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Acceleration of the thrombin inactivation of single chain urokinase-type plasminogen activator (pro-urokinase) by thrombomodulin

Author: Munk, G.A.W. de · Groeneveld, E. · Rijken, D.C.
Type:article
Date:1991
Institution: Instituut voor Verouderings- en Vaatziekten Onderzoek TNO
Source:Journal of Clinical Investigation, 5, 88, 1680-1684
Identifier: 231411
Keywords: Biology · Endothelial cells · Fetomodulin · Fibrinolysis · Protein C · Thrombolytic therapy · Calcium · Human · Kinetics · Molecular Weight · Receptors, Cell Surface · Receptors, Thrombin · Support, Non-U.S. Gov't · Thrombin · Urinary Plasminogen Activator

Abstract

The in vitro effects of thrombomodulin on the inactivation of single chain urokinase-type plasminogen activator (scu-PA) by thrombin were investigated by incubating scu-PA with varying concentrations of human thrombin, in both the absence and presence of soluble rabbit thrombomodulin. 50% inactivation of scu-PA occurred in 45 min at 160 ng/ml thrombin in the absence of thrombomodulin and at 4.6 ng/ml thrombin in the presence of thrombomodulin. No difference was found in either the absence or the presence of thrombomodulin between the inactivation rates of high molecular weight scu-PA, and a low molecular weight scu-PA which lacked the growth factor and kringle domains. Enzyme kinetic experiments with varying concentrations of scu-PA showed that thrombomodulin decreased the K(m) of thrombin for scu-PA from 7.8 to 0.43 μM and increased the k(cat) from 0.30 to 1.2 s-1, corresponding to a 70-fold increase in the second-order rate constant k(cat)/K(m)·SDS- polyacrylamide gel electrophoresis showed that scu-PA was cleaved into two chains upon inactivation by thrombin, and confirmed the acceleration effect of thrombomodulin on inactivation of scu-PA. Thrombomodulin thus not only has anticoagulant properties but is also antifibrinolytic. The acceleration may imply a new mechanism for the regulation of local plasminogen activator activity on the cell surface.