Characterization of Bacillus stearothermophilus cyclodextrin glucanotransferase in ascorbic acid 2-O-alpha-glucoside formation.
In this study, we characterized cyclodextrin glucanotransferase (CGTase) from Bacillus stearothermophilus in L-ascorbic acid-2-O-alpha-D-glucoside (AA-2G) formation and compared its enzymological properties with those of rat intestinal and rice seed alpha-glucosidases which had the ability to form AA-2G. CGTase formed AA-2G efficiently using alpha-cyclodextrin (alpha-CD) as a substrate and ascorbic acid (AA) as an acceptor. Several AA-2-oligoglucosides were also formed in this reaction mixture, and they could be converted to AA-2G by the additional treatment of glucoamylase. The optimum temperature for AA-2G formation was 70 degrees C and its optimum pH was around 5.0. CGTase also utilized beta- and gamma-CDs, maltooligosaccharides, dextrin, amylose, glycogen and starch as substrates, but not any disaccharides except maltose. CGTase showed the same acceptor specificity as two alpha-glucosidases, whereas its hydrolyzing activity towards AA-2G was very low compared with those of alpha-glucosidases. Cleavage profiles of AA-2-oligoglucosides by CGTase present a possible mechanism for AA-2G formation that CGTase transfers a glucose-hexamer to an acceptor at the first step and then a glucose is stepwisely removed from the non-reducing end of the product through glucoamylase-like action of this enzyme. These results indicate that CGTase is able to synthesize AA-2G more efficiently than rat and rice alpha-glucosidases and utilization of this enzyme makes the mass production of AA-2G possible.
Tanaka
M
M
Department of Immunochemistry, Faculty of Pharmaceutical Sciences, Okayama University, Japan.
Muto
N
N
Yamamoto
I
I
D000818:Animals; D001205:Ascorbic Acid / Q000031:analogs & derivatives* / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D001411:Geobacillus stearothermophilus / Q000201:enzymology*; D005087:Glucan 1,4-alpha-Glucosidase / Q000378:metabolism; D005947:Glucose / Q000378:metabolism; D005964:Glucosyltransferases / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D012275:Oryza / Q000201:enzymology; D051381:Rats; D013379:Substrate Specificity; D013696:Temperature; D000520:alpha-Glucosidases / Q000378:metabolism
C065378:ascorbic acid 2-O-glucoside; D005964:Glucosyltransferases; C019963:cyclomaltodextrin glucanotransferase; D000520:alpha-Glucosidases; D005087:Glucan 1,4-alpha-Glucosidase; D005947:Glucose; D001205:Ascorbic Acid