Biosynthesis and metabolism of dipeptidylpeptidase IV in primary cultured rat hepatocytes and Morris hepatoma 7777 cells.
N-Glycosylation, biosynthesis and degradation of dipeptidylpeptidase IV (EC 3.4.14.5) (DPP IV) were comparatively studied in primary cultured rat hepatocytes and Morris hepatoma 7777 cells (MH 7777 cells). DPP IV had a molecular mass of 105 kDa in rat hepatocytes and of 103 kDa in MH 7777 cells as assessed by SDS/PAGE under reducing conditions. This difference in molecular mass was caused by differences in covalently attached N-glycans. DPP IV from hepatoma cells contained a higher proportion of N-glycans of the oligomannosidic or hybrid type and therefore migrated at a slightly lower molecular mass. In both cell types DPP IV was initially synthesized as a 97-kDa precursor which was completely susceptible to digestion with endo-beta-N-acetylglucosaminidase H converting the molecular mass to 84 kDa. The precursor was processed to the mature forms of DPP IV, glycosylated with N-glycans mainly of the complex type with a half-life of 20-25 min. The transit of newly synthesized DPP IV to the cell surface displayed identical or very similar kinetics in both cell types with the major portion of DPP IV appearing at the cell surface after 60 min. DPP IV molecules were very slowly degraded in hepatocytes as well as in hepatoma cells with half-lives of approximately 45 h. Inhibition of oligosaccharide processing with 1-deoxymannojirimycin led to the formation of DPP IV molecules containing N-glycans of the oligomannosidic type. This glycosylation variant was degraded with the same half-life as complex-type glycosylated DPP IV. By contrast, inhibition of N-glycosylation with tunicamycin resulted into rapid degradation of non-N-glycosylated DPP IV molecules in both cell types. Non-N-glycosylated DPP IV could not be detected at the cell surface indicating an intracellular proteolytic process soon after biosynthesis.
Loch
N
N
Institut für Molekularbiologie und Biochemie, Freie Universität Berlin, Federal Republic of Germany.
Tauber
R
R
Becker
A
A
Hartel-Schenk
S
S
Reutter
W
W
D000818:Animals; D001692:Biological Transport; D002478:Cells, Cultured; D018819:Dipeptidyl Peptidase 4; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases / Q000096:biosynthesis* / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D006031:Glycosylation; D007700:Kinetics; D008099:Liver / Q000166:cytology / Q000201:enzymology*; D008114:Liver Neoplasms, Experimental / Q000201:enzymology*; D008970:Molecular Weight; D011134:Polysaccharides / Q000378:metabolism; D011233:Precipitin Tests; D051381:Rats; D014407:Tumor Cells, Cultured
D011134:Polysaccharides; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases; D018819:Dipeptidyl Peptidase 4