A monoclonal antibody directed against the high-affinity lysine-binding site (LBS) of human plasminogen. Role of LBS in the regulation of fibrinolysis.
One of thirty murine monoclonal antibodies, raised by immunization with human plasmin-alpha 2-antiplasmin complex, was found to be directed against the high-affinity lysine-binding site in plasminogen. Indeed, this antibody (MA-HAL) reacted with plasminogen and with a fragment of plasminogen composed of the first three triple-loop structures (LBS I) and was displaced by 6-aminohexanoic acid (50% displacement at 25 microM). In competitive radioimmunoassays the binding of radiolabeled plasminogen to MA-HAL was reduced to 50% with 2.3 microM alpha 2-antiplasmin or 1.3 microM histidine-rich glycoprotein, which corresponds to the known dissociation constants between these ligands and the high-affinity lysine-binding site of plasminogen. MA-HAL did not influence the activation of plasminogen by tissue-type plasminogen activator in the absence of CNBr-digested fibrinogen, but abolished the effect of CNBr-digested fibrinogen on the Michaelis constant of the reaction. MA-HAL reduced the reaction rate between plasmin and alpha 2-antiplasmin by a factor 20 and abolished the binding of plasminogen to fibrin. These results indicate that MA-HAL specifically binds to and masks the high-affinity lysine-binding site of plasminogen. It therefore is a useful tool for the investigation of the role of this structure in the regulation of fibrinolysis, both at the level of fibrin-stimulated activation of plasminogen and of the inhibition of generated plasmin.
D000818:Animals; D000911:Antibodies, Monoclonal; D001665:Binding Sites; D001667:Binding, Competitive; D005337:Fibrin / Q000378:metabolism; D005341:Fibrinolysin / Q000378:metabolism; D005342:Fibrinolysis; D008239:Lysine / Q000378:metabolism*; D008433:Mathematics; D051379:Mice; D008807:Mice, Inbred BALB C; D010958:Plasminogen / Q000276:immunology / Q000378:metabolism*; D011863:Radioimmunoassay; D010959:Tissue Plasminogen Activator / Q000494:pharmacology; D000979:alpha-2-Antiplasmin / Q000378:metabolism
D000911:Antibodies, Monoclonal; D000979:alpha-2-Antiplasmin; D005337:Fibrin; D010958:Plasminogen; D010959:Tissue Plasminogen Activator; D005341:Fibrinolysin; D008239:Lysine