Lactose hydrolysis and formation of galactooligosaccharides by a novel immobilized beta-galactosidase from the thermophilic fungus Talaromyces thermophilus.
beta-Galactosidase from the fungus Talaromyces thermophilus CBS 236.58 was immobilized by covalent attachment onto the insoluble carrier Eupergit C with a high binding efficiency of 95%. Immobilization increased both activity and stability at higher pH values and temperature when compared with the free enzyme. Especially the effect of immobilization on thermostability is notable. This is expressed by the half-lifetime of the activity at 50 degrees C, which was determined to be 8 and 27 h for the free and immobilized enzymes, respectively. Although immobilization did not significantly change kinetic parameters for the substrate lactose, a considerable decrease in the maximum reaction velocity V(max) was observed for the artificial substrate o-nitrophenyl-beta-D-galactopyranoside (oNPG). The hydrolysis of both oNPG and lactose is competitively inhibited by the end products glucose and galactose. However, this inhibition is only very moderate as judged from kinetic analysis with glucose exerting a more pronounced inhibitory effect. It was evident from bioconversion experiments with 20% lactose as substrate, that the immobilized enzyme showed a strong transgalactosylation reaction, resulting in the formation of galactooligosaccharides (GalOS). The maximum yield of GalOS of 34% was obtained when the degree of lactose conversion was roughly 80%. Hence, this immobilized enzyme can be useful both for the cleavage of lactose at elevated temperatures, and the formation of GalOS, prebiotic sugars that have a number of interesting properties for food applications.
Nakkharat
Phimchanok
P
Division of Food Biotechnology, Department of Food Sciences and Technology, BOKU-University of Natural Resources and Applied Life Sciences, Vienna Muthgasse 18, A-1190, Vienna, Austria.
Haltrich
Dietmar
D
D004789:Enzyme Activation; D004800:Enzymes, Immobilized / Q000737:chemistry; D005690:Galactose / Q000138:chemical synthesis*; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007785:Lactose / Q000737:chemistry*; D009844:Oligosaccharides / Q000138:chemical synthesis*; D013379:Substrate Specificity; D032901:Talaromyces / Q000201:enzymology*; D013696:Temperature; D001616:beta-Galactosidase / Q000737:chemistry* / Q000378:metabolism*
D004800:Enzymes, Immobilized; D009844:Oligosaccharides; D001616:beta-Galactosidase; D007785:Lactose; D005690:Galactose