Thermoplastic polyurethane (TPU) with different chemical structures is prepared by the prepolymer method and then mixed with base asphalt to obtain thermoplastic polyurethane modified asphalt (TPU-A). The structure of synthetic TPU was characterized by Fourier transform infrared spectroscopy (FTIR) and thermomechanical properties tests. The road performance of TPU-A is characterized by conventional performance tests, dynamic shear rheometer (DSR) and bending beam rheometer (BBR). The modification mechanism of TPU-A is studied by FTIR, fluorescence microscopy (FM) and electron microscopy (SEM). The results show that hard segment increases the polar groups of TPU and gives it higher thermal stability. The hard segment-rich TPU is disperse evenly as particles in the asphalt phase, which increases the thermal stability and shear deformation capacity of TPU-A. The amount of hard segment in TPU has a critical point (30 wt%) to improve the low and high temperature performance of TPU-A. The soft segment can be inflated by the asphalt to form a physical cross-linking network structure, which advances the low-temperature properties.