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

After incubation of confluent monolayer cultures of human HT-1080 fibrosarcoma cells with purified native human plasminogen in plasminogen-depleted serum-containing medium, bound plasmin activity could be specifically eluted from the cells with tranexamic acid, an analogue of lysine. Dexamethasone reduced the amount of recoverable bound plasmin activity in a dose-dependent manner. Dexamethasone was also found to induce a time- and dose-dependent decrease in the ability of the cells to bind added plasmin. Untreated HT-1080 cells bound added plasmin with a high capacity (600,000 molecules bound per cell), and this decreased to an undetectable level after treatment with 100 nM dexamethasone. The kinetics of the loss of plasmin binding by the dexamethasone-treated sarcoma cells, a clear decrease after 4 h, correlated with those for the loss of cell-bound urokinase (u-PA) activity. Plasmin was not, however, bound to the active site of u-PA: an anti-catalytic monoclonal antibody to u-PA had no effect on plasmin binding. Other glucocorticoids, such as hydrocortisone and corticosterone, had a similar effect to dexamethasone on plasmin binding to HT-1080 cells. The effect of glucocorticoids on the plasmin receptor seemed to occur at least partly via a decrease in the affinity for plasmin, since the Kd for plasmin with untreated cells was 5.4 x 10(-9) M, and with cells treated with 5 nM dexamethasone, the Kd value for plasmin was 1.2 x 10(-7) M. These results show that glucocorticoids induce down-regulation of plasmin receptors on the surface of HT-1080 cells: a novel mechanism, in addition to the known effects of glucocorticoids on u-PA and PA inhibitors, by which human tumor cells may be disarmed of their pericellular proteolytic activity.
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PMID:Down-regulation of plasmin receptors on human sarcoma cells by glucocorticoids. 253 25

Chick embryo fibroblast cultures develop fibrinolytic activity after transformation by Rous sarcoma virus (RSV). This fibrinolytic activity is not present in normal cultures, and it does not appear after infection with either nontransforming strains of avian leukosis viruses or cytocidal RNA and DNA viruses. In cultures infected with a temperature sensitive mutant of RSV the onset of fibrinolysis appears after exposure to permissive temperatures and precedes by a short interval the appearance of morphological evidence of transformation. See PDF for Structure The rate of fibrinolysis in transformed cultures depends on the nature of the serum that is present in the growth medium: some sera (e.g., monkey or chicken serum) promote high enzymatic activity, while others (calf, fetal bovine) do not. Some sera contain inhibitors of the fibrinolysin. Based on the effect of a small number of known inhibitors, at least one step of the fibrinolytic process shows specificity resembling that of trypsin. The sera of sarcoma-bearing chickens contain an inhibitor of the fibrinolysin, whereas normal chicken sera do not. For general discussion, conclusions, and summary see the accompanying paper, part II, (J. Exp. Med. 137:112).
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PMID:An enzymatic function associated with transformation of fibroblasts by oncogenic viruses. I. Chick embryo fibroblast cultures transformed by avian RNA tumor viruses. 434 90

Radiomodifying action of fibrinolysin was studied during radiation treatment in experimental transplanted tumors. It has been established that the application of fibrinolysin prior to radiation increases the inhibition of the growth rate of Pliss lymphosarcoma and Sarcoma 45, as evidenced by the decrease of the tumor volume, the increase of the dose change factor and the animals survival.
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PMID:Enhancement of the radiation effect by fibrinolysin and quantitative pattern of the change of tumor radiosensitivity. 623 71

On the basis of cellular morphology, a subline of mouse sarcoma virus-infected 3T3 cells was selected which released a 48 000-dalton plasminogen activator at an approx. 40-fold higher rate than those of the parent line, and which continued to do so for several months when the cells were maintained in serum-free culture medium. Culture medium (3.5 l) containing 0.6 mg plasminogen activator per l was used to purify 620 micrograms of the enzyme 130-fold with a yield of 32% by affinity chromatography followed by anion exchange chromatography and gel filtration. Crucial for the yield was the use of a non-ionic detergent and of inhibitors of proteolysis to prevent adsorption and degradation, respectively. The purified enzyme was homogeneous as evaluated by SDS-polyacrylamide gel electrophoresis and had an isoelectric point of pH 9.2. The purified enzyme showed characteristics of a trypsin-like serine protease (labeling with [3H]diisopropylphosphorofluoridate which was prevented by p-nitrophenyl-p'-guanidinobenzoate) and converted the single chain of human plasminogen into two chains of plasmin with electrophoretic mobilities identical to those of the chains formed by non-purified enzyme and by human urokinase. In the absence of inhibitors, solutions of purified enzyme were stable for 24 h at 4 degrees C at pH 3-9.
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PMID:Purification and characterization of a plasminogen activator from mouse cells transformed by an oncogenic virus. 625 3

