THE SUBMICROSOMAL LOCALIZATION OF URIDINE 5'-DIPHOSPHATE-GLUCOSE DOLICHYL-PHOSPHATE GLUCOSYLTRANSFERASE AND BILE-ACID GLUCOSYLTRANSFERASE IN THE HUMAN LIVER

Uridine 5'-diphosphate-glucose dolichyl-phosphate glucosyltransferase and bile acid glucosyltransferase were quantitatively determined in su bcellular fractions obtained by differential centrifugation of human l iver homogenate. Both enzymes were exclusively enriched in the microso mal fraction with a recovery of total enzyme activity of 65.9+/-9.9% a nd 69.1+/-13.8%, respectively. Microsomal preparations were further su bfractionated by isopycnic centrifugation on a continuous sucrose dens ity gradient. Both glucosyltransferases closely followed marker consti tuents of endoplasmic reticulum, as shown by similar distribution prof iles in the gradient, but differed in their quantitative distribution among the endoplasmic reticulum membranes. The bile acid glucosyltrans ferase showed an almost identical distribution with NADPH-cytochrome c reductase as marker of smooth endoplasmic reticulum with a modal dens ity of 1.16 g/cm(3). The uridine 5'-diphosphate-glucose dolichyl-phosp hate glucosyltransferase equilibrated at a higher density with a peak at a modal density of 1.174 g/cm(3). Its marked overlap with the distr ibution of NADPH-cytochrome c reductase suggests that the major activi ty of uridine 5'-diphosphate-glucose dolichyl-phosphate glucosyltransf erase is also associated with smooth endoplasmic reticulum membranes, whereas minor proportions of enzyme activity are present in the rough endoplasmic reticulum. Association of both glucosyltransferases with m embranes derived from Golgi-complex or plasma membranes could be exclu ded by treatment of microsomes with membrane reagents prior to isopycn ic centrifugation. Digitonin did not alter the equilibrium densities o f the glucosyltransferases and endoplasmic reticulum markers in contra st to markers of plasma membranes and the Golgi-complex shifting to hi gher densities. The reversed effect was observed in case of pretreatme nt of microsomes with pyrophosphate known to detach ribosomes. Only me mbranes associated with both glucosyltransferases and endoplasmic reti culum marker constituents showed a marked shift towards lower equilibr ium densities, whereas no influence was seen on markers of plasma memb ranes and Golgi-complex. These results indicate that the uridine 5'-di phosphate-glucose dolichyl-phosphate glucosyltransferase and the bile acid glucosyltransferase are exclusively confined to the endoplasmic r eticulum in human liver. (C) Journal of Hepatology.