A 60-day feeding trial has been carried out to access the optimal dietary crude protein (DCP) requirement of juvenile Jerdon’s carp, Hypselobarbus jerdoni. Six isoenergetic (around 400 Kcal DE/100 g), isolipidic (60.80 g/kg), and heteronitrogenous diets were prepared with graded levels of protein, viz., 200, 250, 300, 350, 400, and 450 g/kg (TCP 20-TCP 45 with 50 g/kg incremental level). A completely randomised design (CRD) was used to distribute 270 Jerdon’s carp juveniles (6.12–6.18 g) in six experimental groups in triplicates (15 fish/tank, 200 l water capacity). After 60 days, the fishes of the TCP30 group showed significantly higher weight gain, final body weight, and percentage weight gain (WG%). The fish of the TCP30 group exhibited a significantly higher feed efficiency ratio and specific growth rate. On the other hand, a significantly lower feed conversion ratio is recorded in the TCP30 and TCP35 groups. The protein efficiency ratio of fish was reduced significantly with the elevated DCP level. Whole-body moisture, lipid, and total ash contents of H. jerdoni were remain unaffected by DCP levels. However, DCP levels significantly influenced the whole-body protein of H. jerdoni juveniles, with significantly higher values noticed in the TCP30 and TCP35 groups. The TCP35, TCP40, and TCP45 groups exhibited significantly higher (p<0.05) protease activity, and the amylase activity showed a decreasing trend in response to dietary protein levels. A significantly (p<0.05) higher SOD and CAT activity were observed in the TCP20 and TCP25 groups. However, lower hepatic glutamate pyruvate transaminase and glutamic-oxaloacetic transaminase activity were observed in the TCP30 and TCP35 groups, respectively. Furthermore, based on broken-line linear and second-order polynomial regression with respect to WG%, the optimal dietary crude protein requirement of H. jerdoni cultured for 60 days was found to be 309.72 and 316.40 g/kg.