Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.21.73 (urokinase-type plasminogen activator)
 10,685 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A method for determining initial velocities of the urokinase (EC 3.4.99.26) catalysed converstion of NH2-terminal lysine plasminogen to plasmin (EC 3.4.21.7) is presented. This reaction has been coupled with the hydrolysis of alpha-N-benzyoly-L-arginine ethyl ester, which is catalysed by plasmin, and its rate has been determined from the time course of the overall reaction. The proenzyme-enzyme conversion was found to obey the Michaelis-Menten rate equation. The following values of the kinetic parameters were obtained: the apparent Michaelis constant, Km = 40.7 +/- 6.2 muM; the catalytic constant, kc = 2.59+/-0.31 s(-1), and kc/Km = 6.36-10(4) +/- 0.24-10(4) M(-1)-s(-1).
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PMID:Kinetic studies of the urokinase catalysed conversion of NH2-terminal lysine plasminogen to plasmin. 13 49

An antiserum was raised in rabbits against human plasmin-antiplasmin complex and rendered specific for neoantigens of this complex by absorption with purified plasminogen and plasma. Polystyrene particles were coated with the specific antibodies and used in an agglutination test for the determination of plasmin-antiplasmin complex in the plasma from various patients. Purified plasmin-antiplasma complex at a concentration of 0.1-0.2 mg/l was found to cause a clear agglutination of the particles. Activation of fresh human plasma with urokinase caused progressive generation of agglutinating activity up to a plasma dilution of 1/480. Intravenous infusion of streptokinase into patients resulted in an increase of the plasmin-antiplasmin titre of at least 1/240. Sera from patients with rheumatoid factor also agglutinated the particles but this activity could be removed by absorbing rheumatoid factor on insolubilized human IgG. Out of 101 male and twenty-three female control subjects, only three men had a plasmin-antiplasmin titre above 1/16. Of 230 hospitalized patients, plasmin-antiplasmin titres of 1/40 or more were detected in twenty-five patients. Most of these patients had diseases which are frequently associated with in vivo coagulation or fibrinolysis, but among them there was only one who showed diffuse intravascular coagulation detectable by classical methods. In the absence of an increased plasmin-antiplasmin titre none of the haemostasis analyses were indicative of in vivo coagulation or fibrinolysis. Seven out of eight patients with diffuse intravascular coagulation of various origin had plasmin-antiplasmin titres of 1/80 or 1/160. Thus, the present latex agglutination test, owing to its simplicity and sensitivity, appears to be a practical routine screening test for detecting fibrinolytic activation in plasma.
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PMID:A latex agglutination test for rapid quantitative estimation of the plasmin-antipalsmin complex in human plasma. 13 93

A simple plasminogen determination method is presented. It is based upon the conversion of plasminogen into activator by large and constant amounts of streptokinase. The activator contained in a standard coagulum consisting of bovine fibrin, streptokinase, and a 1:40 dilution of human plasma converts the plasminogen adsorbed on bovine fibrin into plasmin. Lysis of the test coagulum is hereby induced. The speed of such lysis is limited by the concentration of the activator incorporated in the test coagulum. The variable component of the activator being human plasminogen, the speed of lysis is directly dependent upon the concentration of human plasminogen in the standard coagulum. Using the thromboelastograph according to Hartert in recording the test clot lysis times, this method of plasminogen determination was shown to be a simple and quick procedure. The standard deviation ranged from +/- 13,2 tp 68%, depending upon the plasminogen value to be measured (lower rates of error were attached to high, and higher rates of error to low, plasminogen concentrations). The biological variation of plasminogen values in a group of 26 men aged from 40 to 65 years was calculated to be +/- 21%. Both plasminogen and plasmin, its activated form, were exchangeable in the test, i.e. plasminogen determinations performed by activator assay did not differentiate between plasminogen and plasmin. There was no influence by varying anti-SK titers in the plasma up to a circulating antibody content of 2 million. Furthermore, plasma antiplasmins did not affect the plasminogen measuring system. Plasminogen tested by activator assay displayed values closely related to those achieved by immunochemical methods. Plasminogen measurements were performed in patients undergoing streptokinase and urokinase infusion treatment. 5,000 u streptokinase per hour, as well as 270,000 CTA-u urokinase per hour, infused over a period of 2 days produced a fall in plasminogen down to 30-60% of normal. In contrast, 100,000 u streptokinase per hour lowered the plasminogen concentration down to values of below 1%. The foregoing data indicate that plasminogen measurement, according to the principles outlined here (activator assay), may be regarded as a valuable and reliable method for the routine control of streptokinase and urokinase therapy.
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PMID:On the reliability of plasminogen measurement employing the proactivator-activator converting method. 13 61

