[Cultivation optimization of prolyl endopeptidase and high cell density fermentation]

Engineered E. coli BL21/pGEM-PEP could constitutively express recombinant prolyl endopeptidase from Aeromonas punctata, which was extremely effected by culture conditions, so fermentation conditions were optimized to obtain it's high expression level. Firstly, the stability of BL21/pGEM-PEP was investigated, then, culture temperature, pH, time, medium were optimized in shake flaskd. The L9(3(4)) orthogonal experiment confirmed that shaker speed, pH, culture temperature, culture time had high degree statistical meaning. Based on these data, high cell density fermentation of E. coli BL21/pGEM-PEP on NBS BioFlo 3000 5 L fermentor was achieved, after 20 h cultivation, the final density(dwt) was 60OD600(22.5 g/L), the expressed PEP was about 28% of total cellular protein, the yield was 3.15 g per litter broth.