RESUMEN
OBJECTIVES: To display a recombinant avidin fused to the autotransporter ShdA to bind biotinylated molecules on the surface of Escherichia coli. RESULTS: Two chimeric protein constructs containing avidin fused to the autotransporter ShdA were expressed on the surface of Escherichia coli DH5α. One fusion protein contained 476 amino acids of the ShdA α and ß domains, whereas the second consisted of a 314 amino acid from α and truncated ß domains. Protein production was verified by SDS-PAGE using an antibody to the molecular FLAG-tag. The surface display of the avidin-shdA fusion protein was confirmed by confocal microscopy and flow cytometry analysis, and the biotin-binding activity was evaluated by fluorescence microscopy and flow cytometry using biotin-4-fluorescein and biotinylated-ovalbumin (OVA). CONCLUSIONS: Expression of a recombinant avidin with biotin-binding activity on the surface of E. coli was achieved using the autotransporter ShdA. This system is an alternative to bind biotinylated molecules to E. coli.
Asunto(s)
Avidina/metabolismo , Proteínas de la Membrana Bacteriana Externa/metabolismo , Escherichia coli/metabolismo , Proteínas de la Membrana/metabolismo , Proteínas Recombinantes de Fusión/metabolismo , Avidina/química , Avidina/genética , Proteínas de la Membrana Bacteriana Externa/química , Proteínas de la Membrana Bacteriana Externa/genética , Proteínas Bacterianas/genética , Biotina/análogos & derivados , Biotina/metabolismo , Electroforesis en Gel de Poliacrilamida , Escherichia coli/citología , Escherichia coli/genética , Fluoresceínas/metabolismo , Proteínas de la Membrana/química , Proteínas de la Membrana/genética , Microscopía Confocal , Regiones Promotoras Genéticas/genética , Proteínas Recombinantes de Fusión/química , Proteínas Recombinantes de Fusión/genéticaRESUMEN
Human respiratory syncytial virus (HRSV) is a member of the Paramyxoviridae family, which causes lower respiratory tract infections in neonates and children younger than 5 years. Here, we report the complete genome sequence of HRSV, isolated from a nasopharyngeal swab of a pregnant woman with cardiac complications.