Electrophysiological characterization of a novel small peptide from the venom of Conus californicus that targets voltage-gated neuronal Ca 2þ channels Johanna Bernaldez b,1 , Omar López a, 1 , Alexei Licea b , Emilio Salceda a , Rogelio O. Arellano c , Rosario Vega a , Enrique Soto a, * a Instituto de Fisiología, Benemérita Universidad Autónoma de Puebla, Mexico b Laboratorio de Inmunología Molecular y Biotoxinas, Departamento de Biotecnología Marina (C.I.C.E.S.E.), Ensenada, B.C., Mexico c Instituto de Neurobiología, Universidad Nacional Autónoma de México, Querétaro, Mexico article info Article history: Received 8 June 2010 Received in revised form 7 September 2010 Accepted 25 September 2010 Available online 7 October 2010 Keywords: Conus californicus Conotoxins Calcium channel Electrophysiology Xenopus laevis Dorsal-root ganglion neurons abstract Conus californicus belongs to a genus of marine gastropods with more than 700 extant species. C. californicus has been shown to be distantly related to all Conus species, but showing unusual biological features. We report a novel peptide isolated from C. californicus with a significant inhibitory action over neuronal voltage-gated calcium channels. The new toxin is formed by 13-amino acid residues with two disulfide bonds, whose sequence (NCPAGCRSQGCCM) is strikingly different from regular u-conotoxins. In the HPLC purification procedure, the venom fraction eluted in the first 10–15 min produced a significant decrease (54%  3%) of the Ca 2þ current in Xenopus laevis oocytes transfected with purified rat-brain mRNA. A specific peptide obtained from the elution at 13 min decreased the Ca 2þ current in the adult rat dorsal-root ganglion neurons in a primary culture by 34%  2%. The cysteine pattern of this peptide corresponds to the framework XVI described for the M-superfamily of conopeptides and is unprecedented among Conus peptides acting on Ca 2þ channels. Ó 2010 Elsevier Ltd. All rights reserved. 1. Introduction Over 50 million years of evolution, the predatory marine snails of the genus Conus have evolved from primitive gastropods, developing specific toxins used to hunt a remark- ably wide phylogenetic range of prey species including bivalves, cephalopods, crustacea, other gastropods, hemi- chordata, polychaete worms, echiuroids, and other fast- moving prey, such as fish predator species (Nybakken, 1990). The genus Conus is the largest single genus of venemous animals known (Kohn, 1990) with ca. 700 species, and each species could express between 100 and 200 venom peptides (Olivera, 2006). Several studies of the genus Conus show that the venom composition of cone snails possesses a sophisticated biochemical arsenal of conotoxins to rapidly immobilize their prey (Jones and Bulaj, 2000). Due to a rapid divergence in venom peptide sequences as specia- tion occurs, the sequences of peptides are notoriously different when compared different Conus species (Olivera, 2006). Therefore, it has been estimated that the number of different peptides that can be expressed in the venom of living Conus species is around 70,000 (Olivera, 2006). Many of these peptides have specific actions on ion channel and receptor subtypes with a high binding affinity (Jones and Bulaj, 2000). Conopeptides have been divided into two groups according to the number of disulfide bridges present in its structure: the non-disulfide-rich peptides, which lack * Corresponding author. Instituto de Fisiología, BUAP, Apartado Postal 406, 72000 Puebla, Pue, Mexico. Tel.: þ52 222 2295500x7316; fax: þ52 222 2295500x7301. E-mail addresses: esoto@siu.buap.mx, esoto24@gmail.com (E. Soto). 1 These two authors contributed equally to this work. Contents lists available at ScienceDirect Toxicon journal homepage: www.elsevier.com/locate/toxicon 0041-0101/$ – see front matter Ó 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.toxicon.2010.09.015 Toxicon 57 (2011) 60–67