title: Synthesis of galactooligosaccharides by CBD fusion β-galactosidase immobilized on cellulose.
abstract: The β-galactosidase gene (bgaL3) was cloned from Lactobacillus bulgaricus L3 and fused with cellulose binding domain (CBD) using pET-35b (+) vector in Escherichia coli. The resulting fusion protein (CBD-BgaL3) was directly adsorbed onto microcrystalline cellulose with a high immobilization efficiency of 61%. A gram of cellulose was found to absorb 97.6 U of enzyme in the solution containing 100mM NaCl (pH 5.8) at room temperature for 20 min. The enzymatic and transglycosylation characteristics of the immobilized CBD-BgaL3 were similar to the free form. Using the immobilized enzyme as the catalyst, the yield of galactooligosaccharides (GOS) reached a maximum of 49% (w/w) from 400 g/L lactose (pH 7.6) at 45 °C for 75 min, with a high productivity of 156.8 g/L/h. Reusability assay was subsequently performed under the same reaction conditions. The immobilized enzyme could retain over 85% activity after twenty batches with the GOS yields all above 40%.
authors.affiliation: State Key Lab of Microbial Technology and National Glycoengineering Research Center, Shandong University, Jinan 250100, PR China.
authors.forename: Lili
authors.initials: L
authors.lastname: Lu
authors.forename: Shuze
authors.initials: S
authors.lastname: Xu
authors.forename: Renfei
authors.initials: R
authors.lastname: Zhao
authors.forename: Dayu
authors.initials: D
authors.lastname: Zhang
authors.forename: Zhengyi
authors.initials: Z
authors.lastname: Li
authors.forename: Yumei
authors.initials: Y
authors.lastname: Li
authors.forename: Min
authors.initials: M
authors.lastname: Xiao
mesh_terms: D000327:Adsorption / Q000187:drug effects; D061249:Batch Cell Culture Techniques; D002482:Cellulose / Q000494:pharmacology*; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability / Q000187:drug effects; D004800:Enzymes, Immobilized / Q000737:chemistry / Q000378:metabolism*; D005690:Galactose / Q000096:biosynthesis*; D006863:Hydrogen-Ion Concentration / Q000187:drug effects; D007700:Kinetics; D007778:Lactobacillus / Q000187:drug effects / Q000201:enzymology; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000096:biosynthesis*; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D059027:Recycling; D012965:Sodium Chloride / Q000494:pharmacology; D013696:Temperature; D013997:Time Factors; D001616:beta-Galactosidase / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D004800:Enzymes, Immobilized; D009844:Oligosaccharides; D011993:Recombinant Fusion Proteins; D012965:Sodium Chloride; D002482:Cellulose; D001616:beta-Galactosidase; C109691:microcrystalline cellulose; D005690:Galactose