Domain structure of fibronectin and its relation to function. Disulfides and sulfhydryl groups

Abstract

Hamster cell fibronectin is a glycoprotein consisting of two 230,000-dalton subunits in a disulfide-bonded dimer. The molecule is composed of domains which can be separated by partial proteolytic cleavage. The carbohydrates, disulfide bonds, and a single free sulfhydryl group per chain are distributed nonuniformly among these regions. All the interchain disulfides are within 10,000 daltons of the end of the molecule and are removed by mild proteolysis which also generates 200,000- and 25,000-dalton fragments which do not contain interchain disulfides. The 200,000-dalton fragment contains all or most of the carbohydrate side chains, and the free sulfhydryl group, but is relatively poor in cystine. The 25,000-dalton fragment is carbohydrate-free and cystine-rich but has no free sulfhydryl groups. There is heterogeneity in carbohydrate content among the monomeric chains of intact fibronectin and the 200,000-dalton fragments. The gelatin binding site of fibronectin is in the 200,000 fragment. Intact disulfide bonds are required for binding of fibronectin to cells and to gelatin and blockage of the free sulfhydryl groups prevents binding of fibronectin to cells, suggesting that intermolecular disulfide bonding may be important.