Abstract
To prepare a chemically modified urokinase that does not dissociate into two peptide fragments upon reduction of its disulfide bridge, we cross-linked the enzyme intramolecular!) with various bifunctional imidoesters. The enzyme underwent the intramolecular cross-linking most moderately by the reaction at 4 °C for 5 hr with 3 mW dimethyl suberimidate in 0.1 M potassium phosphate buffer (pH 9.0). The cross-linked urokinase isolated by gel filtration with a yield of 25 % showed a specific activity of 76, 000 International Units/mg protein, which corresponds to 53 % of that of the native enzyme. Although the modified enzyme was similar to the native urokinase in some properties such as the autocatalytic self-digestion and the low affinity to fibrin, it showed higher in vivo and in vitro stabilities than the native one.
