Two asialoglycoprotein receptor polypeptides in human hepatoma cells.
Two cDNA clones isolated from a HepG2 lambda gt11 library encode the classical asialoglycoprotein receptor, H1, as well as a homologous membrane glycoprotein, H2 (Spiess, M., and Lodish, H.F. (1985) Proc. Natl. Acad. Sci. U.S.A. 82, 6465-6469). To study the relationship of H2 to H1 and its possible role in receptor-mediated endocytosis of desialyated glycoproteins, we generated anti-peptide antibodies that are specific for each polypeptide. As judged by metabolic labeling of HepG2 cells and specific immunoadsorption, the biosynthesis of H2 is similar to H1 (Schwartz, A.L., and Rup, D. (1983) J. Biol. Chem. 258, 11249-11255); H2 is synthesized as a 43,000-dalton precursor polypeptide containing high mannose-type oligosaccharides, that is processed to a 50,000-dalton mature glycoprotein containing complex-type oligosaccharides. Both H1 and H2 have a half-life of approximately 12 h. Trypsin and neuraminidase digestion of intact cells at 4 and 12 degrees C was used to determine that, at steady state, 50-60 percent of both H1 and H2 are on the cell surface. Furthermore, all of the H2 molecules were digested by extracellular neuraminidase in 1 h at 37 degrees C, indicating that all gain access to the plasma membrane. Both H1 and H2 were purified to homogeneity when Triton X-100-solubilized membrane proteins from [35S]cysteine-labeled cells were subjected to affinity chromatography on galactose-agarose. Since we cannot detect a complex between mature H1 and H2, H2 must be a galactose-binding protein. Both quantitative immunoprecipitation of each polypeptide from HepG2 cells and the recovery of purified H1 and H2 from galactose-agarose affinity chromatography indicate that there is 5-6 times more H1 relative to H2. That H2 is a minor species, compared to H1, might explain why it was not observed until a specific antibody was utilized.
D037263:Asialoglycoprotein Receptor; D006528:Carcinoma, Hepatocellular / Q000032:analysis*; D002478:Cells, Cultured; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D003545:Cysteine / Q000378:metabolism; D004247:DNA / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008113:Liver Neoplasms / Q000032:analysis*; D008970:Molecular Weight; D010455:Peptides / Q000032:analysis* / Q000235:genetics; D011971:Receptors, Immunologic / Q000032:analysis* / Q000235:genetics; D014357:Trypsin / Q000378:metabolism
D037263:Asialoglycoprotein Receptor; D010455:Peptides; D011971:Receptors, Immunologic; D004247:DNA; D014357:Trypsin; D003545:Cysteine