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Query: UNIPROT:P00750 (PLA)
 16,800 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Low molecular weight two-chain urokinase is a 33-kD plasminogen activator, which has no innate affinity for fibrin and consequently, its use to facilitate lysis of blood clots may lead to systemic activation of plasminogen. In order to impart clot affinities to this urokinase form (UK) we have generated two novel fibrin-binding derivatives by partially reducing UK and exchanging the native disulfide-linked peptide A with peptide A analogs. The peptide A analogs contained the fibrin-adherent fibrin-derived sequences, GPRP (derived from positions 17-20 of the fibrinogen alpha chain) or QAGDV (407-411 sequence of the fibrinogen gamma chain), each coupled through amino-hexanoic acid to a synthetic peptide, LKFQCGQK, containing the Leu 144-Lys 158 sequence of the urinary plasminogen activator A Chain. The resultant derivatives contained about 0.4 moles peptide analog/mole UK, were 75% active toward synthetic UK substrates, and were recovered in a nearly 80% yield. The two fibrin peptide derivatives had a five-fold greater affinity for the clots.
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PMID:Insertion of fibrin peptides into urokinase enhances fibrin affinity. 235 45

A chimeric plasminogen activator, GPRP-u-PA (144-411), consisting of the Gly-Pro-Arg-Pro tetrapeptide fused to the N-terminal of a truncated urokinase-type plasminogen activator (comprising Leu 144 through Leu 411), was produced by expression of the corresponding chimeric cDNA in Escherichia coli cells. After renaturation, the chimera was purified to homogeneity with specific amidolytic activity of 100,000 IU/mg protein. The chimera showed 6-fold greater affinity for fibrin clots than native low molecular weight urokinase (LUK) and 1.5-fold greater affinity than a chemical conjugate, GPRP-LUK, generating via coupling Gly-Pro-Arg-Pro tetrapeptide to native low molecular weight urokinase. The chimera had 2 to 3 fold greater fibrinolytic potency than native LUK in vitro. Fibrinogen had no influence on fibrinolysis of the chimera. The chimera consumed much less fibrinogen than native LUK.
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PMID:Characterization of a recombinant chimeric plasminogen activator composed of Gly-Pro-Arg-Pro tetrapeptide and truncated urokinase-type plasminogen activator expressed in Escherichia coli. 867 Feb 47

The present paper shows that conformationally changed fibrinogen can expose the sites A alpha-(148-160) and gamma-(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 degrees C for 25 min) demonstrated the presence of both aggregated and monomeric fibrinogen. The aggregated fibrinogen, but not the monomeric fibrinogen, has exposed the epitopes A alpha-(148-160) and gamma-(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 A alpha-(148-160) and gamma-(312-324) involved in t-PA-stimulation, whereas dilution of GPRP to a concentration that is not 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.
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PMID:Aggregated, conformationally changed fibrinogen exposes the stimulating sites for t-PA-catalysed plasminogen activation. 881 85

The interaction of lipoprotein(a) [Lp(a)] with platelets is not well defined, particularly with regards to the individual contribution of the protein components of Lp(a), the apo B-100 and the apolipoprotein apo(a). This study investigated the binding of different recombinant apo(a) [r-apo(a)] isoforms, to human platelets and its effect on platelet aggregation. Scatchard analysis of saturation binding experiments demonstrated that human platelets display a single class of high affinity r-apo(a) binding sites (71 +/- 46 molec./platelet, Kd = 5.6 +/- 2.0 nmol/L). Platelet activation with strong agonists (thrombin, arachidonic acid) increased 2- to 10-fold the r-apo(a) binding, without affecting the affinity. Competition assays showed that the binding sites are highly specific for r-apo(a) and Lp(a). At high concentration t-PA could also bind to the r-apo(a) binding sites. By contrast, neither fibrinogen nor plasminogen inhibited to the r-apo(a) binding. The lysine analogue EACA inhibits the binding of r-apo(a) to platelets, thus suggesting the involvement of lysine residues in that interaction. Moreover, the r-apo(a) binding to platelets is unlikely mediated by GPIIb/IIIa-attached fibrin since it is not affected by platelet treatment with either LJ-CP8, a monoclonal antibody that specifically blocks fibrinogen binding to GPIIb/IIIa, nor GPRP, an inhibitor of fibrin polymerisation. Finally, we show that the distinct recombinant apo(a) proteins, as well as native Lp(a), promote an aggregation response of platelets to otherwise subaggregant doses of arachidonic acid. This proaggregant effect of r-apo(a) is dependent on its binding to platelets since it requires a minimum incubation time, and it is prevented by EACA at concentration inhibiting the r-apo(a)-platelet interaction. These results suggest that the prothrombotic action of Lp(a) may be in part mediated by modulating the platelet function through the interaction of its apo(a) subunit with a specific receptor at the platelet surface.
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PMID:Binding of recombinant apolipoprotein(a) to human platelets and effect on platelet aggregation. 1134 6

