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 subcellular fractions obtained by differential centrifugation of human liver homogenate. Both enzymes were exclusively enriched in the microsomal fraction with a recovery of total enzyme activity of 65.9+/-9.9% and 69.1+/-13.8%, respectively. Microsomal preparations were further subfractionated by isopycnic centrifugation on a continuous sucrose density gradient. Both glucosyltransferases closely followed marker constituents of endoplasmic reticulum, as shown by similar distribution profiles in the gradient, but differed in their quantitative distribution among the endoplasmic reticulum membranes. The bile acid glucosyltransferase showed an almost identical distribution with NADPH-cytochrome c reductase as marker of smooth endoplasmic reticulum with a modal density of 1.16 g/cm(3). The uridine 5'-diphosphate-glucose dolichyl-phosphate glucosyltransferase equilibrated at a higher density with a peak at a modal density of 1.174 g/cm(3). Its marked overlap with the distribution of NADPH-cytochrome c reductase suggests that the major activity of uridine 5'-diphosphate-glucose dolichyl-phosphate glucosyltransferase 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 membranes derived from Golgi-complex or plasma membranes could be excluded by treatment of microsomes with membrane reagents prior to isopycnic centrifugation. Digitonin did not alter the equilibrium densities of the glucosyltransferases and endoplasmic reticulum markers in contrast to markers of plasma membranes and the Golgi-complex shifting to higher densities. The reversed effect was observed in case of pretreatment of microsomes with pyrophosphate known to detach ribosomes. Only membranes associated with both glucosyltransferases and endoplasmic reticulum marker constituents showed a marked shift towards lower equilibrium densities, whereas no influence was seen on markers of plasma membranes and Golgi-complex. These results indicate that the uridine 5'-diphosphate-glucose dolichyl-phosphate glucosyltransferase and the bile acid glucosyltransferase are exclusively confined to the endoplasmic reticulum in human liver. (C) Journal of Hepatology.