title: Purification and kinetic characterization of a fructosyltransferase from Aspergillus aculeatus.
abstract: A fructosyltransferase present in Pectinex Ultra SP-L, a commercial enzyme preparation from Aspergillus aculeatus, was purified to 107-fold and further characterised. The enzyme was a dimeric glycoprotein (20% (w/w) carbohydrate content) with a molecular mass of around 135 kDa for the dimer. Optimal activity/stability was found in the pH range 5.0-7.0 and at 60 degrees C. It was stable or slightly activated (upto 1.4-fold) in the presence of reducing agents, such as dithiothreitol and 2-mercaptoethanol, and detergents, such as sodium dodecylsulphate and Tween 80. The enzyme was able to transfer fructosyl groups from sucrose as donor producing the corresponding series of fructooligosaccharides: 1-kestose, nystose and fructosylnystose. Using sucrose as substrate, the k(cat) and K(m) values for transfructosylating activity were 1.62+/-0.09 x 10(4)s(-1) and 0.53+/-0.05 M, whereas for hydrolytic activity the corresponding values were 775+/-25s(-1) and 27+/-3 mM. At elevated sucrose concentrations, the fructosyltransferase from A. aculeatus showed a high transferase/hydrolase ratio that confers it a great potential for the industrial production of prebiotic fructooligosaccharides.
authors.affiliation: Departamento de Biocatalisis, Instituto de Catalisis y Petroleoquimica, CSIC, Cantoblanco, 28049 Madrid, Spain.
authors.forename: Iraj
authors.initials: I
authors.lastname: Ghazi
authors.forename: Lucia
authors.initials: L
authors.lastname: Fernandez-Arrojo
authors.forename: Humberto
authors.initials: H
authors.lastname: Garcia-Arellano
authors.forename: Manuel
authors.initials: M
authors.lastname: Ferrer
authors.forename: Antonio
authors.initials: A
authors.lastname: Ballesteros
authors.forename: Francisco J
authors.initials: FJ
authors.lastname: Plou
mesh_terms: D001230:Aspergillus / Q000201:enzymology*; D005503:Food Additives / Q000737:chemistry; D006602:Hexosyltransferases / Q000302:isolation & purification* / Q000378:metabolism*; D013395:Sucrose / Q000378:metabolism; D001426:Bacterial Proteins
chemical_list: D005503:Food Additives; D006602:Hexosyltransferases; D013395:Sucrose; C021451:inulosucrase; D001426:Bacterial Proteins