Biosynthesis and turnover of a Mr = 110,000 glycoprotein localized to the hepatocyte bile canaliculus.
Antiserum was raised in rabbits against a bile canalicular glycoprotein of Mr = 110,000 purified to homogeneity from of rat liver. The antisera specifically immunoprecipitated a Mr = 110,000 polypeptide from hepatocytes metabolically labeled with [35S]methionine. When hepatocytes in primary culture were incubated with tunicamycin before labeling with [35S]methionine in the presence of tunicamycin, the major polypeptide immunoprecipitated by the specific antiserum from Triton X-100 extracts of cells had a molecular weight of 59,000. Enzymatic removal of N-linked carbohydrates from the Mr = 110,000 glycoprotein by N-glycanase digestion also yielded a polypeptide with minimum Mr = 59,000. In pulse-chase experiments using [35S]methionine, the Mr = 110,000 protein detected by the specific antisera first appears as Mr = 85,000 and 75,000 intermediate species which are endoglycosidase H sensitive. The Mr = 85,000 intermediate form is lost first with time followed by the Mr = 75,000 form giving rise to the Mr = 110,000 form that is endoglycosidase H resistant. Neuraminidase digestion of the Mr = 110,000 form generated an Mr 85,000 form but with a different carbohydrate structure than the intermediate Mr 85,000 form detected in the pulse-chase experiments. The time required to accomplish the processing of the Mr = 85,000 and 75,000 forms is relatively slow. Finally, the terminal sugars are added and the mature Mr = 110,000 glycoprotein is rapidly transported to the cell surface. A minimum time of 90 min is required for the Mr = 110,000 bile canalicular glycoprotein to be synthesized, processed, and reach the cell surface which is long relative to the time required (10 min) for another domain-specific protein, the receptor for asialoglycoproteins, to reach the sinusoidal surface. The Mr = 110,000 bile canalicular glycoprotein turns over in the bile canalicular domain with a half-life of 43 h while the asialoglycoprotein receptor turns over in the sinusoidal domain with a half-life of 23 h.
Diamond
M
M
Department of Biological Sciences, State University of New York at Buffalo 14260.
Petell
J K
JK
Doyle
D
D
D000818:Animals; D037263:Asialoglycoprotein Receptor; D001648:Bile Canaliculi / Q000378:metabolism*; D001653:Bile Ducts, Intrahepatic / Q000378:metabolism*; D050260:Carbohydrate Metabolism; D002478:Cells, Cultured; D006023:Glycoproteins / Q000378:metabolism*; D008099:Liver / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D008970:Molecular Weight; D051381:Rats; D011915:Rats, Inbred BUF; D011971:Receptors, Immunologic / Q000032:analysis; D014415:Tunicamycin / Q000494:pharmacology
D037263:Asialoglycoprotein Receptor; D006023:Glycoproteins; D008565:Membrane Proteins; D011971:Receptors, Immunologic; D014415:Tunicamycin