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Cloning and Expression of EGFR Tyrosine Kinase and Construction of the Screening Model for the Enzymatic Inhibitors

Keywords: EGFR tyrosine kinase,Cloning and expresssion,Recombinant protein,The screening model for inhibitors

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As a member of tyrosine kinases (TK) family, Epidermal growth factor receptor(EGFR) has an activity of intrinsic protein tyrosine kinase, which plays an essential role in the regulation of signal transduction in the cells. Due to the abnormal expression of EGFR-TK, the certain type cancers may developed and progressed. Based on that, the inhibitors of EGFR-TK could be effective medicines for the treatment of cancer. In this study, the EGFR-TK domain was amplified by RT-PCR with RNA of HUVCEs cells as the template and expressed in E.coli BL21(DE3) using plasmid pET30a as vector. The recombinant protein was purified with the affinity chromatography (Ni-NTA), which was identified to have kinase activity catalyzing the substrate phosphorylated with ATP in the enzymatic reaction. Using the recombinant EGFR-TK as target, the screening model for enzymatic inhibitors was constructed.


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