Surface roughness of structures is a primary factor that affects the mechanical properties of soil-structure interface. To further study the effect of roughness on shear strength of interface, large-scale direct shear tests were performed on clay-concrete interface under different roughness conditions and the influence mechanism of roughness on peak shear strength of interface was revealed. The results showed that the shear stress-displacement curves of clay-concrete interface exhibited strain-softening under different roughness conditions, and the greater roughness, the more obvious peak point of curve. Increasing roughness could obviously increase the peak shear strength of interface and there existed a critical roughness in terms of its influence on peak shear strength of interface. Morphological characteristics of the shear failure plane of different rough interfaces indicated that the smooth interface mainly occurred shear slip failure during the shearing process, and the friction and occlusion between clay particles and concrete were strengthened with increasing roughness, which resulted in the internal shear failure of clay. The shear strength of interface can be approximately divided into two parts: the shear strength of smooth interface and the shear strength of soil in rough parts. A new peak shear strength model of interface considering roughness was established by introducing a roughness-related coefficient into Jewell’s model and proposing a function to describe the relationship between the coefficient and roughness. Finally, comparison results between calculated value and test value showed that maximum relative error was 11.01% and mean relative error was 4.74%, which verified the accuracy and rationality of the proposed model.