BACKGROUND: Based on controlled 36 h experiments a higher dietary protein intake causes a positive protein balance and a negative fat balance. A positive net protein balance may support fat free mass accrual. However, few data are available on the impact of more prolonged changes in habitual protein intake on whole-body protein metabolism and basal muscle protein synthesis rates. OBJECTIVE: To assess changes in whole-body protein turnover and basal muscle protein synthesis rates following 12 weeks of adaptation to a low versus high dietary protein intake. METHODS: A randomized parallel study was performed in 40 subjects who followed either a high protein (2.4 g protein/kg/d) or low protein (0.4 g protein/kg/d) energy-balanced diet (30/35/35% or 5/60/35% energy from protein/carbohydrate/fat) for a period of 12 weeks. A subgroup of 7 men and 8 women (body mass index: 22.8+/-2.3 kg/m2, age: 24.3+/-4.9 y) were selected to evaluate the impact of prolonged adaptation to either a high or low protein intake on whole body protein metabolism and basal muscle protein synthesis rates. After the diet, subjects received continuous infusions with L-[ring-2H5]phenylalanine and L-[ring-2H2]tyrosine in an overnight fasted state, with blood samples and muscle biopsies being collected to assess post-absorptive whole-body protein turnover and muscle protein synthesis rates in vivo in humans. RESULTS: After 12 weeks of intervention, whole-body protein balance in the fasted state was more negative in the high protein treatment when compared with the low protein treatment (-4.1+/-0.5 vs -2.7+/-0.6 mumol phenylalanine/kg/h;P<0.001). Whole-body protein breakdown (43.0+/-4.4 vs 37.8+/-3.8 mumol phenylalanine/kg/h;P<0.03), synthesis (38.9+/-4.2 vs 35.1+/-3.6 mumol phenylalanine/kg/h;P<0.01) and phenylalanine hydroxylation rates (4.1+/-0.6 vs 2.7+/-0.6 mumol phenylalanine/kg/h;P<0.001) were significantly higher in the high vs low protein group. Basal muscle protein synthesis rates were maintained on a low vs high protein diet (0.042+/-0.01 vs 0.045+/-0.01%/h;P = 0.620). CONCLUSIONS: In the overnight fasted state, adaptation to a low-protein intake (0.4 g/kg/d) does not result in a more negative whole-body protein balance and does not lower basal muscle protein synthesis rates when compared to a high-protein intake. TRIAL REGISTRATION: Clinicaltrials.gov NCT01551238.