Scorpion Leiurus Quinquestriatus Research Articles

Mortality and feeding of mid-stadium larvae of Helicoverpa zea and Heliothis virescens fed a wild strain or a recombinant strain of Baculovirus heliothis expressing an insect-specific toxin of the scorpion Leiurus quinquestriatus hebraeus

A recombinant of the Heliothis/Helicoverpa single-enveloped nucleopolyhedrosis virus (RcHzSNPV), carrying a gene encoding an insect-specific toxin from the scorpion, Leiurus quniquestriatus, herbraeus, significantly increased the rapidity of kill and reduced feeding by larvae af Heliothis virescens and Helicoverpa zea over that of its parental, wind-type strain (WtHzSNPV). The RcHzSNPV at 1.0 occlusion bodies/mm2 killed or paralyzed H. virescens and H. zea larvae ca. 1.5-fold and 1.7-fold quicer, respectively, than the WtHzSNPV strain. Larvae not exposed to virus produced 3-to 5-fold more frass than larvae exposed to RcHzSNPV, and ca. 2-to 3-fold more frass than larvae exposed to WtHzSNPV. Increasing the rapidity of kill and reducing larval feeding are attributes that enhance the use of recombinant baculoviruses as potential biorational pesticides.

Second degree atrio-ventricular block and cardiotoxicity secondary to envenomation by the scorpion Leiurus quinquestriatus (‘Yellow Scorpion’) – an indication for serotherapy?

Second degree atrio-ventricular block and cardiotoxicity secondary to envenomation by the scorpion Leiurus quinquestriatus (‘Yellow Scorpion’) – an indication for serotherapy?

δ-Atracotoxins from Australian Funnel-web Spiders Compete with Scorpion α-Toxin Binding but Differentially Modulate Alkaloid Toxin Activation of Voltage-gated Sodium Channels

delta-Atracotoxins from the venom of Australian funnel-web spiders are a unique group of peptide toxins that slow sodium current inactivation in a manner similar to scorpion alpha-toxins. To analyze their interaction with known sodium channel neurotoxin receptor sites, we studied their effect on [3H]batrachotoxin and 125I-Lqh II (where Lqh is alpha-toxin II from the venom of the scorpion Leiurus quinquestriatus hebraeus) binding and on alkaloid toxin-stimulated 22Na+ uptake in rat brain synaptosomes. delta-Atracotoxins significantly increased [3H]batrachotoxin binding yet decreased maximal batrachotoxin-activated 22Na+ uptake by 70-80%, the latter in marked contrast to the effect of scorpion alpha-toxins. Unlike the inhibition of batrachotoxin-activated 22Na+ uptake, delta-atracotoxins increased veratridine-stimulated 22Na+ uptake by converting veratridine from a partial to a full agonist, analogous to scorpion alpha-toxins. Hence, delta-atracotoxins are able to differentiate between the open state of the sodium channel stabilized by batrachotoxin and veratridine and suggest a distinct sub-conductance state stabilized by delta-atracotoxins. Despite these actions, low concentrations of delta-atracotoxins completely inhibited the binding of the scorpion alpha-toxin, 125I-Lqh II, indicating that they bind to similar, or partially overlapping, receptor sites. The apparent uncoupling between the increase in binding but inhibition of the effect of batrachotoxin induced by delta-atracotoxins suggests that the binding and action of certain alkaloid toxins may represent at least two distinguishable steps. These results further contribute to the understanding of the complex dynamic interactions between neurotoxin receptor site areas related to sodium channel gating.

