Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.21.7 (plasmin)
 9,023 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have previously reported paradoxical prothrombotic effects manifest by elevations of fibrinopeptide A (FPA) after administration of streptokinase to patients with acute myocardial infarction. To characterize mechanisms responsible and their dependence on streptokinase (SK) as opposed to other activators of the fibrinolytic system, the present study was performed to compare effects of streptokinase, tissue-type plasminogen activator (t-PA), and urokinase on plasma and on purified prothrombin in concentrations similar to those achieved pharmacologically. The effects of plasmin were assessed to determine the extent to which the elevations of FPA seen could be attributed to activation of plasminogen. Elevations of FPA were observed after incubation of each of the activators with citrated plasma at 37 degrees C for 60 minutes. However, they were most marked with streptokinase (64.5 +/- 4.6 pmol FPA/ml (mean +/- SE) with 100 IU SK/ml, and 77.6 +/- 5.0 pmol/ml with 500 IU SK/ml). Elevations of FPA induced by streptokinase were attenuated by 100 IU/ml heparin [15.2 +/- 1.9 pmol/ml after 100 IU of SK (p less than 0.001 compared with results with streptokinase without heparin)]. Human plasmin, 2.5 CTA/ml, caused changes similar to those induced by streptokinase. The minimal elevations of FPA induced by t-PA or urokinase (less than 10 pmol/ml without heparin) were not significantly attenuated by heparin. Incubation of barium citrate adsorbed plasma (vitamin K factor depleted) with streptokinase markedly attenuated elevation of FPA. Addition of prothrombin (1.5 microM) and streptokinase (100 IU/ml) to the barium citrate adsorbed plasma elicited elevations of FPA similar to those induced by streptokinase in citrated plasma. Amidolytic activity with the "thrombin" substrate H-D-phenylalanyl-L-pipecolyl-L-arginine-p-nitroanilide dihydrochloride (S-2238) was evident when streptokinase, plasminogen (0.24 microM), and prothrombin (1.5 microM) were incubated in buffer. Thus, concentrations of streptokinase that are low in terms of therapeutic blood levels activate prothrombin in plasma, likely due to activation of plasminogen. Neither tissue-type plasminogen activator nor urokinase in pharmacologically comparable concentrations increase thrombin activity appreciably perhaps because of less intense activation of plasminogen. Consideration of the prothrombotic effects observed may be relevant to selection of specific agents for therapeutic thrombolysis, to appropriate titration of dose, and to the need for the use of heparin conjointly with particular activators of the fibrinolytic system such as streptokinase.
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PMID:Differential effects of activation of prothrombin by streptokinase compared with urokinase and tissue-type plasminogen activator (t-PA). 313 87

We report the isolation of a specific protease zymogen from chicken plasma. The purification procedure involves barium citrate precipitation, ammonium sulfate fractionation, removal of plasminogen and plasmin on lysine-Sepharose, followed by anion and cation exchange, and gel permeation chromatography. Based on quantitative radioimmunoassay the zymogen is present in plasma at a concentration of 160 mg/liter, and it is obtained by our procedure in highly purified form with a yield of 1.4%. The single polypeptide chain contains an NH2-terminal alanine residue. The native molecule migrates in sodium dodecyl sulfate-polyacrylamide gel electrophoresis with an apparent molecular weight of 84,000 under reducing conditions. It can be identified as an inactive proenzyme because it has very low amidolytic activity, does not react with the fluorescent active site titrant 4-methyl-lumbelliferyl p-guanidinobenzoate, and does not incorporate radioactive [3H]diisopropylfluorophosphate. It is very susceptible to limited proteolysis which converts it to an active enzyme with trypsin-like specificity. The active enzyme, likewise a single polypeptide chain, migrates as a doublet with apparent molecular weights of 39,000 and 40,000. Its amidolytic activity with synthetic peptide substrates is at least 40-fold higher than that of the proenzyme, it reacts efficiently with 4-methylumbelliferyl p-guanidinobenzoate, and incorporates [3H]diisopropylfluorophosphate while undergoing irreversible inactivation. The enzyme appears to be a reasonably efficient plasminogen activator in zymographic gels, but not in solution. With human high molecular weight kininogen as substrate the enzyme was about 25% as efficient as human plasma kallikrein. It lacks any plasminogen-independent proteolytic activity with other protein substrates, and it hydrolyzes small peptide substrates designed for both human kallikrein and urinary urokinase, respectively. Inhibition studies with peptide chloromethyl ketones indicate enzymatic properties closer to human plasma kallikrein than to the human plasminogen activator urokinase (EC 3.4.21.31). The chicken plasma enzyme and the plasminogen activator from the conditioned media of Rous sarcoma virus-transformed chick embryo fibroblasts treated with tumor promoter are different by criteria of tryptic peptide maps, and amino acid composition and enzymatic specificity. The designations chicken plasma prekallikrein plasminogen proactivator and chicken plasma kallikrein plasminogen activator are proposed for the zymogen and enzyme forms, respectively. Using rabbit antibodies against the proenzyme we developed a solid phase immunoadsorption procedure that allowed us to isolate the protein with an overall yield of 11.4%.
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PMID:A proenzyme from chicken plasma similar to human plasma prekallikrein. 655 13

