Expression, purification and refolding of streptavidin-tagged human tumor necrosis factor-alpha fusion protein / 南方医科大学学报
Journal of Southern Medical University
; (12): 412-415, 2009.
Artigo
em Chinês
| WPRIM (Pacífico Ocidental)
| ID: wpr-233775
Biblioteca responsável:
WPRO
ABSTRACT
<p><b>OBJECTIVE</b>To study the purification, refolding and bioactivity of streptavidin-tagged human tumor necrosis factor-alpha (SA-TNF-alpha) bi-functional fusion protein.</p><p><b>METHODS</b>SA-TNF-alpha fusion protein was expressed in BL21(DE3) host bacteria, purified using Ni-NTA affinity chromatography and refolded by dilution and dialysis followed by identification using Western blotting. The effect of SA-TNF-alpha fusion protein against L929 cells was evaluated by MTT assay. Flow cytometry was used to analyze the surface modification of biotinylated MB49 tumor cells by SA-TNF-alpha fusion protein.</p><p><b>RESULTS</b>Recombinant SA- TNF-alpha fusion protein was expressed in BL21(DE3) at about 30% of total bacterial protein, with a purity of about 95% after purification. The SA-TNF-alpha fusion protein existed as dimmers, tetramers and higher order structures after refolding. The fusion protein exhibited a bi-functionality by inhibiting L929 cells and SA-mediated high-affinity binding to biotinylated cell surfaces, with an anchor modification rate of above 90%.</p><p><b>CONCLUSION</b>The dimmers, tetramers and higher order structures of the obtained SA-TNF-alpha fusion protein all exhibit a bi-functionality, and may serve as a potential candidate therapeutic agent for tumors.</p>
Texto completo:
Disponível
Contexto em Saúde:
Doenças Negligenciadas
Problema de saúde:
Doenças Negligenciadas
/
Zoonoses
Base de dados:
WPRIM (Pacífico Ocidental)
Assunto principal:
Proteínas Recombinantes de Fusão
/
Química
/
Cromatografia de Afinidade
/
Fator de Necrose Tumoral alfa
/
Dobramento de Proteína
/
Estreptavidina
/
Escherichia coli
/
Genética
/
Metabolismo
/
Métodos
Tipo de estudo:
Estudo prognóstico
Limite:
Humanos
Idioma:
Chinês
Revista:
Journal of Southern Medical University
Ano de publicação:
2009
Tipo de documento:
Artigo