New toxins acting on sodium channels from the scorpion Leiurus quinquestriatus hebraeus suggest a clue to mammalian vs insect selectivity

Two new toxins were purified from Leiurus quinquestriatus hebraeus (Lqh) scorpion venom, Lqh II and Lqh III. Lqh II sequence reveals only two substitutions, as compared to AaH II, the most active scorpion α-toxin on mammals from Androctounus australis Hector. Lqh III shares 80% sequence identity with the α-like toxin Bom III, from Buthus occitanus mardochei. Using bioassays on mice and cockroach coupled with competitive binding studies with 125I-labeled scorpion α-toxins on rat brain and cockroach synaptosomes, the animal selectivity was examined. Lqh II has comparable activity to mammals as AaH II, but reveals significantly higher activity to insects attributed to its C-terminal substitution, and competes at low concentration for binding on both mammalian and cockroach sodium channels. Lqh II thus binds to receptor site 3 on sodium channels. Lqh III is active on both insects and mammals but competes for binding only on cockroach. The latter indicates that Lqh III binds to a distinct receptor site. Thus, Lqh II and Lqh III represent two different scorpion toxin groups, the α- and α-like toxins, respectively, according to the structural and pharmacological criteria. These new toxins may serve as a lead for clarification of the structural basis for insect vs mammal selectivity of scorpion toxins.

Effects of antivenom serotherapy on hemodynamic pathophysiology in dogs injected with L. quinquestriatus scorpion venom

In dogs, scorpion venom causes an immediate increase in cardiac output that declines below baseline values within one hour. We tested the hypotheses that antivenom given before venom injection may prevent changes in cardiac output, while antivenom given after the inotropic stage of envenomation cannot reverse cardiac output decline. Twenty-five anesthetized, mechanically ventilated dogs were given 0.1 mg/kg IV venom of the scorpion Leiurus quinquestriatus. The dogs were randomized into 4 groups: 5 dogs were given venom alone (control group) and 6 dogs were given 6 ml of antivenom one minute before venom injection while 8 and 6 dogs were given 6 ml of antivenom 20 and 60 min after venom injection, respectively. Parameters reflecting respiratory and circulatory functions were measured for 180 min after venom injection. Scorpion venom caused a gradual decrease in heart rate, an initial elevation of systemic and pulmonary blood pressure and cardiac output followed by a decline in these parameters. PO 2, pH and HCO 3 − gradually decreased, while PCO 2 gradually increased from baseline. Antivenom given before venom injection prevented all the effects induced by the venom. Antivenom given at 20 and 60 min after venom injection had no effect on cardiac output and HCO 3 − decline, but caused an increase in heart rate, PO 2 and pH and a decrease in PCO 2. We assume that antivenom clears free toxins from the circulation, and since cardiac output and HCO 3 − did not improve after this clearance, we conclude that following intravenous venom injection, heart and circulation are rapidly affected by the toxins or by other substances released by the venom which do not respond to antivenom. Improvements in respiration and heart rate with antivenom given after venom injection may be secondary to reversion of cholinergic effects of the venom. Improvement in respiration may be also explained by reversion of the toxic effects on Ca 2+ activated K + channels of bronchial smooth muscle. All these effects may be secondary to clearance of toxins by the antivenom.

A depressant insect-selective toxin analog from the venom of the scorpion Leiurus quinquestriatus hebraeus--purification and structure/function characterization.

The scorpion venom-derived excitatory and depressant insect-selective polypeptide neurotoxins modify sodium conductance in insect neuronal membranes and differ greatly in their primary structures and symptoms induced in blow fly larvae. We report here the purification and characterization of a new insect selective toxin, LqhIT5. LqhIT5 is more similar to the excitatory toxins in its mode of action and the depressant toxins in its primary structure. This toxin is a single polypeptide composed of 61 amino acids that are cross linked by four disulfide bonds. When LqhIT5 is injected into blow fly larvae, a fast contraction paralysis occurs without depressant activity. No mammalian toxicity was detected by subcutaneous or intracranial injections of this toxin into mice. Sequence comparison of LqhIT5 and known depressant toxins shows a high degree of similarity among the amino acids located on the C-terminus of the toxins. However, there are some clear differences in the amino acids located close to the N-terminus of the toxins. By the aid of homology modeling, we demonstrated that these amino acids have the same orientation in the tertiary structure of the molecule and are exposed to the environment. The change in the mode of action of LqhIT5 (no depressant activity) by substitutions of a few amino acids located on a specific exposed area of the toxin shed a new light on the structure/function relationship of scorpion toxins. These results caution that similarity in the mechanism of action of scorpion toxins does not always follow from an overall similarity in sequence.

