To investigate the optimal supplementation of dietary exogenous protease in plant-based diets of red swamp crayfish (Procambarus clarkii), crayfish [initial mean weight (9.25±0.20) g ]were fed six isonitrogenous and isoenergetic diets which were formulated to contain graded protease levels (0, 0.1, 0.2, 0.4, 0.8 and 1.6 g/kg protease, respectively) for 8 weeks. Each diet was fed to triplicate tanks with 15 per crayfish in each tank. The results showed that, with the increase of dietary protease supplementation: ① There were no significant difference in survival rate, hepatosomatic index and flesh content among groups. The weight gain rate firstly increased and then decreased, and reached the maximum when dietary protease supplementation level was 0.2 g/kg. The feed conversation ratio reached the lowest value when dietary protease supplementation level was 0.4 g/kg. The protein efficiency and protein deposition rate reached the highest in shrimp fed the diet with 0.4 and 0.2 g/kg protease, respectively. Broken-line model analysis in terms of the weight gain rate, feed conversion ratio and protein efficiency indicated that optimal dietary protease supplement of crayfish was 0.16, 0.24 and 0.23 g/kg, respectively.② The crude protein contents in muscle and whole shrimp firstly increased and then decreased, and the maximum occured at the shrimp fed the diet with 0.2 g/kg protease. ③ The activities of protease, lipase and amylase in intestine and hepatopancreas all reached the maximum at the shrimp fed the diet with 0.2 g/kg protease, which was significantly higher than the control shrimp. ④ The maximum content of total protein in serum were observed in shrimps fed the diets with 0.4 g/kg protease; the activities of glutamic oxaloacetic transaminase and glutamic pyruvic transaminase firstly decreased and then increased, with the lowest value occurs at the shrimp fed the diet with 0.4 g/kg protease; the activities of alkaline phosphatase and acid phosphatase firstly increased and then decreased, and reached the maximum in 0.2 g/kg group and 0.4 g/kg group, respectively. The content of malondialdehyde had the minimum value at the shrimp fed the diet with 0.2 g/kg protease. ⑤ The activities of alkaline phosphatase and acid phosphatase in hepatopancreas firstly increased and then decreased, and reached the maximum at the shrimp fed the diet with 0.2 g/kg protease, which was significantly higher than the control shrimp. In conclusion, under this experimental condition, the appropriate supplementation of dietary protease in red swamp crayfish is from 0.16 to 0.40 g/kg.