiparallel beta sheet (residues 23 to 26 and 28 to 31). The interaction of MTX with the pore region of the Kv1.2 potassium channel has been modeled according to their charge anisotropy. The structure of MTX is similar to other short scorpion toxins despite its peculiar disulfide pairing. Its interaction with the Kv1.2 channel involves a dipole moment, which guides and orients the toxin onto the pore, toward the binding site, and which thus is responsible for the specificity.

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Purification, characterization, and biosynthesis of margatoxin, a component of Centruroides margaritatus venom that selectively inhibits voltage-dependent potassium channels.

Cited by 311 publications

References 74 publications

Computational Simulations of Interactions of Scorpion Toxins with the Voltage-Gated Potassium Ion Channel

Computational Simulations of Interactions of Scorpion Toxins with the Voltage-Gated Potassium Ion Channel

Olfactory bulb‐targeted quantum dot (QD) bioconjugate and Kv1.3 blocking peptide improve metabolic health in obese male mice

Solution structure of maurotoxin, a scorpion toxin fromScorpio maurus, with high affinity for voltage-gated potassium channels

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