The involvement of plasma kinins in the cardiovascular effects of Leiurus quinquestriatus scorpion venom in anaesthetised rabbits

The present study was undertaken to investigate the involvement of kinins in the cardiovascular and respiratory effects of LQQ venom. Blood pressure, heart rate, electrocardiogram (ECG) and respiration were studied in anaesthetised rabbits, in the presence and absence of aprotinin and icatibant, a B 2 bradykinin antagonist. Plasma bradykinin concentrations were also measured following venom injection. LQQ venom caused a triphasic effect on blood pressure comprising an immediate fall, a pronounced rise and a progressive decline until death. Bradycardia, myocardial damage, arrhythmias, respiratory distress and pulmonary oedema were also exhibited. Pretreatment with aprotinin attenuated the venom-induced hypotension, bradycardia, ECG and respiratory changes and prolonged survival. Pretreatment of atropinized animals with icatibant gave similar protection. In animals treated with LQQ venom, plasma bradykinin was significantly higher than controls, although there was considerable inter-animal variation in plasma kinin concentrations and the elevation was seen relatively late after venom administration. The data provides some support for the hypothesis that kinins are involved in the cardiovascular and lethal effects of LQQ venom in rabbits.

Differential involvement of disulfide bridges on the folding of a scorpion toxin.

Leiurotoxin I is a neurotoxin, blocker of Ca(2+)-activated apamin-sensitive K+ channel, purified from the venom of the scorpion Leiurus quinquestriatus hebraeus. It is a 31-residue polypeptide reticulated by three disulfide bridges, i.e. Cys3-Cys21, Cys8-Cys26 and Cys12-Cys28. To investigate the role of these disulfide bridges in the folding of this toxin, analogs lacking one disulfide bridge were synthesized. The structures of two analogs in which two half-cystines were placed by alpha-aminobutyrate residues to suppress one disulfide bridge, were analyzed by 1H NMR. The NMR studies reveal a three-dimensional structure identical with the native toxin for the analog lacking disulfide bridge Cys3-Cys21 and a loss of organized structure for another analog lacking disulfide bridge Cys12-Cys28. These analogs are, respectively, fully active and weakly active (2% of the residual activity) when tested in vitro for their ability to interact with their receptor channel and in vivo for their neurotoxic activity in mice. This suggest that disulfide bridge Cys12-Cys28 is essential for the folding process. In contrast, the lack of disulfide bridge Cys3-Cys21 does not affect the folding and the maintenance of bioactive conformation of Leiurotoxin I.

Identification of Structural Elements of a Scorpion α-Neurotoxin Important for Receptor Site Recognition

alpha-Neurotoxins from scorpion venoms constitute the most studied group of modifiers of the voltage-sensitive sodium channels, and yet, their toxic site has not been characterized. We used an efficient bacterial expression system for modifying specific amino acid residues of the highly insecticidal alpha-neurotoxin LqhalphaIT from the scorpion Leiurus quinquestriatus hebraeus. Toxin variants modified at tight turns, the C-terminal region, and other structurally related regions were subjected to neuropharmacological and structural analyses. This approach highlighted both aromatic (Tyr10 and Phe17) and positively charged (Lys8, Arg18, Lys62, and Arg64) residues that (i) may interact directly with putative recognition points at the receptor site on the sodium channel; (ii) are important for the spatial arrangement of the toxin polypeptide; and (iii) contribute to the formation of an electrostatic potential that may be involved in biorecognition of the receptor site. The latter was supported by a suppressor mutation (E15A) that restored a detrimental effect caused by a K8D substitution. The feasibility of producing anti-insect scorpion neurotoxins with augmented toxicity was demonstrated by the substitution of the C-terminal arginine with histidine. Altogether, the present study provides for the first time an insight into the putative toxic surface of a scorpion neurotoxin affecting sodium channel gating.

Characterization of a new family of toxin-like peptides from the venom of the scorpion Leiurus quinquestriatus hebraeus. 1H-NMR structure of leiuropeptide II.

