The cementum–enamel junction (CEJ) is a unique interface that plays a critical role in effectively transferring force from the top of the tooth to the jawbone. A comparative study of bovine and equine teeth has been carried out experimentally to establish the relationship among the interfacial structure, fracture resistance, and mechanical functions. The CEJs were analyzed by optical microscopy and scanning electron microscopy (SEM). Microhardness and elastic modulus were measured with a Vickers microhardness tester and a nanoindentation system. A modified cantilever beam technique was used to study CEJ fracture process and the toughness (Gc,). It was observed that CEJ is a distinct interface connecting the hard enamel (200–350 kg/mm2) with the soft cementum (∼20–30 kg/mm2). Bovine CEJ was macroscopically flat and smooth. However, under SEM, a close integration of cementum bumps and enamel pits at the 5 μm level and dimples at the 200 nm level were clearly seen. In equine CEJ, the similar flat cementum–enamel interface was interrupted by hemispherical cementum protrusions (average size of 28.3 μm) at high density (340/mm2). Despite the microscopic roughness, bovine CEJ was easy to fracture at relatively low toughness (128 J/m2). The measured toughness for equine CEJ was about seven times that of bovine (1140 J/m2). The high toughness has been attributed to the mechanical interlocking provided by the cementum protrusions. The underlying crack bridging mechanism and the applications to the design of orthopaedic implants are briefly discussed.