RESUMO
This work reports the cloning, expression, and purification of a 42-kDa fragment of the SpaA protein from Erysipelothrix rhusiopathiae, the main antigenic candidate for a subunit vaccine against swine erysipelas. The use of an auto-induction protocol to improve heterologous protein expression in recombinant Escherichia coli cultures was also investigated. The cellular growth pattern and metabolite formation were evaluated under different induction conditions. The His-tagged protein was over-expressed as inclusion bodies, and was purified by a single chromatography step under denaturing conditions. Auto-induction conditions were shown to be an excellent process strategy, leading to a high level of rSpaA expression (about 25 % of total cellular protein content) in a short period of time.
Assuntos
Antígenos de Bactérias/isolamento & purificação , Proteínas de Bactérias/genética , Erysipelothrix/genética , Erisipela Suína/microbiologia , Animais , Antígenos de Bactérias/química , Antígenos de Bactérias/genética , Proteínas de Bactérias/química , Cromatografia de Afinidade , Clonagem Molecular , Escherichia coli/genética , Expressão Gênica , Corpos de Inclusão , Peso Molecular , Desnaturação Proteica , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Suínos , Erisipela Suína/imunologiaRESUMO
Serotypes of 46 previously unclassified isolates of Erysipelothrix rhusiopathiae from porcine tissues in the United States and serotypes of 31 isolates of the organism from porcine tissues received from Puerto Rico were determined. The 46 isolates from the United States were classified in serotype 21. Four isolates (from Georgia, Minnesota, Ohio, and Oklahoma) were tested and found to be pathogenic for swine. Serotypes 1 (subtypes 1a and 1b), 2, 5, 6, and 21 were found in porcine tissues from Puerto Rico. The relative frequency of the various serotypes was similar to that previously reported in the United States.