The venom of the scorpion Androctonus mauretanicus mauretanicus contains a toxin, P05, which is structurally and functionally similar to scorpion leiurotoxin I (87% sequence identity), a blocker of the apamin-sensitive Ca(2+)-activated K+ channels. It is a 31-residue polypeptide cross-linked by three disulfide bridges. A C-terminal carboxyl-amidated analog of P05 (sP05-NH2) was chemically synthesized by the solid-phase technique and fully characterized. Toxicity assays in vivo established that sP05-NH2, like native P05, is a potent and lethal neurotoxic agent in mice (LD50 of 20 ng per mouse). Pharmacological assays in vitro however showed that, unlike P05 which has a binding affinity of 2 x 10(-11) M, sP05-NH2 apparently binds irreversibly to the apamin receptor. Iodination at the C-terminal His gave diiodo-sP05-NH2, which had a binding affinity similar to that of native P05. The disulfide bridge pairings were chemically determined for sP05-NH2 and thereby deduced for P05 and leiurotoxin I: linkages were between Cys3 and Cys21, Cys8 and Cys26, and Cys12 and Cys28. Molecular dynamics refinement of P05 also using data from leiurotoxin I suggests that P05 is mainly composed of a double-stranded, antiparallel beta-sheet (from Leu18 to Val29) linked to an alpha-helix (from Arg6 to Gly16) by two disulfides (Cys8-Cys26 and Cys12-Cys28) and to an extended fragment (from Thr1 to Leu5) by the third disulfide (Cys3-Cys21). In agreement with the model, circular dichroism analysis of sP05-NH2 showed that the toxin structure is highly rigid.(ABSTRACT TRUNCATED AT 250 WORDS)

Three novel peptide inhibitors of the SK,,< channels were purdied to homogeneity from the venom of the scorpion Androctonus ntauretanicus muuretunzcus using one step of RP-HPLC and competrtton assays with ["SI]apamin to rat brain synaptosomes PO,, PO? and PO, have K,, of 100, 100 and 0.02 nM, respectively, for the apamm binding site. The sequence of PO3 was established and compared to that of other scorpion toxins active on K' channels: it contains 31 residues and has a free carboxyl end. It shares sequence srmilarny wnh apamin and leiurotoxm 1.

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A new peptide ligand of the small conductance Ca2+ activated K+ channels has been purified from the venom (obtained by manual rather than electrical stimulation of the scorpion Androctonus mauretanicus mauretanicus), by following the inhibition of the 125I‐apamin binding to its receptor on rat brain synaptosomes. Only one step on a C18 reversed‐phase high‐performance liquid chromatography column was necessary to obtain PO1. Its K0.5 for the apamin binding site was 100 nM. The amino acid sequence of PO1 is different from those of leiurotoxin and PO5. For the first time the same peptide was also purified from the venoms of two other species of North African scorpions, Androctonus austmlis and Buthus occitanus tunetanus. PO1 was chemically synthesized by the solid‐phase technique and fully characterized. A model of PO1 was constructed by amino acid replacement using PO5 nuclear magnetic resonance studies as the starting model. Structure‐activity relationships between these toxins and their receptor are discussed.