A new method is presented for estimating the activator (plasminogen-streptokinase complex) concentration in native plasma of patients undergoing streptokinase infusion. The principle of the method is based on clot lysis time as recorded by the thromboelastograph. The test clot constituents were bovine fibrinogen, bovine plasminogen, EDTA, human plasma (with unknown activator concentrations), and thrombin. In order to obtain a standardization line, urokinase dissolved in NaCl solution was substituted for patients' plasma. Thus, each lysis time could easily be converted into urokinase equivalent (CTA-u/ml). Streptokinase and plasminogen molecules in undiluted patients' plasma were found to exist both in an activator-bound (equimolar plasminogen-streptokinase complex) and in a freely circulating form. This result is in agreement with earlier findings where the activator complex was demonstrated to be a widely dissociated complex in highly diluted plasma of patients, thus displaying an ample proportion of free streptokinase and plasminogen and molecules. Streptokinase treatment using dosage schemes of 100,000 u SK/h, and 200,000 u/h were monitored by quantitative activator, streptokinase, and plasminogen measurements. An average activator concentration of 50-100 CTA-u/ml and a SK-concentration of 7-16 u/ml were recorded during streptokinase infusion. Plasminogen values averaged 0.25%, independent of the amount of streptokinase infused. Each drop in streptokinase was accompanied by a drop in activator during the infusion, and each rise in streptokinase by a rise in activator. There was a strong correlation between streptokinase and activator concentrations in that, on the average, 1 u streptokinase equalled 8.4 CTA-u/ml activator (correlation coefficient r = 0.9) It is concluded that the activator concentration in the plasma of patients undergoing fibrinolytic treatment can easily be adjusted by regulating the hourly streptokinase influx.
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PMID:Studies on activator formation in human plasma with streptokinase. III. Investigation of activator kinetics in undiluted plasma in terms of urokinase equivalents. 13 62

The heavy polypeptide chains of human Glu-plasmin and human Lys-plasmin have been isolated in native solvents, after partial reduction and carboxymethylation of the corresponding plasmins. Two major forms of each heavy chain can be eluted, after adsorption to Sepharose/lysine, utilizing a gradient of epsilon-aminocaproic acid as the eluant. The elution profile of these heavy chains is practically identical to the elution behavior previously observed for human Glu- and Lys-plasminogen, and human Glu- and Lys-plasmin adsorbed to these columns. Sedimentation velocity analysis of the heavy chain of human Glu-plasmin, in the presence of epsilon-aminocaproic acid, demonstrated that a gross conformational alteration occurs in this peptide accompanying binding of this amino acid. A much smaller conformational alteration occurs under similar circumstances with the human Lys-plasmin heavy chain. We find that the NH2-terminal peptide released in the Glu-plasminogen to Lys-plasminogen and Glu-plasmin to Lys-plasmin conversions is also released in the Glu-plasmin heavy chain to Lys-plasmin heavy chain conversion. This reaction is catalyzed at a significant rate only by plasmin and not by urokinase. Finally, no strong interaction between streptokinase and the isolated plasmin heavy chains is observed.
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PMID:Purification and some properties of the Glu- and Lys-human plasmin heavy chains. 13 7

Initial velocities for the urokinase (EC 3.4.99.26)-catalysed conversion of glutamic acid plasminogen to plasmin (EC 3.4.21.7) have been determined at various urokinase and glutamic acid plasminogen concentrations. As has been found for the corresponding reaction with lysine plasminogen this conversion obeys the Michaelis rate equation. The apparent Michaelis constants are of the same order of magnitude for lysine and glutamic acid plasminogens. The difference in conversion rates for the reactions has been shown to be connected with their having different catalytic constants. The data were analysed according to two reaction schemes, in one of which only one peptide bond is split during the glutamic acid plasminogen-plasmin conversion and in the other of which the cleavage of two peptide bonds with the obligatory formation of an intermediate plasminogen is assumed. The results favour the former.
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PMID:Kinetic studies of the urokinase-catalysed conversion of NH2-terminal glutamic acid plasminogen to plasmin. 13 31

