The role of the transferrin homologue, melanotransferrin (p97), in iron metabolism has been studied using the human melanoma cell line, SK-MEL-28, which expresses this antigen in high concentrations. The mechanisms of iron and transferrin uptake were investigated using human transferrin labelled with iodine-125 and iron-59. Internalised and membrane-bound iron and transferrin were separated using the proteinase, pronase. The uptake of iron from transferrin occurred by at least two processes. The first process was saturable and consistent with receptor-mediated endocytosis, involving internalisation of transferrin bound to specific binding sites. Uptake of iron also occurred by a second process which was non-saturable up to 0.06 mg/ml (0.75 microM) and was of higher efficiency than the saturable process. This process of iron uptake may be the dominant one at physiological serum transferrin concentrations. A membrane-bound, pronase-sensitive, temperature-dependent, iron-binding component was also identified. The number of binding sites was estimated to be approx. 340,000 per cell (assuming 2 atoms of iron per site) and it is suggested that this binding component may be melanotransferrin.