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| Expression, Purification and Bioactivity Research of Human Angiostatin Kringles 1-3 |
| LI Zhuang-lin1, YAO Xue-jing2, YUAN Gui-yong1 |
1. Shandong Simcere-Medgenn Bio-Pharmaceutical Co., Ltd., Yantai 264006, China;
2. Yantai Rongchang Bioengineering Co., Ltd., Yantai 264006, China |
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Abstract Human angiostatin is composed of 1-4 kringles of plasminogen, which can inhibit the angiogenesis of tumor, the tumor growth and metastasis through preventing tumor's blood and nutrition from supplying. The complete encoding cDNA of human angiostatin Kringles 1-3 was isolated from human embryo liver with PCR and inserted to pPIC9K vector. Then, K1-3 was expressed in secretory P.pastoris expression system by induction of methanol and the expression yield of K1-3 in 5 L fermenter was 41.2 mg/L. The supernatant of cultivation liquid was collected, then concentrated and dialyzed. Recombinant human K1-3 was purified by affinity chromatography with Lysine Sepharose 4B. As a result, HPLC showed purity of K1-3 was more than 98% and the recovery yield was more than 95%. Recombinant human K1-3 inhibited significantly the migration of human microvascular endothelial cells and half inhibitory concentration was 1.86μg/ml. Research showed that K1-3 suppressed specifically the angiogenesis on the CAMs with 95% inhibition.
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Received: 11 August 2010
Published: 25 January 2011
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