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:3.4.21.68 (tissue plasminogen activator)
11,311 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Unlike most proteases, tissue-type plasminogen activator (t-PA) is not synthesized as an inactive precursor or zymogen. Instead, the single-chain "proenzyme" form of t-PA possesses very significant catalytic activity. Recent investigations of the molecular basis of the unusually high enzymatic activity of single-chain t-PA have focused attention upon Asp-194, a residue that is invariant among chymotrypsin-like enzymes. The critical role of this residue in securing the active conformation of mature chymotrypsin-like enzymes has been discussed extensively. Subsequent work, however, has indicated that this conserved residue can also form interactions that dramatically influence the catalytic activity of serine protease zymogens. While Asp-194 forms interactions that suppress the activity of the zymogen chymotrypsinogen, it may, by contrast, directly promote the catalytically active conformation of single-chain t-PA. To test the hypothesis that Asp-194 promotes the activity of both single- and two-chain t-PA and therefore plays opposing roles in single-chain t-PA and chymotrypsinogen, and also to examine whether this invariant residue plays an essential role in the stimulation of t-PA by fibrin, we used site-directed mutagenesis to construct the following variants of t-PA: t-PA/D194E, t-PA/D194N, t-PA/R15E,D194E, and t-PA/R15E,D194N. In the absence of fibrin, the activity of enzymes carrying a mutation at position 194 was reduced by factors of 1000-2000 compared to wild type t-PA. Similar reductions of activity were observed for both single- and two-chain variants, suggesting an important role for Asp-194 in both forms of the enzyme. The mutated enzymes, however, displayed a dramatically enhanced response to fibrin monomers. While the activity of wild type t-PA was stimulated by fibrin monomers by a factor of 960, the corresponding stimulation factor for the mutated enzymes varied from 498,000-1,050,000.
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PMID:Variants of tissue-type plasminogen activator with substantially enhanced response and selectivity toward fibrin co-factors. 755 5

Endothelial cell differentiation into capillary structures is a complex process that requires the concerted effects of several extracellular matrix proteases, including plasminogen activators. Here, the role of tissue-type plasminogen activator (tPA) and urokinase-type plasminogen activator (uPA) was evaluated in an in vitro model of endothelial morphogenesis involving organization of human umbilical vein endothelial cells into tubular structures when they are cultured on the basement membrane preparation, Matrigel. Both uPA and tPA were detected in HUVEC cultures on Matrigel, and inhibitors of plasminogen activators or of serine proteases decreased the extent of the tube network formed by the cells. The decrease resulting from serine protease inhibitors was additive to that from matrix metalloproteinase inhibitors which have previously been shown to decrease tube formation in this model, suggesting that the two classes of proteases modulate tube formation by distinct mechanisms. Plasminogen activator inhibitor (PAI)-1 decreased tube formation by 50% when added up to 4.5 h after the initiation of an 18 h assay and caused 25% inhibition when added 9.5 h after culture initiation, indicating that the effects of plasminogen activators are not limited to an early event in the differentiation process. Steady-state expression of mRNA for uPA increased during the first several hours of culture on Matrigel, further supporting a role for PA activity throughout the process of tube formation. These findings suggested that PAs may affect multiple events during tube-forming activity. A fucosylated peptide comprising the amino-terminal domain of uPA that binds to the uPA receptor (uPAR) but lacking proteolytic activity enhanced tube formation. In contrast, a defucosylated form of the same peptide had no effect. Since fucosylation of this fragment has been shown to be essential in other models of cell stimulation by uPA-uPAR interaction, these data support the hypothesis that uPA enhances endothelial morphogenesis both through proteolytic activity and via uPAR occupancy. Plasminogen activators could facilitate angiogenesis in vivo.
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PMID:Plasminogen activators augment endothelial cell organization in vitro by two distinct pathways. 755 92

Neuronal degeneration in the hippocampus, a region of the brain important for acquisition of memory in humans, occurs in various pathological conditions, including Alzheimer's disease, brain ischaemia and epilepsy. When neuronal activity is stimulated in the adult rat and mouse hippocampus, tissue plasminogen activator (tPA), a serine protease that converts inactive plasminogen to the active protease plasmin, is transcriptionally induced. The activity of tPA in neural tissue is correlated with neurite outgrowth, regeneration and migration, suggesting that it might be involved in neuronal plasticity. Here we show that tPA is produced primarily by microglia in the hippocampus. Using excitotoxins to induce neuronal cell loss, we demonstrate that tPA-deficient mice are resistant to neuronal degeneration. These mice are also less susceptible to pharmacologically induced seizures than wild-type mice. These findings identify a role for tPA in neuronal degeneration and seizure.
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PMID:Excitotoxin-induced neuronal degeneration and seizure are mediated by tissue plasminogen activator. 756 88

