P05, a new leiurotoxin I-like scorpion toxin: Synthesis and structure-activity relationships of the .alpha.-amidated analog, a ligand of calcium-activated potassium channels with increased affinity

Abstract

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)

Design and Characterization of a Highly Selective Peptide Inhibitor of the Small Conductance Calcium-activated K+Channel, SkCa2

“…Jurkat T-cells were used to assess the effect on endogenous hSKCa2 with symmetric internal and external potassium aspartate (165 mM potassium aspartate). (20,34,36) alterations in the side chains at these positions may allow PO5 to fit more tightly within the channel-binding pocket due to either shorter side-chain size or local change in backbone conformation (37,38). Similar results were obtained with these PO5 mutants on SKCa3 (Fig.…”

“…1 shows the sequence alignment of these toxins (35), three of which (Lei, PO5, and Pi1) contain a motif (RXCQ) reported to be important for binding to SK Ca channels. (20,36). Apamin contains an RRCQ sequence (Fig.…”

Multibranched V3 peptides inhibit human immunodeficiency virus infection in human lymphocytes and macrophages

“…Chemical synthesis of SPCs was performed by the solid-phase technique (23). The peptide chains were elongated stepwise on 4-(oxy-methyl)-phenylacetamidomethyl resin by using optimized t-butyloxycarbonyl-benzyl chemistry as previously described (33). Amino Viruses.…”

“…Such synthetic polymeric constructions (SPCs), consisting of an uncharged poly-Lys core matrix (which accounts for about 15% of the total weight), have been used as immunogens (36) and, in the case of the HIV-1 V3 * Corresponding author. Phone: (33) loop, elicited type-specific responses (29). The rationale for using V3 SPCs instead of monomeric V3 peptides as anti-HIV compounds is that they may have enhanced ligand avidity due to the high local density of binding sites for the potential V3 recognition sites.…”

Two Conserved Arginine Residues from the SK3 Potassium Channel Outer Vestibule Control Selectivity of Recognition by Scorpion Toxins

“…According to these studies, SK channel blockade would occur according to an "intermediate" mode of interaction (19) involving mainly the toxin ␣-helical structure rather than its ␤-sheet structure (20 -22). Two arginyl residues found in an RRCQ motif present in both P05 and apamin are critical for P05 interaction with its target, as described for apamin (23). In contrast, the ␤-sheet structure appears to be important for a "pore-blocking" mode as far as Kv channel block is concerned (19,24,25).…”