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| Expression of Paenibacillus polymyxa EJS-3 Fibrinolytic Enzyme Gene in Pichia pastoris |
| QIAN Hui, ZHANG Chong, LU Zhao-xin, BIE Xiao-mei, ZHAO Hai-zhen, LV Feng-xia |
| College of Food Science and Technology, Nanjing Agricultural University, Nanjing 210095, China |
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Abstract A recombinant plasmid pET-DsbA/PPFE-I was constructed and used as template to amplify fibrinolytic enzyme gene PPFE-I. This was then used in Pichia pastoris expression vector pPICZαA / PPFE-I; pPICZαA / PPFE-I and integrated into the genome of Pichia pastoris SMD1168 by electroporation. Results showed that after methanol induction for 72h, enzyme activity was 286 IU / ml. When compared to the wild type, the enzyme activity had improved 2.6 fold. SDS-PAGE electrophoresis analysis showed that the recombinant fibrinolytic enzyme (rPPFE-I) was expressed. In human fibrin degradation test rPPFE-I was used to firstly degrade the α chain human fibrinogen, followed by β chain, while the γ chain degradation was slowest. Endogenous Paenibacillus polymyxa fibrinolytic enzyme gene expression in Pichia pastoris was achieved, this would provide a new way to develop thrombolytic drug from endophyte sources.
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Received: 22 August 2014
Published: 25 December 2014
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