title: Purification and characterization of a Bacillus sp. SAM1606 thermostable alpha-glucosidase with transglucosylation activity.
abstract: We purified a novel alpha-glucosidase to homogeneity from an Escherichia coli recombinant transformed with the alpha-glucosidase gene from thermophilic Bacillus sp. SAM1606. The enzyme existed as mono- and multimeric forms of a promoter protein with a relative molecular weight of 64,000 and isoelectric point of 4.6. We isolated a monomeric form of the enzyme and characterized it. The enzyme was unique among the known alpha-glucosidases in both broad substrate specificity and high thermostability. The enzyme hydrolysed a variety of O-alpha-D-glucopyranosides such as nigerose, maltose, isomaltose, sucrose, and trehalose efficiently. The molecular activity (k0) and the Michaelis constant (Km) values at 55 degrees C and pH 6.0 for sucrose were 54.6 s-1 and 5.3 mM, respectively. The optimum pH and temperature for hydrolysis were pH 5.5 and 75 degrees C, respectively. The enzyme exhibited a high transglucosylation activity: it reacted with 1.8 M sucrose at 60 degrees C for 70 h to yield oligosaccharides containing theanderose in a maximum yield of 35% (w/w). High thermostability of the enzyme (stable up to 65 degrees C at pH 7.2 for 10 min) permits the transglucosylation reaction at high temperatures, which would be beneficial for continuous production of oligosaccharides from sucrose.
authors.affiliation: Suntory Ltd., Research Center, Osaka, Japan.
authors.forename: M
authors.initials: M
authors.lastname: Nakao
authors.forename: T
authors.initials: T
authors.lastname: Nakayama
authors.forename: M
authors.initials: M
authors.lastname: Harada
authors.forename: A
authors.initials: A
authors.lastname: Kakudo
authors.forename: H
authors.initials: H
authors.lastname: Ikemoto
authors.forename: S
authors.initials: S
authors.lastname: Kobayashi
authors.forename: Y
authors.initials: Y
authors.lastname: Shibano
mesh_terms: D000595:Amino Acid Sequence; D001407:Bacillus / Q000201:enzymology*; D002384:Catalysis; D003001:Cloning, Molecular; D004795:Enzyme Stability; D004926:Escherichia coli; D005959:Glucosidases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D008969:Molecular Sequence Data; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D005959:Glucosidases