title: Purification and characterization of alpha-glucosidase from Torulaspora pretoriensis YK-1.
abstract: alpha-Glucosidase was partially purified 103-fold from a cell-free extract of Torulaspora pretoriensis YK-1 by column chromatography on Toyopearl HW55F, DEAE-Toyopearl 650M, hydroxylapatite and phenyl-Toyopearl 650M. Further purification by preparative polyacrylamide gel electrophoresis (PAGE) gave the homogenous protein, but the specific activity was reduced. The molecular weight of the enzyme was estimated to be 69,000 by SDS-PAGE and 60,000 by gel filtration. Optimum pH and temperature were 6.8 and 35 degrees C, respectively. The enzyme was inhibited strongly by AgNO3, HgCl2, sodium dodecyl sulfate, and N-ethylmaleimide. The Km (mM) for p-nitrophenyl alpha-D-glucopyranoside, maltose, maltotriose, isomaltose, methyl alpha-glucoside, and sucrose were 0.15, 150, 45, 17, 18, and 29, and Vmax (mumol/min/mg protein) for those substrates were 87, 0.23, 2.4, 9.0, 12, and 7.4, respectively. The N-terminal amino acid sequence of the enzyme was PEVKNHPETQPKWWKEATVY. The properties of alpha-glucosidase from T. pretoriensis YK-1 were similar to those from Saccharomyces cerevisiae.
authors.affiliation: Department of Food Science and Technology, Fukuyama University, Hiroshima, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Oda
authors.forename: H
authors.initials: H
authors.lastname: Iwamoto
authors.forename: K
authors.initials: K
authors.lastname: Hiromi
authors.forename: K
authors.initials: K
authors.lastname: Tonomura
mesh_terms: D000595:Amino Acid Sequence; D002845:Chromatography; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D013696:Temperature; D015003:Yeasts / Q000201:enzymology*; D000520:alpha-Glucosidases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism
chemical_list: D000520:alpha-Glucosidases