Efficient expression of recombinant human telomerase inhibitor 1 (hPinX1) in Pichia pastoris
PinX1 encoded by a remarkable tumor suppressor gene and located in human chromosome 8p23 is known as telomerase inhibitor. In recent years, this protein has been of interest as clinically tumor suppressor. Pichia pastoris expression system is preferred to produce heterologous proteins and is suitable for industrial and research purposes. In the present study, human PinX1 gene (hPinX1) was cloned in E. coli One Shot TOP10 cells and overexpressed in P. pastoris strain X-33 intracellularly, using a strong AOX (alcohol oxidase) promoter. The recombinant cells were grown in shaking flask. Induction time, methanol concentration and initial pH were optimized for obtaining high levels of hPinX1 protein production. Recombinant protein production was confirmed by Western blot analysis and the relative expression levels of rhPinX1 were quantified. According to Western blot analysis, molecular mass of produced hPinX1 was determined as 47.5 kDa. At the end of optimization studies, the best fermentation conditions were determined as induction time 48 h, methanol concentration 3% and initial culture pH 5.0. This process would be an applicable way for obtaining recombinant hPinX1 using P. pastoris expression system. This is the first report on recombinant production of hPinX1 in P. pastoris.