title: Immobilized Trienzymatic System with Enhanced Stabilization for the Biotransformation of Lactose.
abstract: The use of ketohexose isomerases is a powerful tool in lactose whey processing, but these enzymes can be very sensitive and expensive. Development of immobilized/stabilized biocatalysts could be a further option to improve the process. In this work, β-galactosidase fromBacillus circulans, l-arabinose (d-galactose) isomerase fromEnterococcus faecium, and d-xylose (d-glucose) isomerase fromStreptomyces rubiginosuswere immobilized individually onto Eupergit C and Eupergit C 250 L. Immobilized activity yields were over 90% in all cases. With the purpose of increasing thermostability of derivatives, two post-immobilization treatments were performed: alkaline incubation to favor the formation of additional covalent linkages, and blocking of excess oxirane groups by reacting with glycine. The greatest thermostability was achieved when alkaline incubation was carried out for 24 h, producing l-arabinose isomerase-Eupergit C derivatives with a half-life of 379 h and d-xylose isomerase-Eupergit C derivatives with a half-life of 554 h at 50 °C. Preliminary assays using immobilized and stabilized biocatalysts sequentially to biotransform lactose at pH 7.0 and 50 °C demonstrated improved performances as compared with soluble enzymes. Further improvements in ketohexose productivities were achieved when the three single-immobilizates were incubated simultaneously with lactose in a mono-reactor system.
authors.affiliation: Cátedra de Bioquímica, Departamento de Biociencias, Facultad de Química, Universidad de la República, Gral Flores 2124, 11800 Montevideo, Uruguay. ptorres@fq.edu.uy.
authors.forename: Pedro
authors.initials: P
authors.lastname: Torres
authors.affiliation: Cátedra de Bioquímica, Departamento de Biociencias, Facultad de Química, Universidad de la República, Gral Flores 2124, 11800 Montevideo, Uruguay. fbatista@fq.edu.uy.
authors.forename: Francisco
authors.initials: F
authors.lastname: Batista-Viera
mesh_terms: D019747:Aldose-Ketose Isomerases / Q000378:metabolism*; D001407:Bacillus / Q000201:enzymology; D001426:Bacterial Proteins / Q000378:metabolism; D001711:Biotransformation; D016984:Enterococcus faecium / Q000201:enzymology; D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000378:metabolism*; D007785:Lactose / Q000378:metabolism*; D011108:Polymers / Q000378:metabolism; D013302:Streptomyces / Q000201:enzymology; D001616:beta-Galactosidase / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D004800:Enzymes, Immobilized; D011108:Polymers; C065843:Eupergit C; D001616:beta-Galactosidase; D019747:Aldose-Ketose Isomerases; C019715:L-arabinose isomerase; C023298:xylose isomerase; D007785:Lactose
keywords: ">d-fructose; ">d-glucose isomerase; ">d-tagatose; ">l-arabinose isomerase; enzyme immobilization; β-galactosidase