Abstract Context: Leptin regulates energy homeostasis by suppressing food intake; however, its role in energy expenditure and fat oxidation remains uncertain in humans. Objective: The aim of the study was to assess 24-h energy metabolism before and after weight loss induced by leptin treatment in congenital leptin-deficient subjects or low-calorie diet in controls. Design and Patients: We measured 24-h energy expenditure, 24-h fat oxidation, and body fat in three null homozygous leptin-deficient obese adults before and after weight loss induced by a 19-wk leptin replacement period (0.02–0.04 mg/kg/d). The same measures were performed in three obese controls pair-matched for sex, age, and weight loss induced by a 10- to 21-wk low-calorie diet. Measurements were preceded for 1 wk of weight stabilization. Energy expenditure was adjusted for fat-free mass, fat mass, sex, and age based on a reference population (n = 842; R2 = 0.85; P < 0.0001). Similarly, fat oxidation was adjusted for fat-free mass, percentage body fat, energy balance, and diet composition during the 24-h respiratory chamber stay (R2 = 0.38; P < 0.0001). Results: Before weight loss, congenital leptin-deficient and control subjects had similar energy expenditure. However, after weight loss (∼15 kg), controls had energy expenditures lower than expected for their new weight and body composition (−265 ± 76 kcal/d; P = 0.04), whereas leptin-treated subjects had values not different from the reference population (−128 ± 119 kcal/d; P = 0.67). Before weight loss, fat oxidation was similar between groups. However, after weight loss, leptin-treated subjects had higher fat oxidation than controls (P = 0.005) and higher than the reference population (P = 0.0001). Conclusion: In congenital leptin-deficient subjects, leptin replacement prevented the decrease in energy expenditure and fat oxidation often observed after weight loss.