A novel cyclodextrin glycosyltransferase from an alkalophilic Bacillus species: purification and characterization

Cyclodextrin glycosyltransferase (E.C. 2.4.1.19) of alkalophilic Bacillus sp. 7-12 was purified by ammonium sulfate precipitation, DEAE-cellulose column chromatography and Sepharose CL-6B column chromatography. The enzyme thus obtained consisted of a single band that did not dissociate into subunits by SDS-polyacrylamide gel electrophoresis (PAGE). The molecular weight of the purified enzyme was determined to be 69,000 Da by SDS-PAGE. The enzyme was stable below 70 °C with an optimum activity at 60 °C, and was stable at a pH range of 6-10 with an optimum pH at 8.5. The enzyme activity was strongly inhibited by MgCl2, ZnCl2, CuSO4, Al2(SO4)3, CoCl2, AgNO3, FeSO4 and slightly inhibited by SnCl2 and MnCl2. CaCl2, KCl, EDTA and DTT had no influence on the enzyme activity. For cyclodextrin production, up to 34% conversion to cyclodextrins was obtained from 10% starch. The enzyme produced α-, β- and γ-cyclodextrins in the ratio of 0.26:1:0.86.