Fermentation, purification and identification of recombinant RGD-Hirudin / 生物工程学报
Chinese Journal of Biotechnology
; (12): 126-129, 2004.
Artigo
em Chinês
| WPRIM (Pacífico Ocidental)
| ID: wpr-305215
Biblioteca responsável:
WPRO
ABSTRACT
Recombinant RGD-Hirudin ( r-RGD-Hirudin ) has double functions anti-thrombin activity and anti-platelet aggregation activity. To identify these functions, the expression plasmid, RGD-Hirudin-pPIC9K, was constructed by inserting cDNA of RGD-hirudin in yeast expression vector pPIC9K. The high expression clone was gained after screening. This clone was fermented for 3 days. The r-RGD-hirudin was secreted into the culture. It was ultra-filtrated from culture supernatant, then after gel filtration chromatography and anion exchange chromatography, the purified r-RGD-hirudin was gained. Its purity was larger than 97% and its specific activity was 12 000 ATU/mg. The yield per liter culture of purified r-RGD-hirudin was 1 g and overall recovery yield was more than 75% . The purified r-RGD-hirudin was identified by reductive SDS-PAGE, anti-thrombin activity assay, anti-platelet aggregation assay, LC/MS and isoelectrofocusing assay. It is proved that r-RGD-Hirudin is ramification of wt-Hirudin and it has anti-thrombin activity and anti-platelet aggregation activity.
Texto completo:
Disponível
Base de dados:
WPRIM (Pacífico Ocidental)
Assunto principal:
Farmacologia
/
Pichia
/
Proteínas Recombinantes
/
Inibidores da Agregação Plaquetária
/
Hirudinas
/
Ratos Sprague-Dawley
/
Fermentação
/
Genética
/
Peso Molecular
Limite:
Animais
Idioma:
Chinês
Revista:
Chinese Journal of Biotechnology
Ano de publicação:
2004
Tipo de documento:
Artigo