title: Response surface methodology: Synthesis of short chain fructooligosaccharides with a fructosyltransferase from Aspergillus aculeatus.
abstract: A transferase was isolated, purified and characterised from Aspergillus aculeatus. The enzyme exhibited a pH and temperature optima of 6.0 and 60 degrees C, respectively and under such conditions remained stable with no decrease in activity after 5h. The enzyme was purified 7.1 fold with a yield of 22.3% and specific activity of 486.1Umg(-1) after dialysis, concentration with polyethyleneglycol (30%) and DEAE-Sephacel chromatography. It was monomeric with a molecular mass of 85kDa and K(m) and V(max) values of 272.3mM and 166.7micromolmin(-1)ml(-1). The influence of pH, temperature, reaction time, and enzyme and sucrose concentration on the formation of short-chain fructooligosaccharides (FOS) was examined by statistical response surface methodology (RSM). The enzyme showed both transfructosylation and hydrolytic activity with the transfructosylation ratio increasing to 88% at a sucrose concentration of 600mgml(-1). Sucrose concentration (400mgml(-1)) temperature (60 degrees C), and pH (5.6) favoured the synthesis of high levels of GF(3) and GF(4). Incubation time had a critical effect on the yield of FOS as the major products were GF(2) after 4h and GF(4) after 8h. A prolonged incubation of 16h resulted in the conversion of GF(4) into GF(2) as a result of self hydrolase activity.
authors.affiliation: Department of Biochemistry, Microbiology and Biotechnology, Rhodes University, P.O. Box 94, Grahamstown 6140, South Africa.
authors.forename: A
authors.initials: A
authors.lastname: Nemukula
authors.forename: T
authors.initials: T
authors.lastname: Mutanda
authors.forename: B S
authors.initials: BS
authors.lastname: Wilhelmi
authors.forename: C G
authors.initials: CG
authors.lastname: Whiteley
mesh_terms: D001230:Aspergillus / Q000201:enzymology*; D004795:Enzyme Stability; D006602:Hexosyltransferases / Q000302:isolation & purification / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008970:Molecular Weight; D009844:Oligosaccharides / Q000096:biosynthesis*; D013696:Temperature; D001426:Bacterial Proteins
chemical_list: D006602:Hexosyltransferases; D009844:Oligosaccharides; C116580:fructooligosaccharide; C021451:inulosucrase; D001426:Bacterial Proteins