To extend our knowledge about the structural features of short scorpion toxins, the ion-exchange fractions obtained from Leiurus quinquestriatus hebraeus venom were investigated by plasma desorption mass spectrometry in order to select low molecular mass polypeptides. Three toxin-like peptides with molecular mass close to 3 kDa, named leiuropeptides I, II and III, were purified and found devoid of any significant toxicity against mammals and insects. Their amino acid sequences revealed a cysteine pattern analogous to that of short-chain scorpion toxins. The solution structure of leiuropeptide II was determined by 2D 1H-NMR spectroscopy and indicated the presence of a helix accommodating a proline, connected to a two-standard beta-sheet by three disulfide bonds. The overall fold of leiuropeptide II is found to be similar to that of leiurotoxin I, a 31-residue toxin present in the same scorpion venom which acts on K+ channels. In order to rationalize the absence of toxicity, the electrostatic potential of leiuropeptide II was compared to that of leiurotoxin I. The peptide is characterized by a large negative zone around Glu4, Asp5 and Asp8 residues, beginning in the neighbourhood of the beta-turn and extending along the helix. In the same area, leiurotoxin I exhibits a positive surface, around Arg6 and Arg13 basic residues, which are essential for its receptor affinity.

Purification, characterization, and synthesis of an inward-rectifier K+ channel inhibitor from scorpion venom.

We have purified a protein inhibitor of an inward-rectifier K+ channel, ROMK1, from the venom of the scorpion Leiurus quinquestriatus var. hebraeus. The inhibitor is Lq2, a previously discovered blocker of voltage- and Ca2+-activated K+ channels. Mutations were made on the channel and the inhibitor, and the resulting effects were examined using an electrophysiological assay. The data show that Lq2 blocks the pore of ROMK1, and that the interaction surface on Lq2 is the same for binding to inward-rectifier, voltage-activated, or Ca2+-activated K+ channels. These findings support the notion that different classes of K+ channels have different gates but a similar K+-selective pore structure.

Solution structure of Lqh-8/6, a toxin-like peptide from a scorpion venom--structural heterogeneity induced by proline cis/trans isomerization.

Lqh-8/6 is a minor fraction isolated from the venom of the scorpion Leiurus quinquestriatus hebraeus. Here we describe the purification, amino acid sequencing and solution structure determination by NMR and molecular modeling of this peptide. Lqh-8/6 is a small polypeptide (38 residues) which contains 8 half-cystines and is highly similar to another venom component, chlorotoxin. Standard homonuclear methods were used to sequentially assign the proton NMR spectra and to collect spatial restraints for structure determination. Two populations, identified early in the assignment step, are in slow interconversion on the NMR timescale. The two conformers were shown to originate from a cis/trans peptidyl-prolyl isomerization. Using a distance geometry program and simulated annealing protocol under the NMR restraints we obtained 10 final structures for the major conformation (trans isomer). None of the structures showed NOE violations larger than 0.05 nm, and the rmsd value relative to the mean structure (considering the main chain atoms in well-defined secondary structure) is 0.07 nm. The three-dimensional structure contains a short alpha-helix strapped on a small antiparallel beta-strand and an N-terminal extended fragment. The sequence/structure and structure/function relationships of the new scorpion toxin-like peptide are discussed in the context of the present structure determination. This toxin shows a stable, highly populated cis conformer of a peptidyl-prolyl peptide bond.

NMR study of the toxin III of the scorpion Leiurus quinquestriatus quinquestriatus: secondary structure and global folding

NMR study of the toxin III of the scorpion Leiurus quinquestriatus quinquestriatus: secondary structure and global folding

Synthesis and characterization of leiurotoxin I analogs lacking one disulfide bridge: evidence that disulfide pairing 3-21 is not required for full toxin activity.