The plasminogen activator (PA)/plasmin pathway has been implicated in a variety of physiologic and pathologic processes that require tissue remodeling and cell motility. The pathway is highly regulated and results in the generation of the broad spectrum serine protease, plasmin, from the zymogen plasminogen. Urokinase-type plasminogen activator (uPA) and tissue-type plasminogen activator (tPA) are produced by osteoblasts, as are the specific inhibitor plasminogen activator inhibitor-1 (PAI-1) and a cellular receptor for uPA. Little is known about regulation of the receptor, but the other 3 components of the pathway are regulated differentially by various osteotropic hormones and local factors. Several roles have been proposed for this pathway in bone. These involve proteolytic activation of procollagenase and latent growth factors, such as transforming growth factor beta (TGF beta) and insulin-like growth factor (IGF), as well as cell motility as a result of pericellular proteolysis. The roles of this pathway in bone remodeling may not be confined to proteolytic events, because uPA has been reported to act as a mitogen on osteoblast-like cells. This effect is independent of proteolytic activity but requires the growth factor domain of uPA to bind to uPA cellular receptors. If the plasminogen activator/plasmin pathway encompasses these roles, it would serve to couple formation and resorption of bone in a highly regulated manner.
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PMID:The plasminogen activator inhibitor system in bone cell function. 764 98

We investigated the effect of gonadotropins on protease that were suggested to be implicated in the invasive activity of the trophoblast. hCG levels ranging from 10 x 10(3) to 333 x 10(3) IU/L produced a dose-dependent inhibition of the in vitro globinolytic activity of the purified proteases trypsin, chymotrypsin, and urokinase, but failed to inhibit plasmin, collagenase, elastase, and tissue-type plasminogen activator. Likewise, FSH inhibited purified trypsin and urokinase, but not plasmin or tissue-type plasminogen activator. Culture medium conditioned with human trophoblast displayed serine protease and urokinase-like activities; exposure of the cultured trophoblast to exogenous hCG markedly suppressed serine protease and urokinase activities in the conditioned medium. A short treatment of the conditioned medium with trypsin abolished the hCG-mediated inhibition of urokinase activity. The present findings offer an explanation for earlier observations that hCG reduced collagenase activity in trophoblasts without affecting the level of collagenase-specific mRNA. The present results are also consistent with the concept that hCG, by its direct ability to inhibit certain serine proteases and urokinase in trophoblast, suppresses a protease-mediated conversion of procollagenase to active collagenase. The ability of hCG to prevent initiation of the collagenolytic cascade suggests that gonadotropins may regulate the transient invasive activity of the trophoblast.
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PMID:Gonadotropin-mediated inhibition of proteolytic enzymes produced by human trophoblast in culture. 768 89

Urokinase plasminogen activator (uPA) is a serine protease involved in cancer invasion and metastasis. uPA mediates its action while attached to a membrane-bound receptor (uPAR). In this investigation we show that uPAR levels correlate with uPA levels in human breast cancers. uPAR levels, however, do not correlate with other components of the plasminogen activator system such as tissue-type plasminogen activator (t-PA), PAI-I or PAI-2. In addition, uPAR levels showed no correlation with tumor size, axillary-node status or estrogen-receptor status. On the basis of an optimum cut-off point, patients with breast cancers containing high levels of uPAR had a worse prognosis than patients with low levels of the receptor. However, as a prognostic marker in breast cancer, uPAR was not as strong as uPA. Our results are consistent with data from model systems suggesting that both uPA and uPAR are necessary for metastasis.
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PMID:Urokinase plasminogen activator and urokinase plasminogen activator receptor in breast cancer. 776 29

We have developed a model whereby the middle cerebral artery in an experimental animal can be occluded by a photochemical reaction between rose bengal and green light. This causes endothelial injury followed by platelet adhesion, aggregation and formation of a platelet-rich thrombus at the site of the photochemical reaction. SUN9216, a modified tissue-type plasminogen activator, is a new thrombolytic agent which consists of the fibrin kringle 1 domain of plasminogen and the two kringles, the serine protease domains of the native tissue-type plasminogen activator. The mannose glycosylation site on the kringle 1 of tissue-type plasminogen activator is modified to yield a compound with a longer half-life in the blood than native tissue-type plasminogen activator. We evaluated the thrombolytic effects of recombinant tissue-type plasminogen activator and SUN9216 in the thrombotically occluded rat middle cerebral artery. SUN9216 was administered by continuous infusion or as a single bolus injection 30 min after the middle cerebral artery had been occluded by a thrombus. Both SUN9216 and recombinant tissue-type plasminogen activator caused reopening of the middle cerebral artery by thrombolysis. The efficacy of SUN9216 was higher than that of recombinant tissue-type plasminogen activator. Further, the area of ischaemic cerebral damage caused by the middle cerebral artery occlusion was significantly (P < 0.05) reduced by SUN9216, but in this respect, recombinant tissue-type plasminogen activator was ineffective.
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PMID:Thrombolytic efficacy of a modified tissue-type plasminogen activator, SUN9216, in the rat middle cerebral artery thrombosis model. 781 75

