title: Solution structure of Pi4, a short four-disulfide-bridged scorpion toxin specific of potassium channels.
abstract: Pi4 is a short toxin found at very low abundance in the venom of Pandinus imperator scorpions. It is a potent blocker of K(+) channels. Like the other members of the alpha-KTX6 subfamily to which it belongs, it is cross-linked by four disulfide bonds. The synthetic analog (sPi4) and the natural toxin (nPi4) have been obtained by solid-phase synthesis or from scorpion venom, respectively. Analysis of two-dimensional (1)H NMR spectra of nPi4 and sPi4 indicates that both peptides have the same structure. Moreover, electrophysiological recordings of the blocking of Shaker B K(+) channels by sPi4 (K(D) = 8.5 nM) indicate that sPi4 has the same blocking activity of nPi4 (K(D) = 8.0 nM), previously described. The disulfide bonds have been independently determined by NMR and structure calculations, and by Edman-degradation/mass-spectrometry identification of peptides obtained by proteolysis of nPi4. Both approaches indicate that the pairing of the half-cystines is (6)C-(27)C, (12)C-(32)C, (16)C-(34)C, and (22)C-(37)C. The structure of the toxin has been determined by using 705 constraints derived from NMR data on sPi4. The structure, which is well defined, shows the characteristic alpha/beta scaffold of scorpion toxins. It is compared to the structure of the other alpha-KTX6 subfamily members and, in particular, to the structure of maurotoxin, which shows a different pattern of disulfide bridges despite its high degree of sequence identity (76%) with Pi4. The structure of Pi4 and the high amounts of synthetic peptide available, will enable the detailed analysis of the interaction of Pi4 with K(+) channels.
authors.affiliation: Unité de RMN des Biomolécules (CNRS URA 2185), Dépt. de Biologie Structurale et Chimie, Institut Pasteur, 75724 Paris Cedex 15, France.
authors.forename: J Iñaki
authors.initials: JI
authors.lastname: Guijarro
authors.forename: Sarrah
authors.initials: S
authors.lastname: M'Barek
authors.forename: Froylan
authors.initials: F
authors.lastname: Gómez-Lagunas
authors.forename: Damien
authors.initials: D
authors.lastname: Garnier
authors.forename: Hervé
authors.initials: H
authors.lastname: Rochat
authors.forename: Jean-Marc
authors.initials: JM
authors.lastname: Sabatier
authors.forename: Lourival
authors.initials: L
authors.lastname: Possani
authors.forename: Muriel
authors.initials: M
authors.lastname: Delepierre
authors.forename: Lourrival
authors.initials: L
authors.lastname: Possani
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D004220:Disulfides / Q000737:chemistry; D004594:Electrophysiology; D007313:Insecta; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry; D015221:Potassium Channels / Q000737:chemistry*; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D012604:Scorpion Venoms / Q000737:chemistry*; D012605:Scorpions
chemical_list: D004220:Disulfides; D010455:Peptides; C114245:Pi4 peptide; D015221:Potassium Channels; D012604:Scorpion Venoms