Genomic organization of the KTX2 gene, encoding a `short' scorpion toxin active on K+ channels

AbstractA single intron of 87 bp, close to the region encoding the C-terminal part of the signal peptide, was found in the gene of the `short' scorpion toxin kaliotoxin 2 of Androctonus australis acting on various types of K+ channels. Its A+T content was particularly high (up to 86%). By walking and ligation-mediated PCR, the promoter sequences of the kaliotoxin 2 gene of Androctonus australis were studied. The transcription unit of the gene is 390 bp long. Consensus sequences were identified. The genes of `short' scorpion toxins active on K+ channels are organized similarly to those of the `long' scorpion toxins active on Na+ channels and not like those of structurally related insect defensins, which are intronless

A new class of scorpion toxin binding sites related to an A-type K+ channel: pharmacological characterization and localization in rat brain

AbstractA new scorpion toxin (3751.8 Da) was isolated from the Buthus martensi venom, sequenced and chemically synthesized (sBmTX3). The A-type current of striatum neurons in culture completely disappeared when 1 μM sBmTX3 was applied (Kd=54 nM), whereas the sustained K+ current was unaffected. 125I-sBmTX3 specifically bound to rat brain synaptosomes (maximum binding=14 fmol mg−1 of protein, Kd=0.21 nM). A panel of toxins yet described as specific ligands for K+ channels were unable to compete with 125I-sBmTX3. A high density of 125I-sBmTX3 binding sites was found in the striatum, hippocampus, superior colliculus, and cerebellum in the adult rat brain

In vivo synergy of cardiotoxin and phospholipase A2 from the elapid snake Naja mossambica mossambica

International audienceThe lethality of elapid snake venoms is due to toxic polypeptides which are alpha-neurotoxins, phospholipases A2 and cardiotoxins. In contrast to alpha-neurotoxins, less is known about the mode of action of phospholipases A2 and cardiotoxins at a cellular and molecular level, although it has been demonstrated that the hemolytic effect of cardiotoxin and its action on muscle can be accelerated in vitro by phospholipase A2. Here we show that when mice are injected i.v. with phospholipase A2, a significant decrease in survival time is observed if a sub-LD50 amount of cardiotoxin is injected simultaneously. Furthermore, the survival time is drastically reduced when phospholipase A2 (2.3 LD50) is injected first, followed 15 min later by doses of cardiotoxin as small as 0.18 LD50. These results strongly suggest a synergy between the two polypeptides

Characterization of elapidae snake venom components using optimized reverse-phase high-performance liquid chromatographic conditions and screening assays for alpha-neurotoxin and phospholipase A2 activities

International audienceThe vast majority of Elapidae snake venoms, genus Naja, includes three classes of toxic polypeptides: alpha-neurotoxins, phospholipases A2, and cardiotoxins. A new experimental approach using reverse-phase high-performance liquid chromatography in particular has been developed, allowing their respective resolution, identification, and quantitation from milligram quantities of venom. First, definition of optimal chromatographic conditions for Naja mossambica mossambica toxins has been ascertained. Different column packing and solvent systems were compared for their efficiency, with particular attention to the ionic strength of the aqueous solvent. A medium-chain alkyl support (octyl) in conjunction with a volatile ammonium formate (0.15 M, pH 2.70)/acetonitrile solvent system was found to be particularly effective. All the components known until now from this venom could be resolved in a single step, and the elution order was alpha-neurotoxins, phospholipases A2, and cardiotoxins with a total recovery of absorbance and toxicity. Then, with these suitable conditions, we describe a new major cardiotoxin molecule in this venom by hydrophobic and not ionic-charge discrimination. Second, specific assays were designed to detect alpha-neurotoxin and phospholipase A2 activities in chromatographic fractions: alpha-neurotoxin activity was determined by competition for the binding of a radiolabeled alpha-neurotoxin to the acetylcholine receptor of the ray electric organ, and phospholipase A2 activity was defined by the enzymatic activity of these toxins with a fluorescent phospholipid as substrate. Finally, the applicability of these new methods to study other Naja snake venoms was demonstrated

Potassium Channels Blockers from the Venom of Androctonus mauretanicus mauretanicus

International audienceK(+) channels selectively transport K(+) ions across cell membranes and play a key role in regulating the physiology of excitable and nonexcitable cells. Their activation allows the cell to repolarize after action potential firing and reduces excitability, whereas channel inhibition increases excitability. In eukaryotes, the pharmacology and pore topology of several structural classes of K(+) channels have been well characterized in the past two decades. This information has come about through the extensive use of scorpion toxins. We have participated in the isolation and in the characterization of several structurally distinct families of scorpion toxin peptides exhibiting different K(+) channel blocking functions. In particular, the venom from the Moroccan scorpion Androctonus mauretanicus mauretanicus provided several high-affinity blockers selective for diverse K(+) channels (SK(Ca), K(v)4.x, and K(v)1.x K(+) channel families). In this paper, we summarize our work on these toxin/channel interactions

Use of high performance liquid chromatography to demonstrate quantitative variation in components of venom from the scorpion Androctonus australis hector

International audienceUsing reverse-phase high performance liquid chromatography (RP-HPLC), to resolve less than 1 mg of scorpion venom, quantitative variations in protein components were demonstrated in Androctonus australis Hector venoms obtained either by electric or manual stimulation. The results support polymorphism of scorpion venom components at an individual level

Structural determinants of fasciculin specificity for acetylcholinesterase

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Crystal structure of mouse acetylcholinesterase

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1.9 A resolution structure of fasciculin 1, an anti-acetylcholinesterase toxin from green mamba snake venom

International audienceThe crystal structure of fasciculin 1, a potent acetylcholinesterase inhibitor from green mamba snake venom, has been solved by the multiple isomorphous replacement method complemented with anomalous scattering and subsequently refined at 1.9-A resolution. The overall structure of fasciculin is similar to those of the short alpha-neurotoxins and cardiotoxins, with a dense core rich in disulfide bridges and three long loops disposed as the central fingers of a hand. A comparison of these three prototypic toxin types shows that fasciculin 1 has structural features that are intermediate between those of the other two molecules. Its core region, which can be defined as a continuous stretch of conserved residues, is very similar to that of erabutoxin b, whereas the orientation of its long loops resembles that of cardiotoxin VII4. This result introduces a new element in the study of phylogenetic relationships of snake toxins and suggests that, after divergency from an ancestral gene, convergent evolution may have played an important factor in the evolution of these proteins. In fasciculin 1, several arginine and lysine residues are well ordered and relatively exposed to the solvent medium and may play a role in the binding to the peripheral site of acetylcholinesterases