PURIFICATION AND STABILITY CHARACTERIZATION OF A CELL REGULATORY SIALOGLYCOPEPTIDE INHIBITOR

Previous attempts to physically separate the cell cycle inhibitory and protease activities in preparations of a purified cell regulatory sia loglycopeptide (CeReS) inhibitor were largely unsuccessful. Gradient e lution of the inhibitor preparation from a DEAE HPLC column separated the cell growth inhibitor from the protease, and the two activities ha ve been shown to be distinct and non-overlapping. The additional purif ication increased the specific biological activity of the CeReS prepar ation by approximately two-fold. The major inhibitory fraction that el uted from the DEAE column was further analyzed by tricine-SDS-PAGE and microbore reverse phase HPLC and shown to be homogeneous in nature. T wo other fractions separated by DEAE HPLC, also devoid of protease act ivity, were shown to be inhibitory to cell proliferation and most like ly represented modified relatives of the CeReS inhibitor. The highly p urified CeReS was chemically characterized for amino acid and carbohyd rate composition and the role of the carbohydrate in cell proliferatio n inhibition, stability, and protease resistance was assessed. (C) 199 5 Wiley-Liss, Inc.