In the present investigation, an ecofriendly magnetic inorganic-protein hybrid system-based enzyme immobilization was developed using partially purified laccase from Trametes versicolor (TvLac), Fe3O4 nanoparticles, and manganese (Mn), and was successfully applied for synthetic dye decolorization in the presence of enzyme inhibitors. After the partial purification of crude TvLac, the specific enzyme activity reached 212 U∙mg total protein-1. The synthesized Fe3O4/Mn3(PO4)2-laccase (Fe3O4/Mn-TvLac) and Mn3(PO4)2-laccase (Mn-TvLac) nanoflowers (NFs) exhibited encapsulation yields of 85.5% and 90.3%, respectively, with relative activities of 245% and 260%, respectively, compared with those of free TvLac. One-pot synthesized Fe3O4/Mn-TvLac exhibited significant improvements in catalytic properties and stability compared to those of the free enzyme. Fe3O4/Mn-TvLac retained a significantly higher residual activity of 96.8% over that of Mn-TvLac (47.1%) after 10 reuse cycles. The NFs showed potential for the efficient decolorization of synthetic dyes in the presence of enzyme inhibitors. For up to five reuse cycles, Fe3O4/Mn-TvLac retained a decolorization potential of 81.1% and 86.3% for Coomassie Brilliant Blue R-250 and xylene cyanol, respectively. The synthesized Fe3O4/Mn-TvLac showed a lower acute toxicity towards Vibrio fischeri than pure Fe3O4 nanoparticles did. This is the first report of the one-pot synthesis of biofriendly magnetic protein-inorganic hybrids using partially purified TvLac and Mn.