Proteolytic activation of purified human procarboxypeptidase U

Carboxypeptidase U (CPU, EC 3.4.17.20) is a recently described basic carboxypeptidase which circulates in plasma as an enzymatically inactive precursor procarboxypeptidase U (proCPU), also known as plasma carboxypeptidase B precursor or thrombin activatable fibrinolysis inhibitor (TAFI). The activation of the zymogen proceeds through a proteolytic cleavage at Arg-92. The active form - CPU - is able to retard the initial phase of fibrinolysis by cleaving C-terminal lysine residues exposed on fibrin partially degraded by the action of plasmin. These C-terminal lysine residues are essential for the high affinity binding of plasminogen to fibrin and the subsequent activation to plasmin. In this report, the activation of purified human proCPU was studied using trypsin and some key proteases of the coagulation and fibrinolytic cascade, i.e., kallikrein, plasmin and thrombin. The most efficient activation is obtained in the presence of thrombin in complex with thrombomodulin. After in vitro activation, CPU is unstable at 37 degrees C (T(1/2)=15 min). Its stability can be improved dramatically using lower temperatures.