title: Characterization, N-terminal sequencing and classification of Tolworthcin 524: A bacteriocin produced by Bacillus thuringiensis subsp. tolworthi.
abstract: Bacteriocins synthesized by entomopathogenic Bacillus thuringiensis are gaining attention owing to their inhibitory effects against a wide variety of pathogenic bacteria. In the present study, we purified and characterized Tolworthcin 524, a bacteriocin synthesized by B. thuringiensis subsp. tolworthi, and compared it with other bacteriocins synthesized by B. thuringiensis. Tolworthcin 524 was separated and purified from the secretome of B. thuringiensis by fast protein liquid chromatography with a gel filtration column to obtain yields of 17% and a specific activity of ∼3600U/mgprotein. The purified product showed two peptides of ∼9 and 6kDa with antimicrobial activity in a gel-screening assay. The purified product was analyzed by two-dimensional electrophoresis and the resolved peptides of ∼9 and 6kDa with isoelectric points of ∼8 were sequenced. Partial sequences (METPVVQPR and DWTCWSCLVCAACS) were obtained suggesting that the ∼9 and 6kDa correspond to the prebacteriocin and mature Tolworthcin 524, respectively. Sequences showed high identity with Thurincin H and Thuricin 17 and had a conserved motif with other bacteriocins of B. thuringiensis. Based on sequence data, Tolworthcin 524 was classified in subclass II.2 (Thuricin-like peptides) of the Bacillus bacteriocin classification scheme. The larger peptide did not harbor a sequence suggestive of a signal peptide neither did it contain the double-glycine (GG) motif characteristic of the secretion leader recognized by the ABC transport system. Implications of these properties in Tolworthcin 524 secretion are discussed.
authors.affiliation: Universidad de Guanajuato Campus Irapuato-Salamanca, División Ciencias de la Vida, Posgrado en Biosciencias, México.
authors.forename: Rubén D
authors.initials: RD
authors.lastname: Pacheco-Cano
authors.affiliation: Aniversidad Autónoma de Coahuila, Escuela de Ciencias Biológicas, Torreón, Coahuila 27104, México.
authors.forename: Norma M
authors.initials: NM
authors.lastname: de la Fuente-Salcido
authors.affiliation: Universidad de Guanajuato Campus Irapuato-Salamanca, División Ciencias de la Vida, Posgrado en Biosciencias, México; Departamento de Alimentos, Irapuato, Guanajuato 36500, México.
authors.forename: Rubén
authors.initials: R
authors.lastname: Salcedo-Hernández
authors.affiliation: Universidad de Guanajuato Campus Irapuato-Salamanca, División Ciencias de la Vida, Posgrado en Biosciencias, México; Departamento de Alimentos, Irapuato, Guanajuato 36500, México.
authors.forename: M Fabiola
authors.initials: MF
authors.lastname: León-Galván
authors.affiliation: California Baptist University, Department of Natural and Mathematical Sciences, 8432 Magnolia Avenue, Riverside, CA 92504, United States; Department of Entomology, University of California, Riverside, Riverside, CA 92521, United States.
authors.forename: Dennis K
authors.initials: DK
authors.lastname: Bideshi
authors.affiliation: Universidad de Guanajuato Campus Irapuato-Salamanca, División Ciencias de la Vida, Posgrado en Biosciencias, México; Departamento de Alimentos, Irapuato, Guanajuato 36500, México.
authors.forename: Gustavo
authors.initials: G
authors.lastname: Hernández-Guzmán
authors.affiliation: Universidad de Guanajuato Campus Irapuato-Salamanca, División Ciencias de la Vida, Posgrado en Biosciencias, México; Departamento de Alimentos, Irapuato, Guanajuato 36500, México. Electronic address: josebar@ugto.mx.
authors.forename: J Eleazar
authors.initials: JE
authors.lastname: Barboza-Corona
mesh_terms: D000595:Amino Acid Sequence; D001413:Bacillus thuringiensis / Q000737:chemistry / Q000378:metabolism*; D001430:Bacteriocins / Q000737:chemistry* / Q000145:classification* / Q000302:isolation & purification; D002851:Chromatography, High Pressure Liquid; D015180:Electrophoresis, Gel, Two-Dimensional; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry
chemical_list: D001430:Bacteriocins; D010455:Peptides; C568940:thurincin H; C059740:thuricin
keywords: Bacillus thuringiensis; Bacteriocin; Secretion peptide; Tolworthcin 524