Preparation of Recombinant HIV-TATm-Survivin (T34A) Protein and Its Proapoptosis Activity to Four Cancer Cell Lines in vitro

In this study, the cDNA encoding survivin was cloned by RT-PCR obtained from human breast cancer cell lines, B-Cap37. After the mutation of survivin (T34A) by site-directed mutagenesis, an expression vector of pRSET-B-HIV-TATm-Survivin (T34A) was constructed by PCR. Subsequently, the resultant plasmid was transformed into E. coli BL21 (DE3). The recombinant HIV-TATm-Survivin (T34A) protein was expressed efficiently by inducing IPTG at a concentration of 0.5 mmol/L, reaching a yield of 650 mg/L (as inclusion body) in fermentation culture. The inclusion bodies were solubilized, refolded, and purified by ion exchange chromatography and size-exclusion chromatography to a purity of 96 %. Remarkable effects of the purified recombinant HIV- TATm-Survivin (T34A) on the morphology of cell line SW1990 and B-Cap37 were observed after being administrated for 4 h. MTT assay showed that the recombinant HIV-TATm-Survivin (T34A) protein could restrain the cell proliferation of SW1990, B-Cap37, and SSMC-7721, in vitro. Apoptosis rate and cell circle of SW1990 and B-Cap37 that were treated with the target protein (final concentration 60 μg/mL) were detected with flow cytometry. The results revealed that the apoptosis rate of SW1990 and B-Cap37 were 25.6 % and 19.3 %, respectively. More than 65 % of cancer cells were arrested at the G1 phase. The study suggested that TATm-Survivin (T34A) protein was a hopeful protein drug for the treatment of cancer by facilitating apoptosis of cancer cells.