Definition of the alpha-KTx 15 subfamily

Abstract

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 μM 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: α-KTx 15. 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 ( K d) was 0.21 nM. This K d value was confirmed by kinetic experiments ( k on=6.0×10 6 M −1 s −1 and k off=6.0×10 −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 ( K i 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 Ca 2+ 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.