How protein metabolism is perturbed during chronic glucocorticoid excess is poorly understood. The aims were to investigate the impact of chronic glucocorticoid excess and restoration of eucortisolemia in Cushing’s syndrome (CS) on whole body protein metabolism. 18 subjects with CS and 18 normal subjects (NS) underwent assessment of body composition using DXA and whole body protein turnover with a 3h constant infusion of l-[13C] leucine, allowing calculation of rates of leucine appearance (LRa), leucine oxidation (Lox) and leucine incorporation into protein (LIP). 10 subjects with CS were restudied after restoration of eucortisolemia. Percentage FM was greater (43.9±1.6% vs 33.8±2.4%, p=0.002) and LBM lower (52.7±1.6% vs 62.1±2.3%, p=0.002) in CS. LBM was significantly correlated (r²>0.44, p<0.005) to LRa, Lox and LIP in both groups. After correcting for LBM, LRa (133±5 vs 116±5µmol/min, p=0.02) and Lox (29±1 vs 24±1µmol/min, p=0.01) were greater in CS. FM significantly correlated (r²=0.23, p<0.05) with LRa and LIP, but not Lox in CS. In multiple regression, LBM was an independent determinant of all 3 indices of leucine turnover, FM of LRa and LIP and CS of Lox. Following restoration of eucortisolemia, Lox was reduced (-7.5±2.6 µmol/min, p=0.02) and LIP increased (+15.2±6.2 µmol/min, p=0.04). In summary, whole body protein metabolism in CS is influenced by changes in body composition and glucocorticoid excess per se, which increases protein oxidation. Enhanced protein oxidation is a likely explanation for the reduced protein mass in CS. Successful treatment of CS reduces protein oxidation and increases protein synthesis to prevent ongoing protein loss.