Abstract:

The β-CGTase gene from Bacillus cereus was cloned in pBV220 plasmid after PCR amplification. The
recombinant plasmid was transformed into E. coli DH5α. After ampicillin-resistance screening and restriction enzyme
digestion analysis, we acquired recombinant E. coli DH5α/pBVcgt. The optimum fermentation conditions in shaking flask
were acquired as follows: OD600 nm = 1.0, initial culture temperature 30 ℃, initial medium pH 8.0, and gradient-temperature
induction at 39 ℃ for 0.5 h followed by 40 ℃ for 0.5 h, 41 ℃ for 1 h and 42 ℃ for 2 h. The activity of β-CGTase under
these fermentation conditions was increased from 445 to 956 U/mL, representing a 2.15-fold increase compared to that
obtained before optimization. Cloning and expression of the β-CGTase gene showed that this enzyme could be expressed
highly in E. coli expression system. Therefore, this study proves the possibility of protein engineering of β-CGTase for largescale
production of β-CD.

Key words: Bacillus cereus, β-cyclodextrin glucanotransferase, engineered bacteria, expression, culture conditions