Preparation and characterization of copper-Brevibacterium cholesterol oxidase hybrid nanoflowers

Abstract

Brevibacterium cholesterol oxidase (COD)-Cu hybrid nanoflowers were prepared, optimized and characterized for structural and catalytic properties. Regarding scanning electron microscopy (SEM), Fourier transform–infrared spectroscopy (FTIR) and X-ray diffraction (XRD) assays, COD molecules were successfully encapsulated with Cu3(PO4)2·3H2O based hybrid nanoflowers. After immobilization in hybrid nanoflowers, the interaction between COD and flavo-cofactor (FAD) was enhanced; and regarding to differential scanning calorimetry (DSC) assay, the Tm value of immobilized COD was increased from 60.5 °C (free enzyme) to 138.49 °C (nanoflowers). Additionally, in activity assay, Cu-COD nanoflowers revealed improved resistance to temperature and pH. After 10 times of recycling, approximately 70% of initial activity of Cu-COD nanoflowers was maintained, while the free COD was inactivated after 3 times of recycling. Furthermore, using cholesterol as substrate, in n-octane/water biphasic reaction system, the stability of Cu-COD nanoflowers was significantly promoted, and the initial conversion ration could be over two times as that of free enzyme. In brief, the hybrid nanoflowers dramatically enhanced the structural and thermo stability, the tolerance to biphasic mixture, and the catalytic efficiency of COD; and Cu-COD nanoflowers should be of great potential in the bioconversion of sterol derivatives.