Abstract
Fibrin interacts with tissue-type plasminogen activator (tPA) via the finger and the kringle 2 domains. Three monoclonal antibodies against tPA, designated MPW3VPA, MPW6VPA, and MPW7VPA, which react with epitopes in the tPA molecule involved in fibrin binding, were characterized. The IgM monoclonal antibody MPW6VPA, directed against an epitope close to the finger and epidermal growth factor domains, stimulated plasminogen activation only in the absence of CNBr-fibrinogen fragments by increasing kcat in a dose-dependent fashion, an effect which was not restricted to the intact molecule. These results suggest that MPW6VPA mimics the initial effect of fibrin bound to the tPA molecule, which results in a change of kcat values. The MPW6VPA effect was reversed by another antibody, MPW3VPA, also directed against epidermal growth factor and finger domains. The latter antibody also inhibited plasminogen activation by tPA in the presence of CNBr-fibrinogen fragments in a dose-dependent, apparently noncompetitive way. No effect of MPW3VPA was seen in the absence of CNBr-fibrinogen fragments. MPW7VPA directed against kringle 2 of tPA inhibited plasminogen activation by tPA only when CNBr-fibrinogen fragments were present. This inhibition was apparently competitive and dose-dependent. These data suggest that MPW3VPA interferes with the first phase of fibrin binding to tPA, whereas MPW7VPA interferes with the second phase of fibrin binding to the tPA molecule via kringle 2, resulting in Km changes.
Highlights
Fibrin interacts with tissue-type plasminogen acti- Fibrin binding to the tPA molecule occurs via the finger vator via the finger and the kringle2 domains. and kringle 2 domains of tPA [4].van Zonneveld et al [5]
CNBr-fibrinogen fragments by increasing katin a dose-dependent fashion, an effect which was not restricted to the intact molecule. These results suggest that MPWGVPA mimics theinitialeffect of fibrin bound to the tPA molecule, which results in a change binds in a first phase to the tPA finger, causing a change in kc, values, and in a second phase to tPA kringle 2 via generated COOH-terminal lysines in the fibrin molecule, resulting in a K, change
Three monoclonal antibodies against fibrin binding domains in the tPAmolecule wereobtained
Summary
Fibrin interacts with tissue-type plasminogen acti- Fibrin binding to the tPA molecule occurs via the finger vator (tPA) via the finger and the kringle2 domains. and kringle 2 domains of tPA [4].van Zonneveld et al [5]. Method [14] and its activity compared with the International Refer-Monoclonal Anti-tPA Antibodies against Fibrin Binding Domains
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