Expression of a novel chimeric protein containing the A chain of tissue-type plasminogen activator and the B chain of pro-urokinase in insect cells using the baculovirus system. |
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| Authors: | C M Zhao H T Zhang M H Hu |
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| Institution: | College of Life Sciences, National Laboratory of Protein Engineering and Plant Genetic Engineering, Peking University, Beijing, P. R. China. |
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| Abstract: | A hybrid cDNA tu-pa, which contains Ser1-Thr263 of tissue-type plasminogen activator (t-PA) and Ser138-Leu411 of pro-urokinase (pro-UK) was constructed and expressed in the Sf9-AcNPV system. The expression level was approximate 2.5 mg/L. Tu-PA was purified via one-step affinity column conjugated with monoclonal antibody against the B chain of pro-UK, which showed a single band of approximate 60 kDa in SDS-PAGE. The specific activity of the chimeric protein on fibrin plate was 200,000 IU/mg protein. Tu-PA had a higher selectivity for fibrin than UK and pro-UK. Its activity can be promoted by CNBr degraded fibrin fragments as t-PA. |
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