We have previously reported the purification of a plasminogen-activating serine protease with an approximate Mr of 48 000 from sarcoma-virus-transformed murine cells. We now report that under serum-free conditions the enzyme is released from the cells in an inactive form. After affinity chromatography with 4-aminobenzamidine-cellulose, ion-exchange chromatography and gel filtration, the proenzyme could be obtained from culture fluid as a pure, homogeneous protein as evaluated by polyacrylamide gel electrophoresis with sodium dodecylsulphate. Proenzyme was quantitatively converted to active enzyme by incubation with catalytic amounts of plasmin. Analysis by polyacrylamide gel electrophoresis with sodium dodecylsulphate under reducing and non-reducing conditions showed that the inactive form consisted of a single polypeptide chain with an Mr of approximately 48 000, while the active form consisted of two chains with Mr values of approximately 18 000 and 29 000 , held together by one or more disulphide bridges. The active-site reagent diisopropylfluorophosphate in radiolabelled form was incorporated into the 29 000-Mr chain of the active enzyme, but not into the inactive form. These findings provide conclusive evidence for the existence of an inactive proenzyme to this plasminogen activator and thus demonstrate and additional step in a cascade-like reaction leading to extracellular proteolysis. Regulatory as well as methodological implications of this findings are discussed.
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PMID:Plasminogen activator released as inactive proenzyme from murine cells transformed by sarcoma virus. 628 7

Previous studies (Miskin, R., T. G. Easton, and E. Reich, 1970, Cell. 15:1301-1312) have shown that sarcoma virus transformation and tumor promoters reduced the cell surface concentration of acetylcholine receptors (AChR) in differentiating chick embryo myogenic cultures. Both of these agents also induced high rates of plasminogen activator (PA) synthesis in myogenic cultures (Miskin, R., T. G. Easton, A. Maelicke, and E. Reich, 1978, Cell. 15:1287-1300), and the present work was performed to establish whether proteolysis might significantly affect receptor metabolism. Proteolysis in myogenic cultures was modulated by one or more of the following: stimulation of PA synthesis, direct addition of plasmin, removal of plasminogen, or addition of plasmin inhibitors. The results were: (a) When the rates of proteolysis were raised either by addition of plasmin or by stimulating PA synthesis in the presence of plasminogen, both the steady-state concentration and the half-life of surface AChR decreased, but the rate of receptor synthesis was unaffected. (b) The magnitude of these effects, and their dependence on added plasminogen, indicated that proteolysis initiated by plasminogen activation could account almost entirely for the reduction in receptor half-life produced by sarcoma virus transformation and phorbol ester. (c) The rate of receptor synthesis, which is also reduced by viral transformation and tumor promoters, was not modified by proteolysis; hence plasmin action may be responsible for a large part, but not all of the change in surface receptor under these conditions. (d) The plasmin catalysed changes in receptor parameters appear to occur in response to modified membrane metabolism resulting from proteolysis of surface components other than AChR itself.
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PMID:Acetylcholine receptor: effects of proteolysis on receptor metabolism. 646 38

Incorporation of the serine protease active site reagent diisopropyl fluorophosphate (DFP) into a plasminogen activator with an Mr of approximately 52000 released from cultured human glioblastoma cells was strongly enhanced by incubation with plasmin. This observation led to the isolation of an inactive form of the enzyme from serum-free conditioned culture fluid by affinity chromatography on a column of a Sepharose-bound monoclonal antibody raised against urokinase. An 831-fold purification was obtained with a yield of 41%. The purified molecule was homogeneous as evaluated by polyacrylamide gel electrophoresis with sodium dodecyl sulfate (NaDodSO4), having one stainable band under nonreducing as well as reducing conditions with an Mr of approximately 52000. It was unable to activate plasminogen, but catalytic amounts of plasmin converted it into active enzyme. After NaDodSO4-polyacrylamide gel electrophoresis, the active enzyme showed one band under nonreducing conditions, but after reduction, two bands with Mr values of approximately 20000 and 32000 were observed. The active enzyme incorporated [3H]DFP into the approximately Mr 32000 band, while no incorporation was observed into the inactive form. These findings show that the Mr 52000 human plasminogen activator exists in a proenzyme form consisting of a single polypeptide chain that by proteolysis between half-cystine residues is converted into the active enzyme consisting of two chains with molecular weights of approximately 20000 and 32000, the active site being on the latter chain. The results are consistent with the active form of the enzyme being identical with the higher molecular weight form of urokinase, and together with recent observations that a murine plasminogen activator is released from sarcoma virus transformed cells as an inactive proenzyme, they suggest that zymogens to plasminogen activators are of more general occurrence.
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PMID:Purification of zymogen to plasminogen activator from human glioblastoma cells by affinity chromatography with monoclonal antibody. 689 Dec 64

