Inhibition of cell surface receptor bound plasmin by α2-macroglobulin

Department of Pathology, University of Virginia Health Sciences Center, Charlottesville 22908.
Journal of Biological Chemistry (Impact Factor: 4.57). 08/1991; 266(19):12329-36.
Source: PubMed


The purpose of this investigation was to characterize the reaction of alpha 2-antiplasmin (alpha 2AP) and alpha 2-macroglobulin (alpha 2M) with human plasmin bound to rat C6 glioma cells and human umbilical vein endothelial cells (HUVECs). Binding of plasmin (0.1 microM) to C6 cells at 4 degrees C did not cause cell detachment, decrease viability or change cell morphology. The KD and Bmax for the binding of diisopropyl phosphoryl plasmin (DIP-plasmin) to C6 cells were 0.9 microM and 2.6 x 10(6) sites/cell. The dissociation rate constants (koff) for 125I-plasmin were 9.7 x 10(-4) and 4.0 x 10(-4) s-1 at 4 degrees C in the presence and absence of 0.3 microM DIP-plasmin, respectively. Similar constants were determined for 125I-plasminogen and 125I-DIP-plasmin. Neither alpha 2AP nor alpha 2M affected the dissociation of DIP-plasmin. C6 cell-associated 125I-plasmin reacted slowly with alpha 2AP; however, the inhibition rate constants exceeded the koff. alpha 2AP-plasmin complex formed after the plasmin dissociated into solution (reaction pathway 1) and by direct reaction of alpha 2AP with cell-associated enzyme (reaction pathway 2). High concentrations of alpha 2AP favored pathway 2. C6 cell-associated plasmin was also protected from inhibition by alpha 2M. While the same pathways were probably involved in this reaction, alpha 2M was less effective than alpha 2AP as an inhibitor of nondissociated plasmin (pathway 2). When C6 cell-bound plasmin reacted with alpha 2AP, alpha 2AP-plasmin complex was recovered primarily in the medium, suggesting dissociation of complexes formed on the cell surface. Plasmin-receptor dissociation and inhibition experiments were performed at 22 degrees and 37 degrees C, confirming the conclusions of the 4 degrees C studies. Comparable results were also obtained using HUVEC cultures. These studies demonstrate that cell-associated plasmin is protected from inhibition by alpha 2M as well as alpha 2AP. At least two reaction pathways may be demonstrated for the inhibition of plasmin that is initially receptor-bound; however, neither pathway is highly effective, accounting for the "plasmin-protective" activity of the cell surface.

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    • "In contrast to the cleavage mechanism by uPA and tPA, streptokinases form a protein complex which activates the enzyme domain of plasminogen (Boxrud and Bock, 2000; Boxrud et al., 2004). Moreover, immobilization of plasmin in fibrin fibers or on surfaces of cells protects the enzyme against inhibition by its major inhibitor the serpin α2 antiplasmin and also against inactivation via several streptokinase variants (Plow et al., 1986; Hall et al., 1991; Cook et al., 2012). The capture of plasminogen to the bacterial surface via non-activating plasminogen binding proteins induces conformational changes, which promote generation of plasmin activity by host-activators, although no protective effect against α2 antiplasmin inhibition is gained (Miles and Plow, 1985; Plow et al., 1986). "
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