The exploration of enzyme-mimic nanocomposites to replace commercial enzymes is challenging in the construction of biosensors with high performance. Herein, an ultrafast response nonenzymatic glucose sensor was developed based on a Cu2O–TiO2/Ti3C2Tx nanocomposite, which was prepared via an in situ redox procedure using Ti3C2Tx nanosheets both as a reducing agent and a carrier substrate. Structural characterizations, including X-ray diffraction (XRD) and transmission electron microscopy (TEM), revealed that Cu2O nanoparticles with a size of 10–30 nm are uniformly distributed on the surface of the Ti3C2Tx nanosheets and form close contacts through Ti–O–Cu bonds, resulting in favorable stability of the system. The unique heterostructure of Cu2O–TiO2/Ti3C2Tx can produce abundant active sites and facilitate electron transfer, demonstrating enhanced catalytic activity in the electrooxidation of glucose. Consequently, the electrochemical sensor fabricated by modifying a glassy carbon electrode (GCE) with Cu2O–TiO2/Ti3C2Tx nanocomposites exhibited excellent analytical properties for glucose detection, including a fast response time of 0.1 s, a low detection limit of 0.67 μM, and a high sensitivity of 267 μA mM–1 cm–2. In addition, the developed method possesses good anti-interference ability and superior stability and thus has been successfully applied for real sample monitoring. This study offers a way to construct Ti3C2Tx-based heterostructures with tunable physiochemical properties and expands their use in the electrochemical field.