Leiurotoxin I (Lei-NH2), a toxin isolated from the venom of the scorpion Leiurus quinquestriatus hebraeus, is a blocker of the apamin-sensitive Ca(2+)-activated K+ channels. It is a 31-residue polypeptide cross-linked by three disulfide bridges which are presumably between Cys3-Cys21, Cys8-Cys26, and Cys12-Cys28. To investigate the role of these disulfides, analogs of Lei-NH2 lacking one disulfide bridge (i.e., [Abu3,21]Lei-NH2, [Abu8,26]Lei-NH2, and [Abu12,28]Lei-NH2) were chemically synthesized by selective replacement of each pair of half-cystines forming a bridge by two alpha-aminobutyrate (Abu) residues. The two disulfide pairings of the main folded form of the synthetic analogs were established by enzymatic proteolysis. They were as expected between Cys8-Cys26 and Cys12-Cys28 for [Abu3,21]Lei-NH2 but were unexpectedly between Cys3-Cys12 and Cys21-Cys28 for [Abu8,26]Lei-NH2 and between Cys3-Cys8 and Cys21-Cys26 for [Abu12,28]Lei-NH2. The synthetic peptides were tested in vitro for their capacity to compete with the binding of [125I]apamin to rat brain synaptosomes and in vivo for their neurotoxicity in mice. In both assays, [Abu3,21]Lei-NH2 exhibited full Lei-NH2-like activity whereas [Abu8,26]Lei-NH2 and [Abu12,28]-Lei-NH2 possessed only residual activities (< 2% native toxin activity). This suggests that disulfide bridge Cys3-Cys21 is not essential per se for high toxin activity. Circular dichroism (CD) spectroscopy of the three analogs showed that only [Abu3,21]Lei-NH2 exhibited a CD spectrum similar to that of Lei-NH2, suggesting they both adopt closely related conformations, in agreement with the pharmacological data. Structural models of the analogs were constructed on the basis of the disulfide pairing assignment and compared with that of Lei-NH2.

Determination of the Three-dimensional Solution Structure of Scyllatoxin by 1H Nuclear Magnetic Resonance

The three-dimensional solution structure of Scyllatoxin (leiurotoxin I) a venom peptide from the scorpion Leiurus quinquestriatus hebraeuswas determined at 1 Aring; resolution by homonuclear protein n.m.r. methods at 500 MHz. Data analysis and structure calculation followed conventional protocols inherent to DIANA and related programs with two exceptions. First, distance constraints were obtained from two-dimensional nuclear Overhauser spectra by a previously described partial relaxation matrix approach. Second, since the pairing pattern of the six cysteine residues was not established a priori, the unequivocal assignment of the disulfide bridges was achieved exclusively from the n.m.r. data by a statistical analysis of preliminary DIANA structures. In the final calculation we used 227 upper distance constraints, 67 torsional constraints from vicinal coupling constants and ten stereospecific assignments of β-methylene protons. Scyllatoxin folds into a compact ellipsoidal shape. An α-helix (defined with 0.24 Å resolution) spanning 2.5 turns from Leu5 till Ser14 is stabilized by Cys8-Cys26 and Cys12-Cys28 disulfide bridges to the carboxy-terminal strand of an anti-parallel β-sheet. The antiparallel β-sheet (defined at 0.66 Å resolution) extends from Leu18 to Val29 with a tight turn at Gly23-Asp24 and displays a right-handed twist. Scyllatoxin competes with the toxins apamin from Apis mellifera melliferaand PO5 from Androctonus mauretanicus mauretanicusfor the same or similar high conductance calcium-activated potassium channels. Consideration of presently known biological activities and three dimensional structures of these toxins suggest a different toxin-receptor interaction of scyllatoxin as compared to apamin and PO5.

