Evaluation of coupling protocols to bind beta-glucosidase on magnetic nanoparticles.
Beta-Glucosidase has been chosen as a model biomolecule to establish a general protocol for binding enzymes on both ferromagnetic and superparamagnetic nano-particles for sensing applications. Using EDC (1-(3-dimethyl-aminopropyl)-3-ethylcarbodiimide) or SMCC (Succinimidyl-4-(N-maleimidomethyl)cyclohexane-1-carboxylate) as coupling agents, we compared two different methods for the fabrication of enzyme-decorated magnetic nanoparticles. We identified the best conditions for the preparation of a responsive bioactive magnetic system comparing different covalent bio-grafting protocols. The enzymatic test has been performed using beta-Glucosidase. The systems were characterized using scanning electron microscopy, infrared spectroscopy, and the enzyme loading was measured by a glucose assay in the presence of the enzyme-decorated magnetic particles. Although the faster response of ferromagnetic particles to the magnetic field, the assay results suggested that the superparamagnetic particles are more efficient carriers. In fact, the best enzymatic activity was measured on superparamagnetic systems that have the further advantage of preventing aggregation induced by the residual magnetization. Hence, beta-Glucosidase coated magnetic nanospheres could provide an attractive system suitable for the cleavage and the rapid evaluation of glycoside levels in natural products, measuring the liberated glucose without the need for specialised instrumentation. Moreover, the magnetic particles allow the subsequent collection of enzymes for further analysis, such as its use in portable fast screening kits or devices.
Raffaele
R
Ricco
Cara M
CM
Doherty
Paolo
P
Falcaro
D015374:Biosensing Techniques / Q000295:instrumentation* / Q000379:methods; D004800:Enzymes, Immobilized / Q000737:chemistry* / Q000378:metabolism; D005947:Glucose / Q000032:analysis / Q000378:metabolism; D058185:Magnetite Nanoparticles / Q000737:chemistry*; D036103:Nanotechnology / Q000379:methods*; D001617:beta-Glucosidase / Q000737:chemistry* / Q000378:metabolism
D004800:Enzymes, Immobilized; D058185:Magnetite Nanoparticles; D001617:beta-Glucosidase; D005947:Glucose