title: Cloning and heterologous expression of a beta-fructofuranosidase gene from Arthrobacter globiformis IFO 3062, and site-directed mutagenesis of the essential aspartic acid and glutamic acid of the active site.
abstract: We have cloned the gene encoding a beta-fructofuranosidase from Arthrobacter globiformis IFO 3062, and subsequently, the gene was heterologously expressed in Escherichia coli. This beta-fructofuranosidase gene encodes a protein of 548 amino acid residues with a calculated molecular mass of 60,519 Da. We have examined the roles of three residues of A. globiformis IFO 3062 beta-fructofuranosidase by site-directed mutagenesis, and found that aspartic acid 130 and glutamic acid 392, which are two of the apparent catalytic residues, are essential for hydrolase activity. This study provides the first experimental evidence showing that these two amino acid residues of beta-fructofuranosidase play a critical role in hydrolyzing sucrose.
authors.affiliation: Department of Applied Bioscience, Graduate School of Agriculture, Hokkaido University, Sapporo 060-8589, Japan.
authors.forename: Naoto
authors.initials: N
authors.lastname: Isono
authors.forename: Takashi
authors.initials: T
authors.lastname: Tochihara
authors.forename: Yuyus
authors.initials: Y
authors.lastname: Kusnadi
authors.forename: Than Than
authors.initials: TT
authors.lastname: Win
authors.forename: Kenji
authors.initials: K
authors.lastname: Watanabe
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Obae
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Ito
authors.forename: Hirokazu
authors.initials: H
authors.lastname: Matsui