STRUCTURE-ACTIVITY-RELATIONSHIPS OF HUMAN EPIDERMAL GROWTH-FACTOR (H-EGF)

The 53 amino acid regulatory peptide, human epidermal growth factor (h-EGF), is a potent mitogen that stimulates cellular proliferation and differentiation in a wide variety of cells. To identify the critical residues that elicit the biological activity of h-EGF, peptides were constructed by stepwise solid-phase synthesis using the Boc-HF strategy. These synthetic peptides were characterized by HPLC, FAB-MS, amino acid analysis and thermolytic digestion. The mitogenic activity of these h-EGF analogues was determined by the stimulation of [H-3]-thymidine uptake into DNA in NIH-3T3 fibroblast cell lines. Substituting Tyr with Phe at position's 37 and 13 had little effect on the mitogenic activity of h-EGF. In contrast, Ala at these positions resulted in a severe loss of activity (20 and 10(3)-fold). These results indicate that the hydrophobicity of the side chain at positions 13 and 37 of h-EGF is essential for its biological activity. A semiconservative substitution of Leu with Ala at position 15 and a conservative change of Lys at position 41 also drastically reduced mitogenic activity (10(4) and 10(5)-fold). Thus, the bulky hydrophobic side chain at position 15 and the guanidino group at position 41 are indispensable in determining the biological activity of h-EGF.