Involvement of the l-arginine-nitric oxide synthase pathway in the relaxant responses of the rat isolated anococcygeus muscle to a scorpion ( Leiurus quinquestriatus quinquestriatus) venom

Venom from several species of scorpions can cause generalized depolarization of peripheral nerves with enhancement of neurotransmitter release. The effects of the venom (LQV) from the scorpion Leiurus quinquestriatus quinquestriatus were investigated using the rat isolated carbachol (CCh) precontracted anococcygeus muscle (Acm) mounted in Krebs solution containing phentolamine (5 μM). LQV (0.2 μg/ml and 1.5 μg/ml) markedly relaxed the tone of the CCh precontracted (non-stimulated) Acm by 31.5 ± 5.1% and 35.5 ± 4.7%, respectively; the onset was immediate after the high dose, followed by a slow and gradual return of the muscle tone to 88.7 ± 1.8% of the initial peak tension in 50.2 ± 4 min. Subsequent doses of LQV produced essentially no appreciable changes in muscle tone, but the addition of l-arginine (250 μM) or, especially, sodium nitroprusside (SNP; 1 μM) produced marked and rapid relaxations of the Acm. Similar results were obtained with LQV during electrical field stimulation (EPS) of the precontracted Acm; however, during the gradual return of the muscle tone from the relaxed state induced by LQV, the EPS-induced relaxant (NANC) responses were also progressively inhibited by 84.9 ± 3.4% and 64.3 ± 4.5 by LQV 0.2 μg/ml and 1.5 μg/ml, respectively. Tetrodotoxin (TTx; 2 μM) or N g -nitro- l-arginine methylester ( l-NAME; 50 μM) markedly inhibited the relaxant responses of the Acm to EPS as well as to LQV but not the responses to SNP: l-arginine (250 μM) partially restored the relaxant (NANC) responses of the Acm to EPS. Thus, the l-arginine-nitric oxide synthase-nitric oxide pathway is involved in mediating the marked relaxant responses of the CCh precontracted Acm to LQV.

Charybdotoxin and its effects on potassium channels

Over the last few years, a considerable amount of information has been obtained regarding K+ channels. Different areas of research have contributed to knowledge in this field. Charybdotoxin (ChTX), a 37-amino acid peptide isolated from venom of the scorpion Leiurus quinquestriatus var. hebraeus, represents a remarkable tool for studying K+ channels. With its use, it has been possible to purify the high-conductance Ca(2+)-activated K+ (maxi-K) channel to homogeneity and determine the subunit composition of this channel. This has led to the discovery of an auxiliary beta-subunit that, when coexpressed with the pore-forming subunit, mSlo, alters the biophysical and pharmacological properties of this latter subunit. With the feasibility of producing large amounts of ChTX by recombinant techniques and the knowledge of the three-dimensional structure of the peptide, it has been possible to carry out site-directed mutagenesis studies and obtain a picture of the interaction surface of the toxin with two channels, maxi-K and Shaker, and to derive a picture of the complementary surface of the receptor in these two channels. Finally, ChTX, and the more selective K+ channel toxins that were subsequently discovered, have provided us with unique tools not only to determine the functional role that K+ channels play in target tissues but also to develop the molecular pharmacology of these channels.

Prejunctional action of the venom from the scorpion Leiurus quinquestriatus on adrenergic transmission

Prejunctional action of the venom from the scorpion Leiurus quinquestriatus on adrenergic transmission

Isolation of crab paralysing polypeptides from the venom of the scorpion Leiurus quinquestriatus hebraeus

Isolation of crab paralysing polypeptides from the venom of the scorpion Leiurus quinquestriatus hebraeus

Ovariuterus and oocyte dimensions in the female buthid scorpion,Leiurus quinquestriatus, H. &amp; E. (Scorpiones: Buthidae), and the effect of higher temperature

Summary The ovariuterus of the scorpion Leiurus quinquestriatus consists of three longitudinal tubes with five tubes interconnecting them. Oocytes of various sizes are situated on very short pedicles on the ventral side of the ovariuterus tubes. About 50 of either large or small oocytes could be observed under the dissecting microscope. Smaller ones (less than 50 pm in diameter) could only be recognized using a scanning electron microscope or in microscopic sections. Such means also allowed the identification of scars left by shrunken pedicles following parturition. The entire ovariuterus of a female was screened under SEM, and all visible oocytes were counted. In females kept at 25°C, growth and maturation of the small oocytes were accelerated, markedly increasing the number of large oocytes. Moreover, a large number of small, yellow, degenerated oocytes was noticed in these females.