The synthesis and turnover of 5'-nucleotidase in primary cultured hepatocytes.
The synthesis and degradation of 5'-nucleotidase has been studied in rat hepatocytes. Primary cultures of rat hepatocytes were established with the cells showing evidence of polarity after 24-36 h in culture. After a 30 h lag period 5'-nucleotidase activity increased to a plateau level similar to the activity found in whole liver. The half life of the enzyme after reaching the plateau of activity was 22.8 h. Pulse-chase biosynthetic labelling studies of 5'-nucleotidase in the cultured hepatocytes using [35S]methionine showed that the 5'-nucleotidase monomer was synthesised as an Mr 67,000 form which was converted to the mature Mr 72,000 form. [35S]Methionine labelling studies in the presence of tunicamycin showed that the unglycosylated protein monomer was an Mr 57,000 form. The immature Mr 67,000 form of 5'-nucleotidase was sensitive to endoglycosidase H, whereas the mature form was sensitive only to endoglycosidase F. The data presented are consistent with 5'-nucleotidase in a polarised cell being synthesised and processed like other membrane glycoproteins, in contrast to earlier reports.
D015720:5'-Nucleotidase; D000118:Acetylglucosaminidase / Q000378:metabolism; D000818:Animals; D002478:Cells, Cultured; D006026:Glycoside Hydrolases / Q000378:metabolism; D006031:Glycosylation; D006207:Half-Life; D007700:Kinetics; D008099:Liver / Q000187:drug effects / Q000201:enzymology*; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D008970:Molecular Weight; D009708:Nucleotidases / Q000096:biosynthesis / Q000378:metabolism*; D051381:Rats; D014415:Tunicamycin / Q000494:pharmacology
D014415:Tunicamycin; D009708:Nucleotidases; D015720:5'-Nucleotidase; D006026:Glycoside Hydrolases; D000118:Acetylglucosaminidase; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase