title: Development of an immunoaffinity method for purification of streptokinase.
abstract: BACKGROUND
  Streptokinase is a potent activator of plasminogen to plasmin, the enzyme that can solubilize the fibrin network in blood clots. Streptokinase is currently used in clinical medicine as a thrombolytic agent. It is naturally secreted by β-hemolytic streptococci.
  
  
  METHODS
  To reach an efficient method of purification, an immunoaffinity chromatography method was developed that could purify the streptokinase in a single step with high yield. At the first stage, a CNBr-Activated sepharose 4B-Lysine column was made to purify the human blood plasminogen. The purified plasminogen was utilized to construct a column that could purify the streptokinase. The rabbit was immunized with the purified streptokinase and the anti-streptokinase (IgG) purified on another streptokinase substituted sepharose-4B column. The immunoaffinity column was developed by coupling the purified anti-Streptokinase (IgG) to sepharose 6MB-Protein A. The Escherichia coli (E.coli) BL21 (DE3) pLysS strain was transformed by the recombinant construct (cloned streptokinase gene in pGEX-4T-2 vector) and gene expression was induced by IPTG. The expressed protein was purified by immunoaffinity chromatography in a single step.
  
  
  RESULTS
  The immunoaffinity column could purify the recombinant fusion GST-SK to homogeneity. The purity of streptokinase was confirmed by SDS-PAGE as a single band of about 71 kD and its biological activity determined in a specific streptokinase assay. The yield of the purification was about 94%.
  
  
  CONCLUSIONS
  This method of streptokinase purification is superior to the previous conventional methods.
authors: Department of Biology, Faculty of Science, Alzahra University, Tehran, Iran.
authors: Zohreh
authors: Z
authors: Karimi
authors: Mohammad
authors: M
authors: Babashamsi
authors: Ezat
authors: E
authors: Asgarani
authors: Ali
authors: A
authors: Salimi
keywords: Chromatography; Purification; Streptokinase; Thrombolytic agent