Identification, cloning and expression analysis of an alpha-CGTase produced by stain Y112

- Cloning and functional expression of a CGTase gene from Bacillus agaradhaerens Y112 in Escherichia coli.
- CGTase can efficiently convert starch to α-CD.
- The recombination enzyme was highly purified using Ni(II)-immobilized metal-affinity chromatography.

Cyclodextrin glycosyltransferase (CGTase) is an enzyme able to convert starch and other substrates into cyclodextrins (CDs). A marine strain Y112 producing α-CGTase was identified as Bacillus agaradhaerens Y112 by physiological and biochemical characterization, and 16S rDNA analysis. The gene coding for α-CGTase was cloned, sequenced and expressed in Escherichia coli BL21 (DE3) cells. Recombinant α-CGTase was purified in one-step chromatographic separation and its purity evaluated by SDS-PAGE, showing the presence of one band with a molecular mass of about 92 kDa. Additionally, enzymatic capability was analyzed by measuring the starch conversion, and resulted in about 45% of CDs obtained after 6 h of cyclodextrin reaction. Of these CDs, mainly α-CD was produced (70% of the total CDs yield), suggesting the potential of this CGTase for industrial applications.