Myosin modulates the fibrinolytic process as a cofactor of the tissue plasminogen activator and as a substrate of plasmin. We report now that myosin is present in arterial thrombi and it forms reversible noncovalent complexes with fibrinogen and fibrin with equilibrium dissociation constants in the micromolar range (1.70 and 0.94 microM, respectively). Competition studies using a peptide inhibitor of fibrin polymerization (glycl-prolyl-arginyl-proline [GPRP]) indicate that myosin interacts with domains common in fibrinogen and fibrin and this interaction is independent of the GPRP-binding polymerization site in the fibrinogen molecule. An association rate constant of 1.81 x 10(2) M(-1) x s(-1) and a dissociation rate constant of 3.07 x 10(-4) s(-1) are determined for the fibrinogen-myosin interaction. Surface plasmon resonance studies indicate that fibrin serves as a matrix core for myosin aggregation. The fibrin clots equilibrated with myosin are stabilized against dissolution initiated by plasminogen and tissue-type plasminogen activator (tPA) or urokinase (at fibrin monomer-myosin molar ratio as high as 30) and by plasmin under static and flow conditions (at fibrin monomer-myosin molar ratio lower than 15). Myosin exerts similar effects on the tPA-induced dissolution of blood plasma clots. Covalent modification involving factor XIIIa does not contribute to this stabilizing effect; myosin is not covalently attached to the clot by the time of complete cross-linking of fibrin. Thus, our in vitro data suggest that myosin detected in arterial thrombi binds to the polymerized fibrin, in the bound form its tPA-cofactor properties are masked, and the myosin fibrin clot is relatively resistant to plasmin.
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PMID:Myosin: a noncovalent stabilizer of fibrin in the process of clot dissolution. 1254 59

Streptococcus agalactiae is an etiological agent of several infective diseases in humans. We previously demonstrated that FbsA, a fibrinogen-binding protein expressed by this bacterium, elicits a fibrinogen-dependent aggregation of platelets. In the present communication, we show that the binding of FbsA to fibrinogen is specific and saturable, and that the FbsA-binding site resides in the D region of fibrinogen. In accordance with the repetitive nature of the protein, we found that FbsA contains multiple binding sites for fibrinogen. By using several biophysical methods, we provide evidence that the addition of FbsA induces extensive fibrinogen aggregation and has noticeable effects on thrombin-catalyzed fibrin clot formation. Fibrinogen aggregation was also found to depend on FbsA concentration and on the number of FbsA repeat units. Scanning electron microscopy evidentiated that, while fibrin clot is made of a fine fibrillar network, FbsA-induced Fbg aggregates consist of thicker fibers organized in a cage-like structure. The structural difference of the two structures was further indicated by the diverse immunological reactivity and capability to bind tissue-type plasminogen activator or plasminogen. The mechanisms of FbsA-induced fibrinogen aggregation and fibrin polymerization followed distinct pathways since Fbg assembly was not inhibited by GPRP, a specific inhibitor of fibrin polymerization. This finding was supported by the different sensitivity of the aggregates to the disruptive effects of urea and guanidine hydrochloride. We suggest that FbsA and fibrinogen play complementary roles in contributing to thrombogenesis associated with S. agalactiae infection.
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PMID:Multiple interactions of FbsA, a surface protein from Streptococcus agalactiae, with fibrinogen: affinity, stoichiometry, and structural characterization. 1704 2