Evidence for the translocation of 5'-nucleotidase across hepatic membranes in vivo.
Hepatic 5'-nucleotidase (EC 3.1.3.5; 5'-ribonucleotide phosphohydrolase) activity has been studied in cisternal elements of the Golgi complex and in secretion vacuoles, both isolated after ethanol administration to rats in vivo. The enzyme in secretion vacuoles was latent, so that a 5-fold increase in activity was observed when incubations were carried out in the presence of detergent; evidence is presented that the latency is caused by the impermeability of the membrane to substrate. Essentially no latency was observed in Golgi cisternae. Confirming the results of Farquhar et al. [(1974) J. Cell Biol. 60, 8-25], reaction product from 5'-nucleotidase was localized by cytochemical procedures on the inside of secretion vacuoles and on the cytoplasmic side of Golgi cisternae. After solubilization in detergent, the enzyme from both fractions reacted almost identically with both antibody to the purified enzyme and concanavalin A. In contrast, when intact fractions were incubated with an excess of antibody or concanavalin A, only 22-23% of the enzyme was inhibited in secretion vacuoles whereas 51-84% was inhibited in Golgi cisternae. Sonication of secretion vacuoles in the presence of antibody or concanavalin A increased the inhibition 2- to 3-fold. It is suggested that during the formation of secretion vacuoles from the Golgi cisternae, 5'-nucleotidase is translocated from the cytoplasmic side of the membrane to the inside.
D000818:Animals; D000937:Antigen-Antibody Reactions; D002462:Cell Membrane / Q000187:drug effects / Q000201:enzymology; D003208:Concanavalin A / Q000494:pharmacology; D000431:Ethanol / Q000494:pharmacology; D006056:Golgi Apparatus / Q000201:enzymology* / Q000648:ultrastructure; D008099:Liver / Q000201:enzymology*; D008566:Membranes / Q000187:drug effects / Q000201:enzymology / Q000648:ultrastructure; D008854:Microscopy, Electron; D008862:Microsomes, Liver / Q000201:enzymology; D009708:Nucleotidases / Q000276:immunology / Q000378:metabolism*; D051381:Rats
D003208:Concanavalin A; D000431:Ethanol; D009708:Nucleotidases