AQUEOUS STABILITY OF HUMAN EPIDERMAL GROWTH-FACTOR-1-48

Human epidermal growth factor 1-48 (hEGF 1-48, Des(49-53)hEGF) is a si ngle chain polypeptide (48 amino acids; 3 disulfide bonds; 5445 Da) po ssessing a broad spectrum of biologic activity including the stimulati on of cell proliferation and tissue growth. In this study, three prima ry aqueous degradation products of hEGF 1-48 were isolated using isocr atic, reverse phase/ion-pair HPLC. The degradation products were chara cterized using amino acid sequencing, electrospray ionization mass spe ctrometry, isoelectric focusing, and degradation kinetics. Results ind icate that hEGF 1-48 degrades via oxidation (Met(21)), deamidation (As n(1)), and succinimide formation (Asp(11)). The relative contribution of each degradation pathway to the overall stability of hEGF 1-48 chan ges as a function of solution pH and storage condition. Succinimide fo rmation at Asp(11) is favored at pH < 6 in which aspartic acid is pres ent mostly in its protonated form. Deamidation of Asn(1) is favored at pH > 6. The relative contribution of Met(21) oxidation is increased w ith decreasing temperature, storage as a frozen solution (-20 degrees C), and exposure to fluorescent light.