We described in a foregoing report findings on serpin, a serine protease inhibitor, newly identified in horseshoe crab (Tachypleus tridentatus) hemocytes and we name it limulus intracellular coagulation inhibitor, LICI (Miura, Y., Kawabata, S., and Iwanaga, S. (1994) J. Biol. Chem. 269, 542-547). This serpin specifically inhibits limulus lipopolysaccharide-sensitive serine protease, factor C. In ongoing studies on limulus serpin, we have found another inhibitor, LICI type-2 (LICI-2), which inhibits not only factor C (k1 = 7.1 x 10(4) M-1 S-1) but also limulus clotting enzyme (k1 = 4.3 x 10(5) M-1 S-1). LICI-2 inhibits mammalian serine proteases, including alpha-thrombin, salivary kallikrein, plasmin, and tissue plasminogen activator. The inactivation of plasmin is the most rapid (k1 = 1.2 x 10(6) M-1 S-1). The purified LICI-2 is a single chain glycoprotein with an apparent M(r) = 42,000. A cDNA for LICI-2 was isolated and the open reading frame coded for a mature protein of 386 amino acids, of which 160 residues were confirmed by peptide sequencing. Although LICI-2 shows significant sequence similarity to the previous limulus serpin, LICI-1 (42% identity), LICI-2 contains a unique putative reactive site, -Lys-Ser-, distinct from that of LICI-1 (-Arg-Ser-). Northern blotting revealed expression of LICI-2 mRNA only in hemocytes, and not in heart, brain, stomach, intestine, coxal gland, and skeletal muscle. The immunoblot of large and small granule components with antiserum against purified LICI-2 suggests that LICI-2 is stored specifically in large granules, as in the case of LICI-1, and is released in response to external stimuli. We propose that the LICIs be classified into a new subfamily of intracellular serpins, regulated secretory serpins.
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PMID:A limulus intracellular coagulation inhibitor type 2. Purification, characterization, cDNA cloning, and tissue localization. 782 80

Plasminogen activator inhibitor 1 (PAI-1), the primary physiological inhibitor of t-PA, is an unusual member of the serpin family of serine protease inhibitors, in that it spontaneously converts to a latent form. Latent PAI-1 has been reported to share characteristics with the cleaved form of other serpins. Here we examine the conformation of four forms of PAI-1, active and latent wild-type, together with a noninhibitory, substrate mutant that is cleavable at P1-P1', and its cleaved product. The circular dichroism spectra of active and latent PAI-1 showed differences consistent with decreased alpha-helix from 26% to 22% and increased beta-sheet from 23% to 34% as active-->latent. Active and substrate PAI-1 were less thermostable than latent PAI-1, which was 50% denatured at 70 degrees C. In contrast, cleaved PAI-1 was very stable, with little loss of structure at 100 degrees C. Cleaved PAI-1 was much more resistant to guanidinium chloride (Gdn-HCl), 50% unfolding requiring 4.5 M Gdn-HCl, while active, latent, and substrate forms of PAI-1 were 50% unfolded in 2-2.5 M Gdn-HCl. The differences in fluorescence emission maxima, latent 339 nm, active 336 nm, substrate 343 nm, and cleaved 333 nm, underline the contrast between latent and cleaved PAI-1. The conformational changes occurring on cleavage are clearly more profound than those seen on transition from active to latent PAI-1.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Conformational studies on plasminogen activator inhibitor (PAI-1) in active, latent, substrate, and cleaved forms. 782 21

Human hepatocyte growth factor (hHGF) has considerable sequence homology with plasminogen and both proteins can be activated by plasminogen activators. The aim of this study was to investigate the relationship between plasma hHGF and fibrinolysis in patients with fulminant hepatic failure (FHF), in whom proteases of coagulation are known to be activated and hHGF levels have been shown to be raised as a consequence of hepatic regeneration. Serum hHGF measured by ELISA was increased in FHF (median 6.67 ng/ml, range 1.2-62 ng/ml), but the values did not correlate with the decreased plasminogen level (median 9%., range 0.7-35.5%) or the level of t-PA which was normal. There was a significant correlation between serum hHGF and increased plasma D-dimer (median 2,163 microgram/l, range 39-7 311 microgram/l), produced by the action of plasmin on fibrin and increased plasma thrombin-antithrombin III complexes (TAT, median 31.7 microgram/l, range 3.7-105 microgram/l). These relationship could be indicative of an involvement of blood coagulation, possibly a specific serine protease, in hHGF activity. After liver transplantation, plasma hHGF was rapidly cleared to almost normal levels, whereas D-dimer and TAT continued to be at elevated levels.
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PMID:Hepatocyte growth factor and plasminogen activation in fulminant hepatic failure. 784 6


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