Transforming growth factor‐β1 binds to immobilized fibronectin

Abstract

We have characterized the interaction of homodimeric porcine transforming growth factor-β1 (TGF-β1) with affinity-purified human plasma fibronectin. Using a solid-phase binding assay, we have demonstrated that TGF-β1 binds to fibronectin immobilized on Immunlon I^TM microtiter plates. TGF-β1 binding increased with time, reaching a plateau after 4–6 h, and was dependent upon the concentration of both labeled TGF-β1 and immobilized fibronectin present. The binding of radiolabeled TGF-β1 to fibronectin was saturable and was reduced 75% in the presence of a 100-fold excess of unlabeled TGF-β1. TGF-β1 bound to fibronectin with an association rate constant (K_a) of 2.96 × 10³ M⁻¹ s⁻¹ and did not readily dissociate under various conditions. The binding of TGF-β1 to fibronection was insensitive to variations in ionic strength over a range of 0.1–1.0 M NaCl and was relatively insensitive to divalent cation concentration in the range of 0.1–10.0 mM as well. These data suggest that the binding of TGF-β1 to fibronectin may not be dependent upon the interaction of charged amino acids within these two molecules. However, the binding of TGF-β1 to fibronectin was strongly pH-dependent and binding decreased dramatically below pH 4.0 and above pH 10.0, suggesting that charged amino acids may influence TGF-β1/fibronectin interactions. The association of TGF-β1 with immobilized fibronectin or other extracellular matrix components and subsequent dissociation under acidic conditions or by an as-yet-unidentified mechanism may play a role in the distribution and/or activity of this potent growth regulator at sites of tissue injury and inflammation in vivo.