Characterization of a neopullulanase and an alpha-glucosidase from Bacteroides thetaiotaomicron 95-1.
Previously, we constructed a gene disruption in the pullulanase I gene of Bacteroides thetaiotaomicron 5482A. This mutant, designated B. thetaiotaomicron 95-1, had a lower level of pullulanase specific activity than did wild-type B. thetaiotaomicron but still exhibited a substantial amount of pullulanase activity. Characterization of the remaining pullulanase activity present in B. thetaiotaomicron 95-1 has identified an alpha(1----4)-D-glucosidic bond cleaving pullulanase which has been tentatively designated a neopullulanase. The neopullulanase (pullulanase II) is a 70-kDa soluble protein which cleaves alpha(1----4)-D-glucosidic bonds in pullulan to produce panose. The neopullulanase also cleaved alpha(1----4) bonds in amylose and in oligosaccharides of maltotriose through maltoheptaose in chain length. An alpha-glucosidase from B. thetaiotaomicron 95-1 was characterized. The alpha-glucosidase was partially purified to a preparation containing three proteins of 80, 57, and 50 kDa. Pullulan and amylose were not hydrolyzed by the alpha-glucosidase. alpha(1----4)-D-Glucosidic oligosaccharides from maltose to maltoheptaose were hydrolyzed to glucose by the alpha-glucosidase. The alpha-glucosidase also hydrolyzed alpha(1----6)-linked oligosaccharides such as panose (the product of the pullulanase II action on pullulan) and isomaltotriose.
Smith
K A
KA
Department of Microbiology, University of Illinois, Urbana 61801.
Salyers
A A
AA
D000681:Amylases / Q000032:analysis; D000688:Amylose / Q000378:metabolism; D001439:Bacteroides / Q000201:enzymology*; D002845:Chromatography; D004591:Electrophoresis, Polyacrylamide Gel; D005936:Glucans / Q000378:metabolism; D005960:Glucosides / Q000378:metabolism; D006026:Glycoside Hydrolases / Q000032:analysis* / Q000302:isolation & purification; D008970:Molecular Weight; D014461:Ultracentrifugation; D000520:alpha-Glucosidases / Q000032:analysis* / Q000302:isolation & purification
D005936:Glucans; D005960:Glucosides; C019502:4-nitrophenyl alpha-glucoside; C009109:pullulan; D000688:Amylose; D000681:Amylases; D006026:Glycoside Hydrolases; C058484:neopullulanase; D000520:alpha-Glucosidases