Effect of monensin on the metabolism, localization, and biosynthesis of N- and O-linked oligosaccharides of galactosyltransferase.

The effect of monensin on the metabolism and glycosylation of the trans-Golgi enzyme galactosyltransferase (GT) was studied in HeLa cells. The synthesis of GT was not affected by monensin; however, in the presence of the drug GT precursor molecules (Mr 54 000 and 47 000) were transported to the Golgi system at a slower rate than in control cells. the half-life of GT as an intrinsic membrane protein of the trans-Golgi system was prolonged in the presence of monensin. Mature GT (Mr 54 000) contained in addition to one N-linked oligosaccharide several O-linked oligosaccharides; the majority of them consisted of the disaccharide galactose-N-acetylgalactosamine containing one or two sialic acid residues. We conclude that GT resides at an intracellular location where significant sialylation occurs. The sialylation of O-linked oligosaccharides was not hampered by the presence of monensin. Two-dimensional gel electrophoresis combined with neuraminidase treatment revealed that GT consists of a number of species in a pI range between 4 and 7. Removal of sialic acid residues resulted in a less acidic pI range of GT but the charge heterogeneity persisted. Immunoelectron microscopy with antibodies against GT showed that monensin affects primarily the GT-containing cisternae.