AmmTx3 has been originally isolated from the venom of the scorpion Androctonus mauretanicus and is now offered by Smartox in its synthetic version. AmmTx3 binds on its target A-type potassium channel with an affinity of 66 pM in rat brain synaptosomes (K_v4 channels). As such, AmmTx3 should represent an excellent tool to study action potential firing frequency, spike initiation and waveform in excitable cells. AmmTx3 was indeed found to block A-type potassium currents from cerebellum granular cells and from striatum neurons but with IC₅₀ values of 130 nM. AmmTx3 appears to be a pore blocker of K_v4.2 and K_v4.3 and seems to require the expression of the K_v4 associated protein DPP6. It should also contribute to our understanding of learning, memory and behavior. AmmTx3 shares very high sequence homology with two other A-type potassium channel blockers, Aa1 from Androctonus australis (94%) and BmTx3 from Buthus martensi (91%), and according to binding experiments the same channel target as well. AmmTx3 is a member of the a-KTX 15 family of toxins and consist of a single chain of 37 amino acid residues, with an N-terminal pyroglutamate, cross-linked by three disulfide bridges.

Description:

AA sequence: Pyr-Ile-Glu-Thr-Asn-Lys-Lys-Cys⁸-Gln-Gly-Gly-Ser-Cys¹³-Ala-Ser-Val-Cys¹⁷-Arg-Lys-Val-Ile-Gly-Val-Ala-Ala-Gly-Lys-Cys²⁸-Ile-Asn-Gly-Arg-Cys³³-Val-Cys³⁵-Tyr-Pro-OHDisulfide bonds: Cys⁸-Cys²⁸, Cys¹³-Cys³³ and Cys¹⁷-Cys³⁵Length (aa): 37Formula: C₁₅₈H₂₆₃N₅₁O₄₇S₆ Molecular Weight: 3821.52  DaAppearance: white lyophilized solidSolubility: water or saline bufferCAS number:Source: SyntheticPurity rate: > 98%

Reference:

Dipeptidyl-peptidase-like-proteins confer high sensitivity to the scorpion toxin AmmTX3 to Kv4-mediated A-type K+ channels

K+ channels containing Kv4.2 and Kv4.3 pore-forming subunits mediate most of the subthreshold-operating somatodendritic A-type K+ current in CNS neurons. These channels are believed to be important in regulating the frequency of repetitive firing, the backpropagation of action potential into dendrites, and dendritic integration and plasticity. Moreover, they have been implicated in several diseases from pain to epilepsy and autism spectrum disorders. The lack of toxins that specifically and efficiently block these channels has hampered studies aimed at confirming their functional role and their involvement in disease. AmmTX3 and other related members of the α-KTX15 family of scorpion toxins have been shown to block the A-type K+ current in cultured neurons, but their specificity has been questioned because the toxins do not efficiently block the currents mediated by Kv4.2 or Kv4.3 subunits expressed in heterologous cells. Here we show that the high-affinity blockade of Kv4.2 and Kv4.3 channels by AmmTX3 depends on the presence of the auxiliary subunits DPP6 and DPP10. These proteins are thought to be components of the Kv4 channel complex in neurons and to be important for channel expression in dendrites. These studies validate the use of AmmTX3 as a blocker of the Kv4-mediated A-type K+ current in neurons.

Maffie JK., et al. (2013) Dipeptidyl-peptidase-like-proteins confer high sensitivity to the scorpion toxin AmmTX3 to Kv4-mediated A-type K+ channels. J Physiol. PMID: 23440961

Scorpion toxins that block transient currents (IA) of rat cerebellum granular cells

