Expression of α-Cyclodextrin Glycosyltransferase Gene of Gebacillius sp. CHB1 in Bacillus subtilis

  Objective  To construct a vector for expressing α-cyclodextrin glycosyltransferase (α-CGTase) gene in Bacillus subtilis.

  Method  The α-CGTase gene from Gebacillius sp. CHB1 was amplified by PCR. The EcoR Ⅰ-digested pBES and Xho Ⅰ-digested α-CGT gene were connected and transformed into B. subtilis RIK1285. Subsequently, fermentation of the recombinant B. subtilis RIK1285/pBE-CGT was optimized.

  Result  (1) The α-CGTase gene was expressed in a fermentation medium to show an enzymatic activity of 2.9 U·mL^-1 by B. subtilis RIK1285/pBE-CGT. (2) Medium TB with the formula of 0.5% glycerol, 1.2% peptone, 2.4% yeast extract, 1.64% K₂HPO₄, and 0.23% KH₂PO₄ was found to be optimal for the fermentation. After fermentation in TB at 37℃ for 24h, the α-CGTase activity reached 5.3 U·mL^-1, which was 8-fold of what the wild Gebacillius sp. CHB1 could generate.

  Conclusion  The engineered B. subtilis RIK1285/pBE-CGT was successfully obtained and the fermentation process optimized.