Immobilization of glycerol dehydrogenase on magnetic silica nanoparticles for conversion of glycerol to value-added 1,3-dihydroxyacetone

Abstract

Glycerol dehydrogenase (GlyDH) which oxidizes glycerol to the value-added chemical, 1,3-dihydroxyacetone, is of interest due to the oversupply of glycerol as a by-product of the biodiesel industry. To exploit the enzymatic oxidation of glycerol industrially, silica coated magnetic Fe3O4 nanoparticles were prepared and then activated with an amino-silane reagent for covalent immobilization of GlyDH via a glutaraldehyde linkage. At the optimal glutaraldehyde concentration of 0.05% (v/v), an enzyme loading of up to 57.5 mg/g-nanoparticles was achieved with 81.1% of the original activity retained. Reaction kinetic analysis indicated that the immobilized GlyDH had almost the same Michaelis-Menten constants for both NAD+ and glycerol as the free GlyDH did. However, after immobilization the turnover number kcat of the GlyDH decreased from 164 s−1 to 113 s−1, and the reaction was 1.3-fold less sensitive to inhibition by DHA, which could compensate the decrease in kcat. The immobilized GlyDH was also less sensitive to changes in pH and temperature, and showed a 5.3-fold improvement in thermal stability at 50°C. Furthermore, excellent reusability was observed such that 10 cycles of re-use only led to 9% loss of enzyme activity.