Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.5 (thrombin)
33,306 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In this report we used a fluorescent flow cytometry-based assay to examine plasminogen binding to platelets in plasma. Our data indicate that platelets activated in platelet-rich plasma (PRP) by adenosine-5'-diphosphate (ADP) or thrombin bind plasminogen to their surface. Fab fragments of the monoclonal antibody LJ-CP8 that are directed against the fibrinogen binding site on the glycoprotein (GP) IIb-IIIa complex inhibit both plasminogen and fibrinogen binding to ADP-stimulated platelets as does 5 mmol/L EDTA. Platelet aggregation and plasminogen and fibrinogen binding are also concurrently inhibited by the Gly-Arg-Asp (RGD) analogue Gly-Arg-Gly-Asp-Ser (GRGDS) when it is added to PRP before ADP stimulation. The scrambled peptide analogue SDGRG has no effect. The monoclonal antibody 6D1, directed against the von Willebrand factor binding site on GPIb, has no effect on plasminogen-platelet binding, nor does antithrombospondin antibody. epsilon-Aminocaproic acid (EACA), however, inhibits plasminogen binding to ADP-activated platelets. These data indicate that plasminogen binds to platelets activated in plasma, that binding occurs on platelet GPIIb/IIIa, and that binding may be mediated via plasminogen association with fibrinogen via lysine binding domains. Finally, we found both plasminogen and fibrinogen on resting platelets in PRP and demonstrated that they are equally displaced by EDTA, LJ-CP8, and 10E5 (an additional anti-GPIIb/IIIa monoclonal antibody). Plasminogen is also equally displaced by EACA. These data suggest that plasminogen is also bound to GPIIb/IIIa on resting platelets, possibly also via interaction with fibrinogen.
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PMID:Plasminogen interactions with platelets in plasma. 317 39

Plasminogen binding sites are expressed by a wide variety of cell types and serve to promote fibrinolysis and local proteolysis. In this study, the recognition specificity of cells for plasminogen has been examined, primarily using platelets as models. Analyses with plasminogen fragments implicated residues 79-337 (or 353), comprising the first three kringles of plasminogen, as a primary recognition site for plasminogen binding to both thrombin-stimulated and nonstimulated platelets. Other regions of plasminogen, namely residues 354-439 and 442-790, can also participate in the interaction, and these other regions contribute differentially to the binding of the ligand to stimulated and nonstimulated platelets. Binding to nucleated cells, with U937 cells serving as the prototype, is dependent upon a recognition specificity similar to that of unstimulated platelets. Binding of Glu-plasminogen, the native form of the molecule, to thrombin-stimulated platelets has been shown previously to require platelet fibrin. By comparing the interaction of Glu-plasminogen and its degradation product, Lys-plasminogen, with thrombin-stimulated platelets, it is concluded that the cell surface uniquely enhances the affinity of Glu-, but not Lys-plasminogen, for fibrin. Finally, we have demonstrated that cellular receptors and interactive sites within plasminogen are available in the plasma environment. Thus, the functions ascribed to cellular plasminogen receptors can occur within a physiologic setting.
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PMID:The cell-binding domains of plasminogen and their function in plasma. 340 57

The investigation of many hemostatic defects in the newborn is limited by the lack of normal reference values. This study was designed to determine the postnatal development of the human coagulation system in the healthy full-term infant. Consecutive mothers of healthy full-term infants born at St Joseph's Hospital in the city of Hamilton were approached for consent. One hundred eighteen full-term infants (37 to 42 weeks' gestational age) were entered into the study. Demographic information and a 2-mL blood sample were obtained in the postnatal period on days 1, 5, 30, 90, and 180. Between 40 and 79 full-term infants were studied on each day for each of the coagulation tests. Plasma was fractionated and stored at -70 degrees C for batch assaying of the following tests: prothrombin time, activated partial thromboplastin time, thrombin clotting time, and factor assays (biologic): fibrinogen, II, V, VII, VIII, IX, X, XI, XII, and high-molecular weight kininogen. Factor XIII subunits A and S, von Willebrand factor, and the inhibitors antithrombin III, alpha 2-antiplasmin, alpha 2-macroglobulin, alpha 1-antitrypsin, C1 esterase inhibitor, protein C, and protein S were measured immunologically. Plasminogen, prekallikrein, and heparin cofactor II were measured by using chromogenic substrates. The large number of infants studied at each time point allowed us to determine the following: the range of normal for each test at five time points in the postnatal period; that coagulation tests vary with the postnatal age of the infant; that different coagulation factors show different postnatal patterns of maturation; and that near-adult values are achieved for most components by 6 months of life. In summary, this large cohort of infants studied consecutively in the postnatal period allowed us to determine the normal development of the human coagulation system in the full-term infant.
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PMID:Development of the human coagulation system in the full-term infant. 359 64

