Expression, Purification and in vitro Activity of Apoptin.
A novel protein named apoptin induces a p53-independent, Bcl-2 insensitive type apoptosis in various human tumor cells but not in normal cells. Apoptin is considered to be a promising candidate for safe and effective anti-tumortherapy. Moreover, apoptin may be important for fundamental studies on molecularbasis of apoptosis and cancer. Apoptin gene was subcloned into prokaryotic expression vector pPROEXHT and pBV220, respectively. The apoptin protein was obtained from pPROEXHT-apoptin expression system and not from pBV220-apoptin the former contains a 6xhistidine affinity tag for ease of purification. Expressed protein was purified by chromatography on a co-chelated affinity column and was renatured by dialysis. The renatured apoptin was able to induce purified Hela nuclei apoptosis in vitro even without the participation of the cytoplasm, showing that apoptin expressed in E.coli was active to induce apoptosis.
Sun
Guo-Jing
GJ
Institute of Radiation Medicine, Academy of Military Medical Sciences, Beijing 100850, China. sunzx@nic.bmi.ac.cn
Tong
Xin
X
Dong
Yan
Y
Sun
Zhi-Xian
ZX