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The sensitivity of soy protein isolate to trypsin was characterized by its flexibility. The effects of different homogenization pressures (0~200MPa) on soy protein isolate flexibility and emulsifying properties were investigated. The changes of soy protein isolate structure and the correlation analysis between flexibility and emulsifying properties (emulsifying activity and emulsion stability) were also investigated. The results showed that when the homogenization pressure was 0~160MPa, the soy protein isolate flexibility was increased with the increase of the homogenization pressure, while the flexibility did not change obviously at 160~180MPa. When the homogenization pressure was 180~200MPa, the soy protein isolate flexibility was declined. The surface hydrophobicity was increased with the increase of homogeneous pressure, while the turbidity was decreased. The changing tendency and flexibility of the emulsification property with homogeneous pressure were consistent with the homogeneous pressure. Correlation analysis suggested that the soy protein isolate flexibility had a linear positive correlation with the emulsifying activity and the emulsifying stability, and the correlation coefficients were 0.893 and 0.938, respectively. Through ultraviolet spectral scanning analysis of soy protein isolate and endogenous tryptophan fluorescence spectroscopy analysis, it was found that as the conformational flexibility of soy protein isolate was increased, the structure became more stretch.