The exploration of low-cost and metal-free nanozymes with oxidase-mimicking activity is highly desired due to their attractive properties and potential applications. However, it is still challenging and remains unexploited to fully realize oxidase-like nanozyme in the emerging covalent organic frameworks (COFs) due to their polymeric nature and weak photoelectric activity. We herein report the first example of the preparation and oxidase-mimicking activity of novel ultrathin two-dimensional (2D) COF (termed as TTPA-COF) nanosheets. The ultrathin TTPA-COF nanosheets with hexagonal layered structure are constructed from two flexible photoactive (diarylamino)benzene-based linkers, and exhibit remarkable catalytic activity toward the oxidation of 3,3',5,5'-tetramethylbenzidine (TMB) in the presence of O₂ due to their large specific surface areas and abundant active sites. Moreover, it is worth noting that the nanozyme activity could be regulated by external light irradiation. Based on the oxidase-mimicking activity of TTPA-COF nanosheets, a green colorimetric sensor is proposed for the sensitive and selective determination of glutathione (GSH) in a wide linear range of 0.5–40 µM with a detection limit of 0.5 µM. This work reported here would open new avenues for the exploration of low-cost and high-efficiency nanozymes, as well as extend the application of 2D COF nanosheets in the fields of catalysis and sensing.