This communication is a revision of the state-of-the-art knowledge of the field of scorpion toxins specific for the K(+)-channels, responsible for the I(A) currents of granular cells of rat cerebellum, maintained in vitro culture. There are 6 members of the sub-family alpha-KTx15 known to affect the I(A) currents. They are: toxins Aa1 from Androctonus australis Garzoni, BmTx3 from Buthusmartensi Karch, AmmTx3 from Androctonus mauretanicus mauretanicus, AaTx1 and AaTx2 from A. australis Garzoni and Discrepin from Tityus discrepans. They share high sequence similarity, apart from Discrepin, which causes an irreversible effect on the I(A) currents and is the most thoroughly studied toxin of the sub-family alpha-KTx15. The three-dimensional structure of Discrepin was determined and a series of mutants were synthesized and assayed in the system with the aim of identifying possible amino acids or sequence segments responsible for the irreversible effect found. In this revision some unpublished original data are also included to foster future work on the field, as well as a short discussion on some relevant aspects still pending and possible limitations associated with the strategy proposed.

Prestipino G., et al. (2009) Scorpion toxins that block transient currents (IA) of rat cerebellum granular cells. Toxicol Lett. PMID: 19429236

Definition of the alpha-KTx15 subfamily

Three novel scorpion toxins, Aa1 from Androctonus australis, BmTX3 from Buthus martensi and AmmTX3 from Androctonus mauretanicus were shown able to selectively block A-type K+ currents in cerebellum granular cells or cultured striatum neurons from rat brain. In electrophysiology experiments, the transient A-current completely disappeared when 1 microM of the toxins was applied to the external solution whereas the sustained K+ current was unaffected. The three toxins shared high sequence homologies (more than 94%) and constituted a new ‘short-chain’ scorpion toxin subfamily: alpha-KTx15. Monoiododerivative of 125I-sBmTX3 specifically bound to rat brain synaptosomes. Under equilibrium binding conditions, maximum binding was 14 fmol/mg of protein and the dissociation constant (Kd) was 0.21 nM. This Kd value was confirmed by kinetic experiments (kon = 6.0 x 10(6) M(-1) s(-1) and koff = 6.0 x 10(-4) s(-1)). Competitions with AmmTX3 and Aa1 with 125I-sBmTX3 bound to its receptor on rat brain synaptosomes showed that they fully inhibited the 125I-sBmTX3 binding (Ki values of 20 and 44 pM, respectively), demonstrating unambiguously that the three molecules shared the same target in rat brain. A panel of toxins described as specific ligands for different K+, Na+ and Ca2+ channels were not able to displace 125I-sBmTX3 from its binding site. Thus, 125I-sBmTX3 is a new ligand for a still unidentified target in rat brain. In autoradiography, the distribution of 125I-sBmTX3 binding sites in the adult rat brain indicated a high density of 125I-sBmTX3 receptors in the striatum, hippocampus, superior colliculus, and cerebellum.

Vacher H., et al. (2004) Definition of the alpha-KTx15 subfamily. Toxicon. PMID: 15208021

Expanding the scorpion toxina-KTX 15 family with AmmTX3 fromAndroctonus mauretanicus

A novel toxin, AmmTX3 (3823.5 Da), was isolated from the venom of the scorpion Androctonus mauretanicus. It showed 94% sequence homology with Aa1 from Androctonus australis and 91% with BmTX3 from Buthus martensi which, respectively, block A-type K+ current in cerebellum granular cells and striatum cultured neurons. Binding and displacement experiments using rat brain synaptosomes showed that AmmTX3 and Aa1 competed effectively with 125I-labelled sBmTX3 binding. They fully inhibited the 125I-labelled sBmTX3 binding (Ki values of 19.5 pm and 44.2 pm, respectively), demonstrating unambiguously that the three molecules shared the same target in rat brain. The specific binding parameters of 125I-labelled AmmTX3 for its site were determined at equilibrium (Kd = 66 pm, Bmax = 22 fmol per mg of protein). Finally, patch-clamp experiments on striatal neurons in culture demonstrated that AmmTX3 was able to inhibit the A-type K+ current (Ki = 131 nm).

Vacher H., et al. (2002) Expanding the scorpion toxina-KTX 15 family with AmmTX3 fromAndroctonus mauretanicus. Eur. J. Biochem. PMID: 12473099