Seven laboratories collaborating in a study of two intermediate purity plasminogen preparations (64/23, 63/6) observed that the amount of activator (urokinase or streptokinase) and the time of activation of plasminogen influenced the amount of plasmin generated. Using casein and a synthetic polypeptide (S-2251) as substrates, the authors subsequently showed that complete activation of plasminogen was difficult to achieve without acitivity losses due to plasmin autodigestion. Comparison of the polypeptide subunits (on SDS electrophoresis) of the various plasminogen activation mixtures with their plasmin activity allowed the conclusion that at maximum generation of plasmin from plasminogen, some plasminogen remains in the form of an inactive plasminogen intermediate (PLG-i).
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PMID:Activation of plasminogen as a feature in its assay. 14 Jan 14

Trypsin, thrombin, fibrinolysin, papain, chymothrypsin and urokinase were immobilized on aminopolystyrene resin by the reaction of diazocoupling. An activation of prothrombin and plasminogen and also hydrolysis of fibrin by immobilized enzymes were studied. The immobilized enzymes hydrolyzed N-benzoyl-1-arginine ethyl ester and L-tyrosine ethyl ester. The only preparation of immobilized thrombin possessed the coagulational activity. After the covalent binding trypsin and plasmin maintained the capacity to cause a fibrinolysis. Immobilized trypsin, plasmin, papain, chymotrypsin and urokinase exhibited the fibrinolytic effect due to convertion of plasminogen into plasmin.
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PMID:[Blood coagulating properties of immobilized proteases]. 14 May 25

Vascular endothelial cells derived from rabbit vena cava and maintained in continuous culture exhibited properties characteristic of the intact endothelium. These cells were used as a model for characterizing the fibrinolytic components specified by the endothelium. Endothelial cells in culture digested radiolabeled fibrinogen. Digestion resulted from the synthesis and secretion of a plasminogen activator. Fibrinolysis was not detected when cells were grown in medium lacking plasminogen, indicating the absence of plasminogen-independent fibrinolytic enzymes. Phorbol-myristate-acetate increased extracellular plasminogen activator activity dramatically. This increase was prevented when actinomycin D or cycloheximide was included in the growth medium, indicating that new gene expression was required for it. Intracellular plasminogen activator could not be detected unless the cell extracts were exposed briefly to mildly acidic conditions. Mixing experiments between acid-treated and untreated extracts suggested that the cells contained a potent, acid-labile inhibitor of fibrinolysis. As little as 10 mug of protein from whole cell extracts inhibited both cell and urokinase-mediated fibrinolysis by more than 70%. Cell fractionation studies localized the inhibitor to the cytosol whereas plasminogen activator activity was restricted to the membrane-rich fraction. This membrane fraction did not require acidification for activity, suggesting that the inhibitor had been removed and that acidification did not activate a plasminogen proactivator. These observations demonstrate that regulation of endothelial fibrinolytic activity is far more complex than had been anticipated and raise several uncertainties in regard to detecting the presence of plasminogen activators in cells and tissues.
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PMID:Synthesis of a fibrinolytic activator and inhibitor by endothelial cells. 14 64

In 20 consecutive rheumatoid arthritis patients, 14 women and 6 men, age 26--76, average 62 years, the concentration of the recently found "primary plasmin inhibitor" and phase proteins was estimated in plasma and synovial fluid. In 12 patients a complex between the inhibitor and plasmin could be demonstrated by crossed immunoelectrophoresis into immunoglobulins against the primary plasmin inhibitor and immunoglobulin against plasminogen. Only free inhibitor was found in corresponding plasma. All plasminogen present in synovial fluid could be activated to plasmin upon addition of urokinase (24 nM/1). In those patients where enzyme-inhibitor complex in synovial fluid was present, a higher concentration of phase proteins in synovial fluid was found, indicating an increased degree of inflammation despite identical scores in the Lansbury clinical index in the two groups. From these experiments it was concluded that the fibrinolytic capacity in rheumatoid synovial fluid is not decreased. It is suggested that the fibrin-like material in synovial tissue and upon the synovial membrane is a poor substrate for plasmin.
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PMID:The primary plasmin inhibitor in rheumatoid synovial fluid. 14 41


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