Yamada K M, Kennedy D W, Kimata K, Pratt R M
J Biol Chem. 1980 Jul 10;255(13):6055-63.
Fibronectin is a major cell-surface glycoprotein which has been reported to interact with glycosaminoglycans. A nitrocellulose filter-binding assay was developed to quantitate these interactions at physiological pH and ionic strength. Fibronectin isolated from chick embryo fibroblasts binds both hyaluronic acid and heparin; heparan sulfate is bound less efficiently, and chondroitin sulfate and glycopeptides are bound minimally. The binding of hyaluronic acid and heparin to fibronectin is saturable and reversible and occurs at separate binding sites. The binding of both molecules to fibronectin is not blocked by EDTA or by other glycosaminoglycans, and is only moderately inhibited by elevated ionic strength. Scatchard analyses revealed nonlinear, high affinity binding to fibronectin with a KD of approximately 10(-7) to 10(-8) M for these glycosaminoglycans. The affinity for heparin was utilized for the isolation of heparin-binding domains of fibronectin on heparin-agarose affinity columns. Heparin-binding proteolytic fragments with apparent molecular weights of 160,000 and 50,000 were isolated following hydrolysis of fibronectin by chymotrypsin or pronase, respectively. The possible involvement of such high affinity binding sites of fibronectin in the binding of glycosaminoglycans to the cell surface or in the organization of extracellular matrices is discussed.
纤连蛋白是一种主要的细胞表面糖蛋白，据报道它可与糖胺聚糖相互作用。已开发出一种硝酸纤维素滤膜结合测定法，用于在生理pH值和离子强度下定量这些相互作用。从鸡胚成纤维细胞中分离出的纤连蛋白可结合透明质酸和肝素；硫酸乙酰肝素的结合效率较低，而硫酸软骨素和糖肽的结合最少。透明质酸和肝素与纤连蛋白的结合是可饱和且可逆的，并且发生在不同的结合位点。这两种分子与纤连蛋白的结合不会被EDTA或其他糖胺聚糖阻断，仅在离子强度升高时受到适度抑制。Scatchard分析显示，这些糖胺聚糖与纤连蛋白呈非线性、高亲和力结合，其解离常数（KD）约为10^(-7)至10^(-8) M。利用对肝素的亲和力，在肝素-琼脂糖亲和柱上分离纤连蛋白的肝素结合结构域。分别用胰凝乳蛋白酶或链霉蛋白酶水解纤连蛋白后，分离出表观分子量为160,000和50,000的肝素结合蛋白水解片段。本文讨论了纤连蛋白的这种高亲和力结合位点在糖胺聚糖与细胞表面结合或细胞外基质组织中的可能作用。
