Revista: | Electronic journal of biotechnology |
Base de datos: | PERIÓDICA |
Número de sistema: | 000358458 |
ISSN: | 0717-3458 |
Autores: | Díaz, Mauricio1 Venturini, Elena2 Marchetti, Stefano2 Arenas, Gloria3 Marshall, Sergio H3 |
Instituciones: | 1Pontificia Universidad Católica de Valparaíso, Escuela de Ingeniería Bioquímica, Valparaíso. Chile 2Universita di Udine, Dipartimento di Scienze Agrarie e Ambientali, Udine, Friuli Venezia Giuli. Italia 3Pontificia Universidad Católica de Valparaíso, Facultad de Ciencias, Valparaíso. Chile |
Año: | 2012 |
Volumen: | 15 |
Número: | 2 |
País: | Chile |
Idioma: | Inglés |
Tipo de documento: | Artículo |
Enfoque: | Experimental, aplicado |
Resumen en inglés | Different strategies have been used to overcome the difficulties to produce antimicrobial peptides. Here we used Intein Mediated Purification with an Affinity Chitin-binding Tag (IMPACT-System, New England Biolabs) for the expression of the antimicrobial peptide cecropin to reduce its sensitivity to intracellular proteases and use its inducible self-cleaving capability to remove the carrier. Cecropin was cloned into suitable expression vector pTYB11, and expression induced by IPTG in Escherichia coli ER2566. The use of 22ºC induction allowed the expression of cecropin with its intein carrier in soluble form. Cell extracts were purified by chitin affinity chromatography and intein-mediated splicing of the target protein was achieved by thiol addition, obtaining a final yield of 2.5 mg cecropin/l. Cecropin cleaved from the intein had its proper biologically active form, showing a micromolar antimicrobial activity against Vibrio ordalii, Vibrio alginolyticus and Escherichia coli |
Disciplinas: | Biología |
Palabras clave: | Biotecnología, Bioquímica, Péptidos antimicrobianos, Cecropinas, Expresión génica, Fusión celular, Escherichia coli |
Keyword: | Biology, Biotechnology, Biochemistry, Antimicrobial peptides, Cecropins, Gene expression, Cell fusion, Escherichia coli |
Texto completo: | Texto completo (Ver HTML) |