Cloning and characterization of two alpha-glucosidases from Bifidobacterium adolescentis DSM20083.
Two alpha-glucosidase encoding genes (aglA and aglB) from Bifidobacterium adolescentis DSM 20083 were isolated and characterized. Both alpha-glucosidases belong to family 13 of the glycosyl hydrolases. Recombinant AglA (EC 3.2.1.10) and AglB (EC 3.2.1.20), expressed in Escherichia coli, showed high hydrolytic activity towards isomaltose and pnp-alpha-glucoside. The K(m) for pnp-alpha-glucoside was 1.05 and 0.47 mM and the V(max) was 228 and 113 U mg(-1) for AglA and AglB, respectively. Using pnp-alpha-glucoside as substrate, the pH optimum for AglA was 6.6 and the temperature optimum was 37 degrees C. For AglB, values of pH 6.8 and 47 degrees C were found. AglA also showed high hydrolytic activity towards isomaltotriose and, to a lesser extent, towards trehalose. AglB has a high preference for maltose and less activity towards sucrose; minor activity was observed towards melizitose, low molecular weight dextrin, maltitol, and maltotriose. The recombinant alpha-glucosidases were tested for their transglucosylation activity. AglA was able to synthesize oligosaccharides from trehalose and sucrose. AglB formed oligosaccharides from sucrose, maltose, and melizitose.
van den Broek
L A M
LA
Laboratory of Food Chemistry, Wageningen University, P.O. Box 8129, 6700 EV Wageningen, The Netherlands.
Struijs
K
K
Verdoes
J C
JC
Beldman
G
G
Voragen
A G J
AG
D000595:Amino Acid Sequence; D001644:Bifidobacterium / Q000145:classification / Q000201:enzymology*; D003001:Cloning, Molecular; D005960:Glucosides / Q000378:metabolism; D006031:Glycosylation; D006923:Hymecromone / Q000031:analogs & derivatives* / Q000378:metabolism; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000032:analysis; D016415:Sequence Alignment; D013379:Substrate Specificity; D000516:alpha-Amylases / Q000032:analysis / Q000235:genetics; D000520:alpha-Glucosidases / Q000032:analysis* / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification
D005960:Glucosides; D011994:Recombinant Proteins; C014110:4-methylumbelliferyl glucoside; D006923:Hymecromone; D000516:alpha-Amylases; C049150:maltose-6'-phosphate glucosidase; D000520:alpha-Glucosidases