Heat and autoclave resistance of cell-spreading activity of vitronectin

We have investigated the heat- and autoclave-resistant properties of the cell-spreading activity of vitronectin, a cell-spreading glycoprotein in animal blood plasma. Vitronectin heated at 100 degrees C for 10 min or autoclaved at 121 degrees C at 1.2 kg/cm2 for 20 min retained the same cell-spreading activity as native vitronectin. In contrast, fibronectin and type-I collagen treated in the same way lost their activity almost completely. GRGDSP remarkably inhibited the cell-spreading activity of native, heated and autoclaved vitronectins. GRGESP did not inhibit the activity of native vitronectin, but, unexpectedly, partially inhibited the activity of both heated and autoclaved vitronectins. In SDS-polyacrylamide gel analysis under reducing conditions, vitronectin heated at 100 degrees C migrated mainly as a monomer, but autoclaved vitronectin migrated at both the top and front of the gel instead of at the position of the monomer. The change in molecular size during the heat- and autoclave treatments was partially prevented by adding 10 mM dithiothreitol or 2% 2-mercaptoethanol to the protein solution.