A mechanism by which platelets might participate in fibrinolysis by binding plasminogen and influencing its activation has been examined. Binding of radioiodinated human Glu-plasminogen to washed human platelets was time-dependent and was enhanced 3-9-fold by stimulation of platelets with thrombin but not with ADP. The interaction with both stimulated and unstimulated cells was specific, saturable, divalent ion-independent, and reversible. The platelet-bound ligand had the molecular weight of plasminogen, and no conversion to plasmin was detected. Scatchard analyses provided evidence for a single class of plasminogen-binding sites on both stimulated and unstimulated cells. The Kd for thrombin-stimulated platelets was 2.6 +/- 1.3 microM, and 190,000 +/- 45,000 molecules were bound per cell, whereas unstimulated platelets bound 37,000 +/- 10,500 molecules/cell with a Kd of 1.9 +/- 0.15 microM. Plasminogen binding was inhibited in a dose-dependent manner by omega-aminocarboxylic acids at concentrations consistent with a requirement for an unoccupied high affinity lysine-binding site for plasminogen binding to the cells. When platelet-bound plasminogen was incubated with tissue plasminogen activator, urokinase, or streptokinase, gel analysis established that plasmin was preferentially associated with the platelet relative to the supernatant. Plasminogen and plasmin interacted with thrombin-stimulated platelets with similar binding characteristics, and there was no evidence for a binding site for plasmin which did not also bind plasminogen. Therefore, the results suggest that plasminogen activation is enhanced on the cell surface. In sum, these results indicate that platelets bind plasminogen at physiologic zymogen concentrations and this interaction may serve to localize and promote plasminogen activation.
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PMID:Binding and activation of plasminogen on the platelet surface. 392 Feb 16

In an earlier article we have reported a technique which allows the measurement of haemostasis from whole blood by the "tube bleeding" principle. The technique is now progressed by using non-anticoagulated blood and therefore clotting-time is also measured. We report here that the technique allows a simple thrombolytic assay to perform from the same blood sample used for haemostasis measurement by detecting the length of time until the haemostatic plugs crack or are expelled. Evidence is presented that this "spontaneous" expulsion is mainly due to thrombolysis. Generation of thrombin during haemostasis is indicated by the inhibitory effect of hirudin. 125I-fibrinogen accumulates (28-fold) in the haemostatic plugs. Plasminogen activators (streptokinase and urokinase) reduce while inactivation of plasminogen by specific antibody greatly prolongs the expulsion-time. The effect of cytochalasin B shows that the retraction of the haemostatic plug is not a decisive factor of the expulsion. As the technique allows the simultaneous measurement of haemostasis, thrombolysis and clotting, it seems ideal for monitoring thrombolytic therapy.
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PMID:A new, ideal technique to monitor thrombolytic therapy. 395 59

Urokinase-activated human plasma was analysed by acetic acid/urea/polyacrylamide-gel electrophoresis. The bands representing plasminogen, the plasmin-alpha 2-plasmin inhibitor and plasmin-alpha 2-macroglobulin complexes were identified by immunoprecipitation with specific antibodies and by comparison with purified components. Plasminogen and the plasmin-inhibitor complexes were isolated from plasma or thrombin-clotted plasma containing 125I-labelled Glu-plasminogen (residues 1-790) and urokinase. The plasma was kept at 37 degrees C for 0.5 and 10 times the lysis time of the clotted plasma, the clotted plasma until lysis. The plasmin heavy chain from the plasmin-inhibitor complexes was subsequently prepared. Only in one case could a low-grade proteolytic conversion of Glu- forms into Lys/Met/Val-forms (residues 77-790, 68-790 and 78-790 respectively) during the preparations be detected. Sodium dodecyl sulphate/polyacrylamide-gel electrophoresis and N-terminal sequence analysis of the purified plasminogen and plasmin heavy chain showed the following. The plasminogen in plasma was on the Glu- form. Glu-plasmin constituted 0.74 and 0.58 of the plasmin bound to the alpha 2-plasmin inhibitor in plasma after brief and prolonged activation respectively. The rest was Lys/Met/Val-plasmin. The clotted plasma contained about equal amounts of Glu-plasminogen and Lys/Met/Val-plasminogen, and predominantly Lys/Met/Val-plasmin complexed to alpha 2-plasmin inhibitor and alpha 2-macroglobulin. The results of the analysis of the purified material substantiated the identity of radioactive protein bands in the gel after acetic acid/urea/polyacrylamide-gel electrophoresis.
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PMID:Identification of molecular forms of plasminogen and plasmin-inhibitor complexes in urokinase-activated human plasma. 620 93

