The fermentation characteristics of milk protein concentrate (MPC) with different concentration of lactose was studied after fermentation using Lactobacillus bulgaricus and Streptococcus thermophilus. The results showed that lactose concentration affected the time of fermentation entering stable phase and pH value. With the increase of lactose concentration, the final pH value of MPC fermentation was decreased (4.14-4.59). The microrheological parameters indicated that the solid-liquid balance and fluidity index of MPC fermentation product were increased with the increase of lactose concentration, while the macroscopic viscosity index was decreased. The texture characteristics showed that the water retention rate of MPC fermented product increased with the increase of lactose concentration, and the gumminess and elasticity increased first and then decreased, while hardness and viscosity were not affected. The results of GC-MS showed that the acid compounds and ketone compounds were the main volatile substances in the MPC fermentation product. The content of acid compounds were decreased, while the content of lactose metabolites such as 2,3-butanedione and 3-hydroxy-2-butanone were increased accompanied by the lactose increased. The results of this study provided basic data for further study of the fermentation characteristics of MPC and broadened the application of MPC in different fermented dairy products.