Role of carbohydrate in biological function of the adhesive glycoprotein fibronectin.

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Abstract
The role of the carbohydrate moiety in the biological activity of fibronectin in vitro was studied by using tunicamycin to inhibit the glycosylation of this glycoprotein. Tunicamycin is a glucosamine-containing antibiotic that specifically inhibits glycosylation of protein asparaginyl residues mediated by dolichol pyrophosphate. Fibronectin synthesized in the presence of 0.5 .mu.g of tunicamycin per ml was not glycosylated, as determined by amino sugar analysis, lack of incorporation of [14C]glucosamine and [3H]mannose, and concanavalin A binding studies. Nonglycosylated fibronectin that was isolated from chicken embryo fibroblasts and added to [SV-40-transformed 3T3] cells in vitro was as effective as the glycosylated protein in promoting a more normal fibroblastic phenotype, including cell flattening, elongation of cell processes and parallel alignment of cells. The nonglycosylated protein was as effective as the glycosylated species in mediating cell attachment to collagen and spreading on plastic, and in agglutination of formalin-fixed sheep erythrocytes. The non-glycosylated protein was twice as sensitive as the glycosylated protein to proteolytic hydrolysis in vitro as was suggested by previous studies with intact cells. The carbohydrate moiety of fibronectin is not required for the mediation of a number of biological activities characteristic of this glycoprotein.