Characterizing the thermal maturation of organic matter is essential to oil and gas resource evaluation and generation mechanisms, but is extremely challenging in nanoscale vitrinite-dominated formations (few suitable vitrinites in micron scales for traditionally microscope petrographic analyses), forming a critical issue in oil and gas geochemistry requiring new solutions. Here we considered Permian source rocks in the Junggar Basin, China, as a case study for the application of a new method for characterizing maturity based on nanomechanical properties. Atomic force microscopy (AFM) was used to characterize the nanomechanical properties of vitrinite at different maturity levels, and the results were combined with a micro-Fourier-transform infrared (FTIR) study to examine the chemical structural response to changes in mechanical properties. The Young's modulus of vitrinite increases in an S-shaped trend with maturity, reflecting differences in chemical structure at different evolutionary stages. In the early stage of maturity (< 0.7% R_o), vitrinite is relatively soft with a heterogeneous mechanical structure. On reaching the oil-generation threshold (∼0.7% R_o), the removal of aliphatic structures leads to a significant increase in Young's modulus. At the oil-generation peak (∼1.0% R_o), the Young's modulus increase slightly. When the vitrinite enters the gas window (> 1.3% R_o), the higher aromatization level results in a high Young's modulus, with mechanical structure becoming more homogeneous. The results show that maturity can be effectively characterized on the basis of the nanomechanical properties of vitrinite. This method has unique advantages in determining the maturity of organic matter with vitrinites being dominated by nanoscale particles, which cannot be easily measured by conventional microscopes. Thus, the AFM method is a useful supplement to the traditional vitrinite reflectance measurement.