Abstract
Glucosyltransferase and glucanotransferase involved in the production of cyclic tetrasaccharide (CTS; cyclo {→6}-α-D-glucopyranosyl-(1→3)-α-D-glucopyranosyl-(1→6)-α-D-glucopyranosyl-(1→3)-α-D-glucopyranosyl- (1→)) from α-1,4-glucan were purified from Bacillus globisporus C11. The former was a 1,6-α-glucosyltransferase (6GT) catalyzing the α-1,6-transglucosylation of one glucosyl residue to the nonreducing end of maltooligosaccharides (MOS) to produce α-isomaltosyl-MOS from MOS. The latter was an isomaltosyl transferase (IMT) catalyzing α-1,3-, α-1,4-, and α,β-1,1-intermolecular transglycosylation of isomaltosyl residues. When IMT catalyzed α-1,3-transglycosylation, α-isomaltosyl-(1→3)-α-isomaltosyl-MOS was produced from α-isomaltosyl-MOS. In addition, IMT catalyzed cyclization, and produced CTS from α-isomaltosyl-(1→3)-α-isomaltosyl-MOS by intramolecular transglycosylation. Therefore, the mechanism of CTS synthesis from MOS by the two enzymes seemed to follow three steps:
1) MOS→α-isomaltosyl-MOS (by 6GT),
2) α-isomaltosyl-MOS→α-isomaltosyl-(1→3)-α- isomaltosyl-MOS (by IMT), and
3) α-isomaltosyl-(1→3)-α-isomaltosyl-MOS→CTS +MOS (by IMT).
The molecular mass of 6GT was estimated to be 137 kDa by SDS-PAGE. The optimum pH and temperature for 6GT were pH 6.0 and 45°C, respectively. This enzyme was stable at from pH 5.5 to 10 and on being heated to 40°C for 60 min. 6GT was strongly activated and stabilized by various divalent cations. The molecular mass of IMT was estimated to be 102 kDa by SDS-PAGE. The optimum pH and temperature for IMT were pH 6.0 and 50°C, respectively. This enzyme was stable at from pH 4.5 to 9.0 and on being heated to 40°C for 60 min. Divalent cations had no effect on the stability or activity of this enzyme.
