The recombinant Escherichia coli-derived protease domain of tissue-type plasminogen activator is a potent and fibrin specific fibrinolytic agent

The protease domain of human tissue-type plasminogen activator is expressed in Escherichia coli as inclusion bodies. Subsequently it is converted into its active form by an in vitro folding process. In this study we compare the in vitro activity and the specificity in a plasma milieu of the protease domain with that of the recombinant tissue-type plasminogen activator isolated from Chinese hamster ovary cells. The amidolytic activity of the single chain and the two chain form of the protease is comparable with recombinant tissue-type plasminogen activator thus indicating that the active site of both enzymes is similar. The plasmin forming activity and the activity in a static fibrin clot lysis assay of the protease domain is reduced by a factor of 240 and 10, respectively, as compared to recombinant tissue-type plasminogen activator. This may be a consequence of the missing affinity to fibrin. In contrast, the protease is equipotent and at high concentrations even more potent than the recombinant tissue-type plasminogen activator in a dynamic plasma model. Furthermore, the in vitro analysis of several coagulation parameters revealed no significant differences between the protease domain and the recombinant tissue-type plasminogen activator from Chinese hamster ovary cells. In conclusion, our data demonstrate that in plasma the protease domain is a potent plasminogen activator with a similar fibrin specificity as CHO-t-PA.