The mechanistic target of rapamycin (mTOR) is a key regulator of protein synthesis in skeletal muscle particularly sensitive to nutrient availability. It is not fully understood how carbohydrate and amino acid availability results in activation of mTOR or the downstream propagation of intracellular signals resulting in cell growth in human skeletal muscle. Recently, it has emerged that mTOR localisation to the surface of the lysosome, and translocation to the cell membrane in its active form may be critical for its nutrient-stimulated downstream effects. We utilized novel biochemical approaches to study mTOR localization and protein complex formation in human skeletal muscle in the fasted state and 1h and 3h post consumption of a protein-carbohydrate beverage (Gatorade Recover®, Gatorade, IL, USA) providing 20/44/1g of protein/carbohydrate/fat. In contrast to cell and rodent studies, we observed mTOR to interact with the lysosomal marker LAMP2 in basal conditions and mTOR/LAMP2 complexes translocating to the cell periphery following nutrient stimulation. This redistribution of mTOR coincided with increased mTOR activity as assessed via kinase activity and immunoblotting of proximal targets of mTOR. Collectively our results provide information about the nutrient-sensing mechanisms regulating the activation of mTOR in skeletal muscle, suggesting that mTOR cellular localisation may be fundamentally important for the initiation of the protein synthetic response following nutrient stimulation in human skeletal muscle. It is anticipated that further understanding of these mechanisms will optimize our ability to combine the two potent activators of skeletal muscle protein synthesis – resistance exercise and nutrition – to maximize the growth response in humans.