Functional role of laminin carbohydrate

Previous work showed that tunicamycin suppresses glycosylation of laminin. In the present work, the role of glycosylation in the secretion of laminin and in the disulfide bonding of laminin subunits was studied, using tunicamycin to inhibit glycosylation. Tunicamycin inhibited extensively the secretion of laminin into culture medium and extracellular matrix even though the treated cells contained higher concentrations of laminin than the control cells. The laminin subunits synthesized in the presence of tunicamycin were disulfide bonded. Thus, suppression of glycosylation did not adversely affect disulfide bonding of the subunits, but did decrease the secretion of laminin. Glycosidases were also used to remove the carbohydrate of laminin to study the role of carbohydrate in the stability of laminin and in its interaction with another extracellular matrix component, heparin. The glycosidases removed about 73% of [3H]glucosamine. Both glycosidase-treated and untreated laminin were stable when incubated with cell lysate or culture medium. The glycosidase-treated laminin bound as efficiently as the untreated laminin to heparin. These results suggest that the presence of a carbohydrate moiety, at least at the level found in untreated laminin, is not essential in binding to heparin or in protecting laminin from proteolytic degradation in the cell or culture medium.