It is reported that fluorescent microtubules may break under irradiation. However, the process and mechanism are not well understood. In this paper, we study the process of light-induced microtubule breaking by optical tweezers and find the breakup of protofilaments in microtubule is not simultaneous but proceeds step-by-step. After a series of biological experiments, we confirm that free radicals are the main reason for the photosensitive breaking of microtubule. According to the results of optical tweezers experiments, the mechanism of free radicals acting on microtubule is infered.