Production of a biologically active epidermal growth factor fusion proteinwith high collagen affinity

Collagen is generally incapable of capturing polypeptides such as growth fa ctors in a specific manner. In this study, we established a collagen-bindin g growth factor (FNCBD-EGF) consisting of epidermal growth factor (EGF) and the fibronectin collagen-binding domain. A typical yield of FNCBD-EGF was approximately 200 mug/ml culture in an Escherichia coli expression system. This fusion protein bound to gelatin and fibrillar collagen sponges, and th e bound protein was not effectively eluted even with 2 M NaCl, In addition, FNCBD-EGF bound to type I, II, III, or TV collagen-coated plates, and the specificity of binding was confirmed by competitive inhibition using fibron ectin. FNCBD-EGF substantially stimulated cell growth after binding to coll agen-coated culture plates, whereas EG;F had no effect, indicating that thi s fusion protein acted as a collagen-associated growth factor, In an animal model of impaired wound healing, FNCBD-EGF, but not EGF, was retained with collagen sponges at wound sites 4 d after implantation, and repair of epid ermis was observed underneath the sponges, These results suggested that our fusion protein with high collagen affinity would be useful for wound heali ng.