Role of tyrosine-5 in the cytoplasmic tail of the macrophage asialoglycoprotein receptor in the rapid internalization of ligands.
A macrophage asialoglycoprotein binding protein (M-ASGP-BP), which is an endocytic receptor specific for Gal/GalNAc-terminated glycoproteins, was shown to be functionally active as a homooligomer (hexamer or octamer) of a single polypeptide chain of 42 kDa [Ozaki, K., Ii, M., Itoh, N., & Kawasaki, T. (1992) J. Biol. Chem. 267, 9229-9235]. In various endocytic receptors, a four-amino-acid sequence, Tyr-X-Y-Phe, in the cytoplasmic domain has been identified as an internalization signal [Pearse, B.M.F. & Robinson, M.S. (1990) Annu. Rev. Cell Biol. 265, 151-171]. The amino-terminus of the M-ASGP-BP deduced from its cDNA sequence was found to contain the sequence, Tyr5-Glu6-Asn7-Phe8, in its cytoplasmic tail. This was confirmed by the fact that the recombinant M-ASGP-BP isolated from transfected COS-1 cells was found to have the amino-terminal sequence, Thr2-Met3-Ala4-Tyr5-Glu6-Asn7-Phe8. The role of this presumptive internalization signal in the cytoplasmic tail was studied by measuring the endocytic activity of the wild-type and mutant M-ASGP-BPs expressed on COS-1 cells through transfection with the wild-type and mutant cDNAs prepared by oligonucleotide-directed mutagenesis, respectively. On the deletion of Tyr5 or replacement of it with alanine, the internalization of asialoorosomucoid (ASOR) decreased to approximately one-fourth that in the case of the wild-type molecule. On replacement of Tyr5 with phenylalanine, the internalization proceeded at a rate similar to that in the case of the wild-type molecule.(ABSTRACT TRUNCATED AT 250 WORDS)
Ozaki
K
K
Department of Biological Chemistry, Faculty of Pharmaceutical Sciences, Kyoto University.
Itoh
N
N
Kawasaki
T
T
D000595:Amino Acid Sequence; D000818:Animals; D001212:Asialoglycoproteins / Q000378:metabolism*; D001483:Base Sequence; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D003593:Cytoplasm; D004247:DNA; D037102:Lectins; D008264:Macrophages / Q000737:chemistry*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010649:Phenylalanine / Q000737:chemistry; D011956:Receptors, Cell Surface; D014162:Transfection; D014443:Tyrosine / Q000737:chemistry / Q000235:genetics / Q000502:physiology*
D001212:Asialoglycoproteins; D002352:Carrier Proteins; C023339:Clec10a protein, rat; D037102:Lectins; D011956:Receptors, Cell Surface; D014443:Tyrosine; D010649:Phenylalanine; D004247:DNA