Aggregated, conformationally changed fibrinogen exposes the stimulating sites for t-PA-catalysed plasminogen activation
article
The present paper shows that conformationally changed fibrinogen can expose the sites Aα-(148-160) and γ-(312-324) involved in stimulation of the tissue-type plasminogen activator (t-PA)-catalysed plasminogen activation. The exposure of the stimulating sites was determined by ELISA using mABs directed to these sites, and was shown to coincide with stimulation of t-PA-catalysed plasminogen activation as assessed in an assay using a chromogenic substrate for plasmin. Gel permeation chromatography of fibrinogen conformationally changed by heat (46.5°C for 25 min) demonstrated the presence of both aggregated and monomeric fibrinogen. The aggregated fibrinogen, but not the monomeric fibrinogen, had exposed the epitopes Aα-(148-160) and γ-(312-324) involved in t-PA stimulation. Fibrinogen subjected to heat in the presence of 3 mM of the tetrapeptide GPRP neither aggregates nor exposes the rate-enhancing sites. Thus, aggregation and exposure of t-PA-stimulating sites in fibrinogen seem to be related phenomena, and it is tempting to believe that the exposure of stimulating sites is a consequence of the conformational changes that occur during aggregation, or self-association. Fibrin monomers kept in a monomeric state by a final GPRP concentration of 3 mM do not expose the epitopes Aα-(148-160) and γ-(312-324) involved in t-PA-stimulation, whereas dilution of GPRP to a concentration that is no longer anti-polymerizing, results in exposure of these sites. Consequently, the exposure of t-PA-stimulating sites in fibrin as well is due to the conformational changes that occur during self-association. Chemicals/CAS: Antibodies, Monoclonal; Biopolymers; Epitopes; Fibrinogen, 9001-32-5; glycyl-prolyl-arginyl-proline, 67869-62-9; Oligopeptides; Plasminogen, 9001-91-6; Tissue Plasminogen Activator, EC 3.4.21.68
Topics
Gel permeation chromatographyPlasminogen activationProtein aggregationAmino Acid SequenceAnimalsAntibodies, MonoclonalBinding SitesBiopolymersChromatography, GelEnzyme-Linked Immunosorbent AssayEpitopesFibrinogenHeatHumansKineticsMolecular Sequence DataOligopeptidesPlasminogenProtein ConformationRabbitsTissue Plasminogen Activator
TNO Identifier
233211
ISSN
03406245
Source
Thrombosis and Haemostasis, 75(2), pp. 326-331.
Pages
326-331
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