Coagulation and fibrinolysis profiles of naturally menopausal women receiving conjugated estrogens (0.625 or 1.25 mg for 21 of 28 days) and medroxyprogesterone acetate (10 mg for seven of 28 days) for 18 months were compared with those of similar women receiving no hormone therapy. Tests indicative of the dynamics of the coagulation cascade, ongoing intravascular coagulation, and anticoagulation were performed. Hormone therapy had no effect on prothrombin times, activated partial thromboplastin times, or thrombin times. There was no evidence of intravascular coagulation in any of the groups as assessed by platelet counts, fibrinogen antigen and activity, and fibrin degradation products. Antithrombin III antigen and activity, alpha 1-antitrypsin antigen, and alpha 2-macroglobulin antigen, the natural inhibitors of coagulation, were also unaffected by hormone therapy. Plasminogen antigen levels were unaffected, but plasminogen activity was enhanced in the hormone-treated groups, suggesting a stimulatory effect on fibrinolysis. These data indicate that in terms of the coagulation system, healthy women can safely use a combined regimen of conjugated estrogens and medroxyprogesterone acetate.
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PMID:Combination estrogen and progestogen replacement therapy does not adversely affect coagulation. 662 49

We prospectively studied the hemostatic system of ten persons bitten by the Eastern diamondback rattlesnake (Crotalus adamanteus) during 1978-1980. Blood was drawn when the patients arrived in the emergency room and every 6 hr thereafter. All envenomated victims developed incoagulable blood (defined by a thrombin time greater than or equal to 120 sec, normal less than 20 sec). Platelet counts and plasma levels of antithrombin III and factors II and VIII were not drastically altered, which distinguished this disorder from classic disseminated intravascular coagulation. Fibrinogen levels were markedly decreased (mean coagulable level of 0 mg/dl and antigenic levels of 99 mg/dl). Plasminogen levels were 20% of normal, alpha-2-plasminogen inhibitor was 17% of normal, and plasminogen activator was 20 times normal. Levels of fibrin degradation products peaked at a mean of 7,680 micrograms/ml. The magnitude and duration of the coagulopathy were proportional to the clinical severity of envenomation. Treatment with antivenin blunted the coagulopathy. Because venom from the Eastern diamondback rattlesnake does not directly activate plasminogen, we conclude that coagulopathy following envenomation by that reptile appears to be due to partial proteolysis of fibrinogen with secondary activation of plasminogen by released plasminogen activator, probably of endothelial origin.
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PMID:Mechanism of defibrination in humans after envenomation by the Eastern diamondback rattlesnake. 685 33

Screening coagulation tests, coagulation factors, and components of fibrinolysis and kinin generation were examined in 21 healthy adult cats. Observed ranges for screening tests were: prothrombin time, 7.3 to 11.4 s; activated partial thromboplastin time, 10.6 to 14.9 s; and thrombin time, 10.7 to 18.9 s. Functional coagulation factors II, V, VII, VIII, IX, X, XI, and XII were assayed and expressed as percentage of normal. Although individual factors varied, the observed range for factors assayed was 37% to 208% of normal. Fibrinogen ranged from 50 to 165 mg/dl. Plasminogen and antithrombin III ranged from 50% to 200% and 89% to 111% of normal, respectively. Plasma kallikrein ranged from 0.3 to 3.9 mukat/L. Fibrin(ogen) degradation products and fibrin monomers were examined with variable and inconsistent results.
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PMID:Coagulation, fibrinolysis, and kinin generation in adult cats. 710 32

Plasminogen activator inhibitor-1 (PAI-1), an important risk factor for thrombotic diseases, is a member of the superfamily of serine proteinase inhibitors. To define structural rearrangements occurring during interaction between PAI-1 and its target proteinases we have raised monoclonal antibodies against the PAI-1/t-PA complex. Thirteen out of 401 monoclonal antibodies reacted preferentially with the PAI-1/t-PA complex as compared to free PAI-1 or free t-PA. Detailed characterization revealed the presence of two non-overlapping neoantigenic epitopes in the PAI-1/t-PA complex. Both neoantigenic epitopes were also exposed after complex formation between PAI-1 and either urokinase-type plasminogen activator, plasmin or thrombin as well as after cleavage of the reactive site loop of non-inhibitory substrate type PAI-1 variants. Thus, we have identified two neoantigenic epitopes, localized entirely in PAI-1, and commonly exposed after complex formation of active PAI-1 with various proteinases or after cleavage of substrate PAI-1. These monoclonal antibodies should facilitate further studies on the mechanism of interaction between various PAI-1 forms and its target proteinases.
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PMID:Characterization of common neoantigenic epitopes generated in plasminogen activator inhibitor-1 after cleavage of the reactive center loop or after complex formation with various serine proteinases. 749 51


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