title: Isolation, purification and functional characterization of alpha-BnIA from Conus bandanus venom.
abstract: We report the isolation and characterization by proteomic approach of a native conopeptide, named BnIA, from the crude venom of Conus bandanus, a molluscivorous cone snail species, collected in the South central coast of Vietnam. Its primary sequence was determined by matrix-assisted laser desorption/ionization time-of-flight tandem mass spectrometry using collision-induced dissociation and confirmed by Edman's degradation of the pure native fraction. BnIA was present in high amounts in the crude venom and the complete sequence of the 16 amino acid peptide was the following GCCSHPACSVNNPDIC*, with C-terminal amidation deduced from Edman's degradation and theoretical monoisotopic mass calculation. Sequence alignment revealed that its -C1C2X4C3X7C4- pattern belongs to the A-superfamily of conopeptides. The cysteine connectivity of BnIA was 1-3/2-4 as determined by partial-reduction technique, like other α4/7-conotoxins, reported previously on other Conus species. Additionally, we found that native α-BnIA shared the same sequence alignment as Mr1.1, from the closely related molluscivorous Conus marmoreus venom, in specimens collected in the same coastal region of Vietnam. Functional studies revealed that native α-BnIA inhibited acetylcholine-evoked currents reversibly in oocytes expressing the human α7 nicotinic acetylcholine receptors, and blocked nerve-evoked skeletal muscle contractions in isolated mouse neuromuscular preparations, but with ∼200-times less potency.
authors.affiliation: CNRS, Centre de Recherche de Gif - FRC3115, Institut de Neurobiologie Alfred Fessard - FRC2118, Laboratoire de Neurobiologie et Développement - UPR3294, F-91198 Gif-sur-Yvette, France; University of Nha Trang, Institute of Biotechnology and Environment, Nha Trang, Khanh Hoa 57000, Viet Nam. Electronic address: Bao.Nguyen@inaf.cnrs-gif.fr.
authors.forename: Bao
authors.initials: B
authors.lastname: Nguyen
authors.affiliation: CNRS, Centre de Recherche de Gif - FRC3115, Institut de Chimie des Substances Naturelles - UPR 2301, F-91198 Gif-sur-Yvette, France.
authors.forename: Jean-Pierre
authors.initials: JP
authors.lastname: Le Caer
authors.affiliation: CNRS, Centre de Recherche de Gif - FRC3115, Institut de Neurobiologie Alfred Fessard - FRC2118, Laboratoire de Neurobiologie et Développement - UPR3294, F-91198 Gif-sur-Yvette, France.
authors.forename: Romulo
authors.initials: R
authors.lastname: Aráoz
authors.affiliation: CEA, iBiTec-S, Service d'Ingénierie Moléculaire des Protéines, F-91191 Gif-sur-Yvette, France.
authors.forename: Robert
authors.initials: R
authors.lastname: Thai
authors.affiliation: University of Nha Trang, Institute of Biotechnology and Environment, Nha Trang, Khanh Hoa 57000, Viet Nam.
authors.forename: Hung
authors.initials: H
authors.lastname: Lamthanh
authors.affiliation: CNRS, Centre de Recherche de Gif - FRC3115, Institut de Neurobiologie Alfred Fessard - FRC2118, Laboratoire de Neurobiologie et Développement - UPR3294, F-91198 Gif-sur-Yvette, France.
authors.forename: Evelyne
authors.initials: E
authors.lastname: Benoit
authors.affiliation: CNRS, Centre de Recherche de Gif - FRC3115, Institut de Neurobiologie Alfred Fessard - FRC2118, Laboratoire de Neurobiologie et Développement - UPR3294, F-91198 Gif-sur-Yvette, France. Electronic address: Jordi.Molgo@inaf.cnrs-gif.fr.
authors.forename: Jordi
authors.initials: J
authors.lastname: Molgó
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002853:Chromatography, Liquid; D052078:Conus Snail / Q000737:chemistry*; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D008978:Mollusk Venoms / Q000737:chemistry* / Q000302:isolation & purification* / Q000633:toxicity; D017386:Sequence Homology, Amino Acid; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D008978:Mollusk Venoms; C000600076:alpha-BnIA, Conus bandanus
keywords: Alpha-conotoxin; Conus bandanus; Mass spectrometry; Nicotinic acetylcholine receptors