title: A novel conotoxin framework with a helix-loop-helix (Cs alpha/alpha) fold.
abstract: Venomous predatory animals, such as snakes, spiders, scorpions, sea anemones, and cone snails, produce a variety of highly stable cystine-constrained peptide scaffolds as part of their neurochemical strategy for capturing prey. Here we report a new family of four-cystine, three-loop conotoxins (designated framework 14). Three peptides of this family (flf14a-c) were isolated from the venom of Conus floridanus floridensis, and one (vil14a) was isolated from the venom of Conus villepinii, two worm-hunting Western Atlantic cone snail species. The primary structure for these peptides was determined using Edman degradation sequencing, and their cystine pairing was assessed by limited hydrolysis with a combination of CNBr and chymotrypsin under nonreducing, nonalkylating conditions in combination with MALDI-TOF MS analysis of the resulting peptidic fragments. CD spectra and nanoNMR spectroscopy of these conotoxins directly isolated from the cone snails revealed a highly helical secondary structure for the four conotoxins. Sequence-specific nanoNMR analysis at room temperature revealed a well-defined helix-loop-helix tertiary structure that resembles that of the Cs alpha/alpha scorpion toxins kappa-hefutoxin, kappa-KTx1.3, and Om-toxins, which adopt a stable three-dimensional fold where the two alpha-helices are linked by the two disulfide bridges. One of these conotoxins (vil14a) has a Lys/Tyr dyad, separated by approximately 6A, which is a conserved structural feature in K(+) channel blockers. The presence of this framework in scorpions and in cone snails indicates a common molecular imprint in the venom of apparently unrelated predatory animals and suggests a common ancestral genetic origin.
authors.affiliation: Department of Chemistry and Biochemistry and Center of Excellence in Biomedical and Marine Biotechnology, Florida Atlantic University, Boca Raton, Florida 33431, USA.
authors.forename: Carolina
authors.initials: C
authors.lastname: Möller
authors.forename: Sanaz
authors.initials: S
authors.lastname: Rahmankhah
authors.forename: Janelle
authors.initials: J
authors.lastname: Lauer-Fields
authors.forename: Jose
authors.initials: J
authors.lastname: Bubis
authors.forename: Gregg B
authors.initials: GB
authors.lastname: Fields
authors.forename: Frank
authors.initials: F
authors.lastname: Marí
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002942:Circular Dichroism; D020916:Conotoxins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D052078:Conus Snail / Q000737:chemistry*; D003553:Cystine / Q000737:chemistry; D019077:Helix-Turn-Helix Motifs; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009498:Neurotoxins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D019906:Nuclear Magnetic Resonance, Biomolecular; D010455:Peptides / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D020916:Conotoxins; D009498:Neurotoxins; D010455:Peptides; D003553:Cystine