title: Molecular cloning and in silico characterization of knottin peptide, U2-SCRTX-Lit2, from brown spider (Loxosceles intermedia) venom glands.
abstract: Inhibitor cystine knots (ICKs) are a family of structural peptides with a large number of cysteine residues that form intramolecular disulfide bonds, resulting in a knot. These peptides are involved in a variety of biological functions including predation and defense, and are found in various species, such as spiders, scorpions, sea anemones, and plants. The Loxosceles intermedia venom gland transcriptome identified five groups of ICK peptides that represent more than 50 % of toxin-coding transcripts. Here, we describe the molecular cloning of U2-Sicaritoxin-Lit2 (U2-SCRTX-Lit2), bioinformatic characterization, structure prediction, and molecular dynamic analysis. The sequence of U2-SCRTX-Lit2 obtained from the transcriptome is similar to that of μ-Hexatoxin-Mg2, a peptide that inhibits the insect Nav channel. Bioinformatic analysis of sequences classified as ICK family members also showed a conservation of cysteine residues among ICKs from different spiders, with the three dimensional molecular model of U2-SCRTX-Lit2 similar in structure to the hexatoxin from μ-hexatoxin-Mg2a. Molecular docking experiments showed the interaction of U2-SCRTX-Lit2 to its predictable target-the Spodoptera litura voltage-gated sodium channel (SlNaVSC). After 200 ns of molecular dynamic simulation, the final structure of the complex showed stability in agreement with the experimental data. The above analysis corroborates the existence of a peptide toxin with insecticidal activity from a novel ICK family in L. intermedia venom and demonstrates that this peptide targets Nav channels.
authors.affiliation: Department of Cell Biology, Federal University of Paraná, Jardim das Américas, 81531-990, Curitiba, Paraná, Brazil.
authors.forename: Gabriel Otto
authors.initials: GO
authors.lastname: Meissner
authors.affiliation: Laboratory of Computational Biology and Bioinformatics, Federal University of ABC, Santo André, São Paulo, Brazil.
authors.forename: Pedro Túlio
authors.initials: PT
authors.lastname: de Resende Lara
authors.affiliation: Laboratory of Computational Biology and Bioinformatics, Federal University of ABC, Santo André, São Paulo, Brazil.
authors.forename: Luis Paulo Barbour
authors.initials: LP
authors.lastname: Scott
authors.affiliation: Laboratory of Computational Biology and Bioinformatics, Federal University of ABC, Santo André, São Paulo, Brazil.
authors.forename: Antônio Sérgio Kimus
authors.initials: AS
authors.lastname: Braz
authors.affiliation: Department of Cell Biology, Federal University of Paraná, Jardim das Américas, 81531-990, Curitiba, Paraná, Brazil.
authors.forename: Daniele
authors.initials: D
authors.lastname: Chaves-Moreira
authors.affiliation: Department of Cell Biology, Federal University of Paraná, Jardim das Américas, 81531-990, Curitiba, Paraná, Brazil.
authors.forename: Fernando Hitomi
authors.initials: FH
authors.lastname: Matsubara
authors.affiliation: Department of Cell Biology, Federal University of Paraná, Jardim das Américas, 81531-990, Curitiba, Paraná, Brazil.
authors.forename: Eduardo Mendonça
authors.initials: EM
authors.lastname: Soares
authors.affiliation: Department of Cell Biology, Federal University of Paraná, Jardim das Américas, 81531-990, Curitiba, Paraná, Brazil.
authors.forename: Dilza
authors.initials: D
authors.lastname: Trevisan-Silva
authors.affiliation: Department of Cell Biology, Federal University of Paraná, Jardim das Américas, 81531-990, Curitiba, Paraná, Brazil.|Laboratory of Molecular Immunopathology, Department of Clinical Pathology, Clinical Hospital of Federal University of Paraná, Curitiba, Paraná, Brazil.
authors.forename: Luiza Helena
authors.initials: LH
authors.lastname: Gremski
authors.affiliation: Department of Cell Biology, Federal University of Paraná, Jardim das Américas, 81531-990, Curitiba, Paraná, Brazil.
authors.forename: Silvio Sanches
authors.initials: SS
authors.lastname: Veiga
authors.affiliation: Department of Cell Biology, Federal University of Paraná, Jardim das Américas, 81531-990, Curitiba, Paraná, Brazil. olgachaim@ufpr.br.
authors.forename: Olga Meiri
authors.initials: OM
authors.lastname: Chaim
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D057771:Cystine-Knot Miniproteins / Q000737:chemistry*; D008958:Models, Molecular; D062105:Molecular Docking Simulation; D017434:Protein Structure, Tertiary; D013111:Spider Venoms / Q000737:chemistry*; D013112:Spiders / Q000737:chemistry*
chemical_list: D057771:Cystine-Knot Miniproteins; D013111:Spider Venoms
keywords: Brown Spider; ICK; Insecticide toxin; Molecular modeling; Peptide structure; Venom