Plasminogen activation by receptor-bound urokinase. A kinetic study with both cell-associated and isolated receptor.
The specific cellular receptor for urokinase-type plasminogen activator (uPA) is found on a variety of cell types and has been postulated to play a central role in the mediation of pericellular proteolytic activity. We have studied the kinetics of plasminogen (Plg) activation catalyzed by uPA specifically bound to its receptor on the human monocytoid cell-line U937 and demonstrate this process to have properties differing widely from those observed for uPA in solution. The solution-phase reaction was characterized by a Km of 25 microM and for the cell-associated reaction this fell 40-fold to 0.67 microM, below the physiological Plg concentration of 2 microM. A concomitant 6-fold reduction in kcat resulted in an increase in the overall catalytic efficiency, kcat/Km, of 5.7-fold. This high affinity Plg activation was abolished in the presence of a Plg-binding antagonist. In contrast to intact cells, purified uPA receptor (isolated from phorbol 12-myristate 13-acetate-stimulated U937 cells) was observed to partially inhibit uPA-catalyzed Plg activation, although activity against low molecular weight substrates was retained. Therefore, the cellular binding of Plg appears to be of critical importance for the efficient activation of Plg by receptor-bound uPA. Plasmin generated in the cell-surface Plg activation system described here was also observed to be protected from its principal physiological inhibitor alpha-2-antiplasmin. Together, these data demonstrate that the cell surface constitutes the preferential site for Plg activation when uPA is bound to its specific cellular receptor, which therefore has the necessary characteristics to play an efficient role in the generation of pericellular proteolytic activity.
Ellis
V
V
Finsen Laboratory, Rigshospitalet, Copenhagen, Denmark.
Behrendt
N
N
Danø
K
K
D000933:Antifibrinolytic Agents / Q000494:pharmacology; D002384:Catalysis; D002460:Cell Line; D004591:Electrophoresis, Polyacrylamide Gel; D005341:Fibrinolysin / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D010958:Plasminogen / Q000378:metabolism*; D010960:Plasminogen Activators / Q000378:metabolism*; D014568:Urokinase-Type Plasminogen Activator / Q000378:metabolism*
D000933:Antifibrinolytic Agents; D010958:Plasminogen; D010960:Plasminogen Activators; D005341:Fibrinolysin; D014568:Urokinase-Type Plasminogen Activator