In order to study the effect of ultrasonic pretreatment on the structure of peanut protein isolate microgel particles (PPIMP) and the characteristics of Pickering emulsion stabilized by it, the structural changes of PPIMP after ultrasonic treatment at a 500 W power for 10, 20, 30 and 40 min were evaluated in terms of its particle size, zeta potential, surface hydrophobicity, infrared spectrum, and emulsifying properties and the particle size distribution, stability index and rheological properties of the Pickering emulsion were systematically investigated. The results showed PPIMP pretreated by ultrasonic for 20 min had the smallest particle size, and the largest surface hydrophobicity, emulsifying capacity and emulsion stability. Meanwhile, the relative content of α-helix in PPIMP increased, the relative content of β-sheet decreased, the hydrogen bonds between protein molecules were destroyed, and the protein molecules were unfolded. In addition, Pickering emulsions with oil phase volume fractions of 65%, 70%, 75%, and 80% stabilized by PPIMP exhibited a typical non-Newtonian pseudoplastic behavior, and the Pickering emulsion with 80% phase oil presented the best stability among all prepared Pickering emulsions. The results of this study confirmed that PPIMP can be used as an effective Pickering emulsion stabilizer, which is helpful for the preparation of PPIMP-based Pickering emulsion with high stability and its application in the food industry.