title: Molecular cloning of an inulin fructotransferase (depolymerizing) gene from Arthrobacter sp. H65-7 and its expression in Escherichia coli.
abstract: The gene encoding an extracellular inulin fructotransferase (depolymerizing) (inulase II) (EC 2.4.1.93), designated ift gene, was cloned from the genomic DNA of Arthrobacter sp. H65-7, and expressed in Escherichia coli for the first time. Sequence analysis showed a single open reading frame consisting of 1314 base pairs that encoded a signal peptide of 32 amino acids and a mature protein of 405 amino acids. The primary structure showed a homology of 49.8% with that of the inulin fructotransferase (DFA I-producing) (EC 2.4.1.200) from Arthrobacter globiformis S14-3. E. coli cells carrying the ift gene produced the active enzyme under control of the lac promoter. The expression of the ift gene was improved by a plasmid, pIFT-B, in which the ift gene was immediately downstream from the lac promoter. An E. coli transformant carrying pIFT-B expressed twice as much activity of inulase II as that of the original strain, Arthrobacter sp. H65-7. Most of the enzyme activity existed within the cells.
authors.affiliation: Department of Bioscience and Chemistry, Faculty of Agriculture, Hokkaido University, Sapporo, Japan.
authors.forename: H
authors.initials: H
authors.lastname: Sakurai
authors.forename: A
authors.initials: A
authors.lastname: Yokota
authors.forename: F
authors.initials: F
authors.lastname: Tomita
mesh_terms: D000595:Amino Acid Sequence; D001173:Arthrobacter / Q000201:enzymology* / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D015964:Gene Expression Regulation, Bacterial; D006602:Hexosyltransferases / Q000096:biosynthesis* / Q000235:genetics*; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
chemical_list: D011994:Recombinant Proteins; D006602:Hexosyltransferases; C021450:inulin fructotransferase (depolymerizing)