Protein Z was purified from human plasma by a four-step procedure which included barium citrate adsorption, ammonium sulfate fractionation, DEAE-Sepharose chromatography, and blue agarose chromatography with a yield of 20%. It is a 62,000 mol wt protein with an extinction coefficient of 12.0. The concentration of Protein Z in pooled, citrated plasma is 2.2 micrograms/ml and its half-life in patients starting warfarin anticoagulation therapy is estimated to be less than 2.5 d. The NH2-terminal sequence is Ala-Gly-Ser-Tyr-Leu-Leu-(Gla)-(Gla)-Leu-Phe-(Gla)-Gly-Asn-Leu. Neither Protein Z nor its cleavage products, which were obtained by treatment of Protein Z with thrombin or plasmin, incorporated [3H]diisopropyl fluorophosphate. The physiological function of Protein Z remains unknown.
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PMID:Human Protein Z. 670 12

Fibrinopeptides A and B were removed from purified human fibrinogen by bovine thrombin, whereas the snake venom protease batroxobin only split fibrinopeptide A from fibrinogen. Aggregation of the resulting desAB- and desA-fibrin monomers was evaluated by recording the turbidity of incubation mixtures. Fibrin assembly was strongly accelerated by increasing the calcium concentration from 10(-5) to 10(-3) M. Fragment D was obtained from fibrinogen by proteolytic degradation with plasmin in the presence of Ca2+. At a 4-fold molar concentration relative to fibrinogen, fragment D dramatically inhibited fibrin polymerization at up to 10(-4) M Ca2+. This anticlotting activity was, however, much less pronounced at 10(-3) M Ca2+. The thrombin clotting time, measured on human plasma, was prolonged by fragment D in a dose-dependent manner. In citrate-containing plasma, the fibrinogen clotting was significantly delayed by an equimolar concentration of fragment D. In barium sulfate-adsorbed oxalated plasma, containing 2.5 mM Ca2+, the same amount of fragment D hardly affected fibrin polymerization. We conclude that fragment D has no important anticlotting effect under physiological conditions. The synthetic peptide Gly-Pro-Arg, corresponding to the amino-terminal sequence of the fibrin alpha-chain, inhibited aggregation of both desA-fibrin and desAB-fibrin at 10(-3) M Ca2+. The inhibition of desAB-fibrin polymerization by Gly-Pro-Arg was abolished at 10(-5) M Ca2+. In addition, Gly-Pro-Arg depressed the anticlotting activity of fragment D at low calcium concentration. An analogue of the amino-terminus of fibrin beta-chain, Gly-His-Arg, strongly accelerated aggregation of desA-fibrin monomers, but only moderately enhanced polymerization of desAB-fibrin monomers at 10(-5) M Ca2+, both in the presence and in the absence of fragment D. This activating effect of Gly-His-Arg was abolished at 10(-3) M Ca2+. It is suggested that the binding of calcium, Gly-His-Arg, and possibly also Gly-Pro-Arg, induces a conformational change in fibrin monomers and thus accelerates the polymerization process.
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PMID:Inhibition of fibrin polymerization by fragment d is affected by calcium, Gly-Pro-Arg and Gly-His-Arg. 682 84

