Fig. 3: Enzymatic characterization of GOx/CuSAE.

a, b Measuring NOx-like activity of CuSAE based on the conversion of NADPH. The experiment was performed by incubation of 200 μg/mL NADPH with different concentrations of CuSAE (0–100 μg/mL) (a) or different pH (7.4, 6.0, and 4.5) (b). After incubation for 30 min, the concentration of the remaining NADPH was measured based on its absorption at 340 nm. c EPR spectra detecting O₂^•- by using 1,3-diphenylisobenzofuran as a trapping agent. The experiment was performed on 100 μg/mL CuSAE and 200 μg/mL NADPH. d, e POD-like activity of CuSAE measured with TMB chromogenic assay at different pH (d) or different concentration of CuSAE (e). f EPR spectra detect •OH radical by using 1,3-diphenylisobenzofuran as a trapping agent. The assay was performed on 10 mM H₂O₂ and 60 μg/mL CuSAE. g GSHOx-like activity of CuSAE measured with DTNB assay. The assay was performed on GSH (0.15 mM) and different concentrations of CuSAE (0–90 μg/mL) in the presence of DTNB (2.5 mM). The peak at 412 nm indicates the reduced TNB from GSH reduction. h pH variation in the GOx/CuSAE-catalyzed oxidation of glucose. The reaction was performed by incubation of GOx/CuSAE (80 μg/mL) with different concentrations of glucose for 12 h. i Catalytic activities of free GOx and GOx/CuSAE after storage. GOx (80 μg/mL) and GOx/CuSAE (with 80 μg/mL GOx) were incubated in weakly acidic conditions (pH 6.0) at 37 °C for different times. After the incubation, their catalytic activity was analyzed by incubation with glucose (10 mM) for 2 h, and the generation of H₂O₂ production was measured titanium sulfate assay. Data are shown as mean ± s.d. from three independent measurements. For statistical analyses, One-way ANOVA Dunnett correction (a, d, e, and h), or Two-sided Student’s t-test (i) was used on the results from three independent experiments (n  =  3). Source data are provided as a Source Data file.