McGraw T E, Pytowski B, Arzt J, Ferrone C
Department of Pathology, Columbia University College of Physicians and Surgeons, New York 10032.
J Cell Biol. 1991 Mar;112(5):853-61. doi: 10.1083/jcb.112.5.853.
Site-specific mutagenesis has been used to define the sequences required for efficient internalization of the human transferrin receptor. It has previously been shown that the sole cytoplasmic tyrosine, at position 20, is required for efficient internalization. When two other cytoplasmic aromatic residues, the phenylalanines at positions 13 and 23, are substituted with alanines internalization is also reduced. The phenylalanine 23 mutation decreases the internalization rate constant approximately threefold, and mutation of phenylalanine 13 decreases it by approximately twofold. The mutation at position 23 has as serious an effect on internalization as substitution with a nonaromatic amino acid for the single tyrosine. These results demonstrate the importance of several aromatic amino acids in maintaining efficient internalization of the transferrin receptor. Substitution of a tyrosine at a second site, for a serine at position 34, within the cytoplasmic domain of a transferrin receptor with a nonaromatic amino acid at position 20, results in a complete reversion of the internalization-defective phenotype. This reversion is completely dependent upon a tyrosine, as phenylalanine substituted at position 34 does not revert the internalization-defective phenotype. This result demonstrates that a tyrosine placed outside of its native context can still function in the internalization of the transferrin receptor, suggesting a flexibility in surrounding sequences required for efficient internalization.
位点特异性诱变已被用于确定人转铁蛋白受体有效内化所需的序列。此前已表明，第20位唯一的胞质酪氨酸是有效内化所必需的。当另外两个胞质芳香族残基，即第13位和第23位的苯丙氨酸被丙氨酸取代时，内化也会减少。苯丙氨酸23突变使内化速率常数降低约三倍，苯丙氨酸13突变使其降低约两倍。第23位的突变对内化的影响与用非芳香族氨基酸取代单个酪氨酸一样严重。这些结果证明了几种芳香族氨基酸在维持转铁蛋白受体有效内化中的重要性。在转铁蛋白受体胞质结构域内，将第34位的酪氨酸替换为丝氨酸，同时在第20位用非芳香族氨基酸取代，会导致内化缺陷表型完全恢复。这种恢复完全依赖于酪氨酸，因为在第