The effect of a 17-kringle form of recombinant apo(a) [r-apo(a)] on in vitro fibrin clot lysis was studied. In these assays, fibrin clots were formed in the wells of microtiter plates, and lysis of the clots was monitored by measurement of the turbidity at 405 nm. The results indicate that r-apo(a) produces a dose-dependent antifibrinolytic effect in clots formed using either purified components or barium-adsorbed plasma. This effect was found to be independent of clot structure, since lysis of clots formed using both high and low concentrations of thrombin was prolonged by r-apo(a) to the same extent. The two components of the antifibrinolytic effect of r-apo(a) were determined to be (i) attenuation of tPA-mediated plasminogen activation (the major component) and (ii) inhibition of plasmin degradation of fibrin, although r-apo(a) did not directly attenuate plasmin activity, as measured by S-2251 hydrolysis. r-Apo(a) interfered most substantially with tPA-mediated activation of Glu-plasminogen and less substantially with tPA-mediated Lys-plasminogen activation and urokinase-mediated activation of plasminogen. In summary, we have demonstrated that apo(a) is able to attenuate fibrin clot lysis in vitro, primarily as a consequence of the interference by apo(a) with tPA-mediated Glu-plasminogen activation. These studies illuminate possible mechanisms by which Lp(a) may contribute to the development of vascular disease in vivo.
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PMID:Antifibrinolytic effect of recombinant apolipoprotein(a) in vitro is primarily due to attenuation of tPA-mediated Glu-plasminogen activation. 771 Oct 34

The effect of human activated protein C (APC) on tissue plasminogen activator (tPA)-induced fibrinolysis was studied in cell free plasma and in a system of purified components. Clots were produced by adding plasma or a solution of fibrinogen and plasminogen to the wells of a microtiter plate containing small separated aliquots of Ca2+, thrombin, and tPA, plus and minus APC. Initial clotting and subsequent fibrinolysis were monitored continuously by turbidity. The lysis time of dialyzed normal human plasma (NHP) was longer than that of dialyzed barium citrate-adsorbed plasma (BAP). APC had no effect on the lysis time of BAP but shortened the lysis time of NHP to that of BAP. Two fractions were produced from material eluted from the barium citrate pellet by precipitation of selective components with polyethylene glycol 8000 (PEG). One fraction comprised materials which precipitated at 5% PEG (5% PF) and the other materials which precipitated between 5 and 40% PEG (5-40% PF). Both fractions together, but neither alone, prolonged the lysis time of BAP, an effect which could be reversed by APC. Fractionation of the 5% PF showed that the component with the required activity has properties of the procoagulant surface and can be replaced with vesicles of phosphatidylcholine/phosphatidylserine (PCPS). In addition, the 5-40% PF can be replaced with either the combination of purified coagulation Factors II, IX, and X or Factor II plus the prothrombin activator Factor Xa. When Factor Xa was used as the activator in BAP plus PSPC vesicles, a dose-dependent saturable increase in lysis time was observed with a half-maximal increase occurring at 32 pM Factor Xa. This effect was eliminated by APC. In a system of purified components comprising PCPS vesicles, Factors V and II, protein S, plasminogen and fibrinogen; the prothrombin activators Factor Xa and ecarin both induced a prolongation of the lysis time. APC prevented prolongation by Factor Xa but not by ecarin. The time courses of the generation of thrombin and plasmin during fibrinolysis of clots produced from systems of purified components in the presence and absence of APC, and with Factor Xa as the prothrombin activator, were determined by standardized activity assays using chromogenic substrates. In the absence of APC the lysis time was 145 min, and prothrombin was quantitatively converted to thrombin. In the presence of APC, however, the lysis time was reduced to 100 min with no evidence for the activation of prothrombin.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:The effect of activated protein C on fibrinolysis in cell-free plasma can be attributed specifically to attenuation of prothrombin activation. 847 6