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Expression，Purification of Fusion Protein TGFα-PE40 and the Cytotoxicity of TGFα-PE40 on Tumor Cells
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Abstract:
Objective：To express and purify transforming growth factor α (TGFα)-pseudomonas exotoxin 40 and investigate its cytotoxic effect on cancer cells overexpressing epidermal growth factor (EGF) receptor. Methods：Recombinant plasmid pV28 was constructed by inserting the gene coding TGFα-PE40 into the vector pET28a Expression of fusion protein was conducted using the host BL21. Production of the recombinant protein was induced by IPTG, following extraction and purification of inclusion bodies with His-tag purification system. Cell viability assay (by MTT) was performed to determine the cytotoxic effect of TGFα-PE40 on cancer cells (A431 and SK-OV3).Results：Recombinant plasmid pV28, which expresses TGFα-PE40, was constructed successfully. Purity of TGFα-PE40 was about 98% after a purification procedure using His-tag column. Cytotoxic experiment showed that at a concentration of 0.86±0.07 μg/ml, TGFα-PE40 could reduce 50% viability of A431, which has high expression of EGFR. Whereas the IC50 for ovarian cancer cell SK-OV3, which expresses less EGFR, was 6.37±2.18 μg/ml.There was a significant difference between these two groups (P<0.05). Conclusion：The cytotoxicity ability of the fusion protein on tumor cells depends on the number of the EGFR presented on tumor cells.
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