Immobilization of β-d-galactosidase from Kluyveromyces lactis on functionalized silicon dioxide nanoparticles: characterization and lactose hydrolysis.
β-D-Galactosidase (BGAL) from Kluyveromyces lactis was covalently immobilized to functionalized silicon dioxide nanoparticles (10-20 nm). The binding of the enzyme to the nanoparticles was confirmed by Fourier transform-infrared spectroscopy (FTIR) and scanning electron microscopy (SEM). Functionalized nanoparticles showed 87% immobilization yield. Soluble and immobilized enzyme preparation exhibited pH-optima at pH 6.5 and 7.0, respectively, with temperature optima at 35 and 40°C, respectively. Michaelis constant (K(m)) was 4.77 and 8.4mM for free and immobilized BGAL, respectively. V(max) for the soluble and immobilized enzyme was 12.25 and 13.51 U/ml, respectively. Nanoparticle immobilized BGAL demonstrated improved stability after favoring multipoint covalent attachment. Thermal stability of the immobilized enzyme was enhanced at 40, 50 and 65°C. Immobilized nanoparticle-enzyme conjugate retained more than 50% enzyme activity up to the eleventh cycle. Maximum lactose hydrolysis by immobilized BGAL was achieved at 8h.
Centre for Biotechnology, Chemistry and System Biology (BioDeakin), Institute for Technology and Research Innovation (ITRI), Deakin University, Geelong, Australia.
Madan Lal
ML
Verma
Colin James
CJ
Barrow
J F
JF
Kennedy
Munish
M
Puri
D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000737:chemistry* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D007716:Kluyveromyces / Q000201:enzymology*; D007785:Lactose / Q000737:chemistry* / Q000378:metabolism; D053758:Nanoparticles / Q000737:chemistry* / Q000648:ultrastructure; D012822:Silicon Dioxide / Q000737:chemistry*; D013696:Temperature; D013997:Time Factors; D001616:beta-Galactosidase / Q000737:chemistry* / Q000378:metabolism
D004800:Enzymes, Immobilized; D012822:Silicon Dioxide; D001616:beta-Galactosidase; D007785:Lactose