A cholesterol oxidase (COD) was hybridized with Ca , Zn , Al , Fe and Mn . After precipitation with PO at 4 °C for 72 h, the resulting pellets were freeze-dried. In scanning electron microscopy assays, the metal-COD complexes revealed flower-like or granular structures after hybridization. Fourier transform infrared spectroscopy assay revealed the characteristic peaks of both the enzyme and metal materials. X-ray diffraction analysis indicated that COD was encapsulated in CaHPO ·2H O-, Zn (PO ) ·4H O-, AlPO -, FeP - and Mn (PO ) ·3H O-based nanostructures, respectively. Differential scanning calorimetry assay indicated significant increases in thermo-denaturation temperatures from 60.5 °C to 167.02 °C, 167.02 °C, 137.70 °C, 172.85 °C and 160.99 °C, respectively. Using steroid derivatives as substrates, this enzyme could convert cholesterol, pregnenolone, dehydroepiandrosterone, ergosterol, -sitosterol and stigmasterol to related single products. Hybridization in metal-based nanostructures could significantly enhance the initial conversion ratio and reaction stability of the enzyme. In addition, substrate selectivity could be affected by various metal materials. Briefly, using Ca , Zn , Al , Fe and Mn as hybrid raw materials could help to encapsulate COD in related metal-enzyme nanostructures, and could help to promote the stability and tolerant properties of the enzyme, while also enhancing its catalytic characteristics.