A new subfamily of conotoxins belonging to the A-superfamily

Two novel conotoxins from vermivorous cone snails Conus pulicarius and Conus tessulatus designated as Pu14 1 and ts14a were identified by cDNA cloning and peptide purification respectively The signal sequence of Pu14 1 is identical to that of alpha-conotoxins while its predicted mature peptide pu14a shares high sequence similarity with ts14a with only one residue different in their first intercysteine loop which contains 10 residues and is rich in proline Both pu14a and ts14a contain four separate cysteines in framework 14 (C-C-C-C) Peptide pu14a was chemically synthesized air oxidized and the connectivity of its two disulfide bonds was determined to be C1-C3 C2-C4 which is the same as found in alpha-conotoxins The synthetic pu14a induced a sleeping symptom in mice and was toxic to freshwater goldfish upon intramuscular injection Using the Xenopus oocyte heterologous expression system 1 mu M of pu14a demonstrated to inhibit the rat neuronal alpha 3 beta 2-containing as well as the mouse neuromuscular alpha 1 beta 1 gamma delta subtypes of nicotinic acetylcholine receptors and then rapidly dissociated from the receptors However this toxin had no inhibitory effect on potassium channels in mouse superior cervical ganglion neurons According to the identical signal sequence to alpha-conotoxins the unique cysteine framework and molecular target of pu14a we propose that pu14a and ts14a may represent a novel subfamily in the A-superfamily designated as alpha 1-conotoxins (C) 2010 Elsevier Inc All rights reserved