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

Collagen is generally incapable of capturing polypeptides such as growth factors in a specific manner. In this study, we established a collagen-binding 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 the 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 fibronectin. FNCBD-EGF substantially stimulated cell growth after binding to collagen-coated culture plates, whereas EG;F had no effect, indicating that this 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 epidermis was observed underneath the sponges, These results suggested that our fusion protein with high collagen affinity would be useful for wound healing.