Radiation therapy is applied in the treatment of head and neck cancer patients. However, oral-health-related side effects like hyposalivation and ahigher prevalence of caries have been shown. This study aims to assess the influence of different radiotherapy doses on the mechanical properties, roughness, superficial microstructure, and crystallinity of the enamel and dentin of human premolar teeth. Specimens (n = 25) were categorized into five groups based on the radiation dose received (0, 10, 30, 50, and 70 Gy). The enamel and dentin of these specimens were subjected to amicrohardness tester, profilometer, atomic force microscopy (AFM), scanning electron microscopy (SEM), and X‑ray diffraction (XRD) before and after different irradiation doses and compared to hydroxylapatite in each group. The data were analyzed using repeated-measures analysis of variance (ANOVA). Therapeutic radiation doses of 30, 50, and 70 Gy led to adecrease in the microhardness and an increase in the average roughness of the enamel, and rougher surfaces were observed in the mixed three-dimensional images. Moreover, in the dentin, asimilar outcome could be observed for more than 10 Gy. The main crystalline phase structure remained hydroxylapatite, but the crystallinity decreased and the crystalline size increased above 10 Gy. The superficial micromorphology revealed granulation, fissures, and cracks in adose-dependent manner. Radiation below 70 Gy had little effect on the hydroxylapatite concentration during the whole experiment. Above aradiation dose of 30 Gy, the micromorphology of the tooth enamel changed. This occurred for dentin above 10 Gy, which indicates that dentin is more sensitive to radiotherapy than enamel. The radiation dose had an effect on the micromorphology of the hard tissues of the teeth. These results illustrate the possible mechanism of radiation-related caries and have guiding significance for clinical radiotherapy.