ABSTRACT
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 beta-hemolytic streptococci. 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-Ac-tivated 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. 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%. This method of streptokinase purification is superior to the previous conventional methods.