Ligand-induced internalization of epidermal growth factor (EGF) receptors via a high affinity saturable pathway requires sequences located in the carboxyl terminus distal to the tyrosine kinase domain. Three regions were found to contain endocytic motifs as defined by their ability to restore internalization function to EGF receptors truncated at the distal border of the kinase domain at residue 958. Deletional analysis identified the sequence 996QQGFF as essential for function of the region encompassing residues 993-1022. QQGFF and the deduced sequence of the region encompassing residues 973-991 (973FYRAL) could effectively replace the endogenous endocytic code of the transferrin receptor (YTRF). FYRAL and YTRF were less active than QQGFF when substituted into region 993-1022 of the EGF receptor, but a synthetic sequence (NNAYF), predicted to have structural features of a tight turn, effectively replaced QQGFF for EGF receptor internalization. Whereas EGF receptor sequences functioned effectively in the transferrin receptor, function of these sequences in the EGF receptor was strictly dependent on intrinsic tyrosine kinase activity as demonstrated kinetically and by immunofluorescence using semithin cryosections. Ligand-dependent endocytosis and down-regulation of the EGF receptor thus require multiple sequence motifs that are exchangeable between ligand-dependent and -independent receptors, but that require intrinsic tyrosine kinase activity for function in the context of the EGF receptor.