Design and Identification of a High Efficient Formic Acid Cleavage Site For Separation of Fusion Protein

文献类型： 外文期刊

第一作者： Zhang, Huaguang

作者： Zhang, Huaguang;Li, Mei;Shi, Shuangfeng;Yin, Chao;Jia, Shirong;Wang, Zhixing;Liu, Yuhui

作者机构：

关键词： Fusion protein;Asp-Pro bond;Formic acid cleavage;Cleavage efficiency

期刊名称：PROTEIN JOURNAL （ 影响因子：2.371； 五年影响因子：1.929 ）

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收录情况： SCI

摘要： The release of target protein with high efficiency and low cost from expressed fusion protein is a key requirement for commercial production of target proteins. To establish such a cleavage system, we have designed four formic acid (FA) cleavage sites C1 (DPDPDP), C2 (DPPDPP), C3 (DDDDPI) and C4 (IVDPNP), which was placed in between the E and G fusion protein. Four expression vectors were individually constructed and expressed in Escherichia coli. Purified proteins were reacted with a series of FA concentrations or under different temperatures followed by SDS-PAGE gel electrophoresis to verify the degree of cleavage efficiency. Results showed that the C2 was the most efficient site compared with the other three. After optimization of cleavage conditions for E-C2-G, the cleavage efficiently could reach as high as 87.3 % within 2.5 h in 37 % FA at 45 A degrees C. Comparing with previous reports, a significant reduction (26 %) of FA concentration at a lower temperature in a short duration of reaction (18 times less) was achieved. We believe the cleavage site of DPPDPP identified in this study can be used in the large-scale production of valuable fusion proteins to save the cost, time and energy.

分类号： Q51

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