PC12 cells were stably transfected with expression vectors containing rat tissue plasminogen activator (tPA) under control of either a cytomegalovirus or rous sarcoma virus promoter. Cell lines were characterized using protease assays, ELISAs, immunoblots, northern blots, and Southern blots. Control PC12 cells or cells containing vectors alone released about 1 pg tPA/cell/24 h, whereas cells stably transfected with a tPA cDNA released 2-5 pg tPA/cell/24 h. A strong correlation existed between the amount of tPA released and the ability of cells to degrade extracellular matrix. Experiments with protease inhibitors and antibodies against tPA and plasminogen indicated that degradation of matrix involved tPA-generated plasmin and that the amount of matrix degraded was dependent on the amount of tPA released. Cells expressing high levels of tPA migrated on a three-dimensional matrix about twice as fast as control cells and regenerated neurites within three-dimensional gels of Matrigel to a greater extent than control cells. Antibodies that inhibited tPA and plasminogen decreased migration and neurite regeneration, indicating that tPA was involved in both events, PC12 cells overexpressing tPA should provide a useful model system for investigating neural functions of tPA including its role in migration and regeneration.
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PMID:PC12 cells overexpressing tissue plasminogen activator regenerate neurites to a greater extent and migrate faster than control cells in complex extracellular matrix. 783 49

Hepatocyte growth factor/scatter factor (HGF/SF) is a pleiotropic effector of cells expressing the Met tyrosine kinase receptor. Although HGF/SF is synthesized by mesenchymal cells and acts predominantly on epithelial cells, we have recently demonstrated that human sarcoma cell lines often inappropriately express high levels of Met and respond mitogenically to HGF/SF. In the present report we show that HGF/SF-Met signalling in the human leiomyosarcoma cell line SK-LMS-1 enhances its in vivo tumorigenicity, an effect for which the mitogenicity of this signalling pathway is likely to play a role. In addition, we found that HGF/SF-Met signalling dramatically induces the in vitro invasiveness and in vivo metastatic potential of these cells. We have studied the molecular basis by which HGFSF-Met signalling mediates the invasive phenotype. A strong correlation has previously been demonstrated between the activation of the urokinase plasminogen activator (uPA) proteolysis network and the acquisition of the invasive-metastatic phenotype, and we show here that HGF/SF-Met signalling significantly increases the protein levels of both uPA and its cellular receptor in SK-LMS-1 cells. This results in elevated levels of cell-associated uPA and enhanced plasmin-generating ability by these cells. These studies couple HGF/SF-Met signalling to the activation of proteases that mediate dissolution of the extracellular matrix-basement membrane, and important property for cellular invasion-metastasis.
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PMID:Enhanced tumorigenicity and invasion-metastasis by hepatocyte growth factor/scatter factor-met signalling in human cells concomitant with induction of the urokinase proteolysis network. 862 56

In our studies, designed to synthesize an active center-directed plasmin (PL) inhibitor, N-(4-aminomethylbenzoyl)-4-(3-picolyloxy)-L-phenylalanine n-hexylamide dihydrochloride (PASI-535) was found. We characterized PASI-535 and analyzed the actions of PL, comparing with those of PASI-535 and tranexamic acid (t-AMCHA). (1) PASI-535 strongly inhibited not only fibrinolysis (IC50: 2.9 x 10(-6) M) but also amidolysis (Ki value: 2.9 x 10(-6) M) and fibrinogenolysis (IC50: 4.5 x 10(-6) M) induced by PL. While t-AMCHA which strongly inhibited fibrinolysis (IC50: 6.0 x 10(-5) M), rarely inhibited amidolysis (Ki value: 4.0 x 10(-2) M) and fibrinogenolysis (IC50: 1.0 x 10(-2) M). (2) PL is able to liberate kinins by degrading kininogen. This kinin-generation by PL was inhibited by 2 x 10(-5) M PASI-535. However, it was little inhibited even by 1 x 10(-3) M t-AMCHA. (3) The inhibitory effect of PASI-535 and t-AMCHA on tumor growth was studied. In sarcoma-180 bearing mice, ascites retention and the increase of tumor cells were markedly suppressed by subcutaneous injection of PASI-535, either 30 mg/kg/day or 50 mg/kg/day, for 5 days, and the inhibitory effect was dose-dependent. Although t-AMCHA also reduced both ascites retention and the increase of tumor cells, it needed approximately 40 times (2 g/kg/day) the amount of PASI-535 to exert these effects. PASI-535 may be a useful tool in analyzing the multiplicity of PL actions. Moreover, PASI-535 can be used as an antifibrinolytic drug which has a mechanism of function different from that of t-AMCHA.
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PMID:Use of an active center-directed plasmin inhibitor elucidates the multiplicity of plasmin actions. 873 12


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