Structural domains of heparan sulphate for specific recognition of the C-terminal heparin-binding domain of human plasma fibronectin (HEPII).
Heparan sulphate (HS) is an abundant polysaccharide component of the pericellular domain and is found in most soft tissues and all adherent cells in culture. It interacts with a wide spectrum of proteins including polypeptide growth factors and glycoproteins of the extracellular matrix. These interactions might influence fundamental cellular activities such as adhesion, growth and migration. HS might therefore represent a highly adaptive mechanism by which cells respond to their environment. The present study shows that the interaction between fibroblast HS, metabolically labelled with [3H]glucosamine, and the C-terminal heparin-binding domain of human plasma fibronectin (HEPII), is determined by distinct regions of the polysaccharide chain. By using a very sensitive affinity-chromatography method and specific polysaccharide scission it was shown that the HEPII-binding regions of HS reside within sulphated domains that are resistant to degradation by heparinase III. In addition, optimal binding was achieved with specific heparinase III-resistant fragments of 14-16 monosaccharides in length. The affinity of HS for HEPII was significantly decreased when the polysaccharide was cleaved with heparinase I. Chondroitin sulphate and dermatan sulphate were poor competitive inhibitors of [3H]HS binding to HEPII whereas unlabelled HS and heparin gave a strong inhibitory activity, with heparin being the most potent inhibitor. These findings suggest that the interaction between HEPII and HS is specific and requires extended sequences of seven to eight N-sulphated disaccharides in which a proportion of the iduronate residues are sulphated at C-2. The results have important implications for the functions of HS in cell adhesion and migration.
Walker
A
A
CRC Department of Medical Oncology, Christie Hospital NHS Trust, Manchester, U.K.
Gallagher
J T
JT
D000818:Animals; D001665:Binding Sites; D002478:Cells, Cultured; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D005353:Fibronectins / Q000097:blood* / Q000737:chemistry; D006493:Heparin / Q000378:metabolism*; D019763:Heparin Lyase; D006497:Heparan Sulfate / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D011133:Polysaccharide-Lyases / Q000378:metabolism
D005353:Fibronectins; D006493:Heparin; D019763:Heparin Lyase; D006497:Heparan Sulfate; D011133:Polysaccharide-Lyases